Hidden hotspots of amphibian biodiversity in China.

Identifying and protecting hotspots of endemism and species richness is crucial for mitigating the global biodiversity crisis. However, our understanding of spatial diversity patterns is far from complete, which severely limits our ability to conserve biodiversity hotspots. Here, we report a comprehensive analysis of amphibian species diversity in China, one of the most species-rich countries on Earth. Our study combines 20 y of field surveys with new molecular analyses of 521 described species and also identifies 100 potential cryptic species. We identify 10 hotspots of amphibian diversity in China, each with exceptional species richness and endemism and with exceptional phylogenetic diversity and phylogenetic endemism (based on a new time-calibrated, species-level phylogeny for Chinese amphibians). These 10 hotspots encompass 59.6% of China's described amphibian species, 49.0% of cryptic species, and 55.6% of species endemic to China. Only four of these 10 hotspots correspond to previously recognized biodiversity hotspots. The six new hotspots include the Nanling Mountains and other mountain ranges in South China. Among the 186 species in the six new hotspots, only 9.7% are well covered by protected areas and most (88.2%) are exposed to high human impacts. Five of the six new hotspots are under very high human pressure and are in urgent need of protection. We also find that patterns of richness in cryptic species are significantly related to those in described species but are not identical.

The highest-elevation frog provides insights into mechanisms and evolution of defenses against high UV radiation.

Defense against ultraviolet (UV) radiation exposure is essential for survival, especially in high-elevation species. Although some specific genes involved in UV response have been reported, the full view of UV defense mechanisms remains largely unexplored. Herein, we used integrated approaches to analyze UV responses in the highest-elevation frog, Nanorana parkeri. We show less damage and more efficient antioxidant activity in skin of this frog than those of its lower-elevation relatives after UV exposure. We also reveal genes related to UV defense and a corresponding temporal expression pattern in N. parkeri. Genomic and metabolomic analysis along with large-scale transcriptomic profiling revealed a time-dependent coordinated defense mechanism in N. parkeri. We also identified several microRNAs that play important regulatory roles, especially in decreasing the expression levels of cell cycle genes. Moreover, multiple defense genes (i.e., TYR for melanogenesis) exhibit positive selection with function-enhancing substitutions. Thus, both expression shifts and gene mutations contribute to UV adaptation in N. parkeri. Our work demonstrates a genetic framework for evolution of UV defense in a natural environment.

A comprehensive molecular phylogeny of the genus Sylvirana (Anura: Ranidae) highlights unrecognized diversity, revised classification and historical biogeography.

The genus Sylvirana includes 12 species widely distributed in South China and Southeast Asia. The phylogenetic relationships and species diversity for Sylvirana and allied genera remain unresolved and controversial due to insufficient data and incomplete taxon sampling. Using a combined dataset of mitochondrial genes (16S and COI) and 101 nuclear genes obtained through the amplicon sequence capture approach, we generated the most comprehensive phylogenetic analysis for the genus Sylvirana to date, inferring diversity, phylogenetic relationships, and historical biogeography with unprecedented levels of taxon and geographic sampling. Our results conservatively reveal six undescribed species, mostly distributed in peninsular Indochina. Phylogenetic analyses strongly support the non-monophyly of Sylvirana with respect to Pterorana. Additionally, phylogenetic results place Sylvirana guentheri and Pelophylax lateralis into genus Humerana, supporting the inclusion of Hylarana latouchii, Papurana milleti, and Hylarana attigua within Pterorana + Sylvirana. The long-disputed species of Hylarana bannanica (previously Sylvirana) cluster with genus Papurana. Because the results of multiple non-monophyletic genera create taxonomic confusion, we suggest relegating all genera to subgenus rank of Hylarana. Sylvirana is a junior synonym of the Pterorana. Biogeographically, we trace the origin of Pterorana to Southeast Asia during the early Miocene, with subsequent dispersal thereafter. Our study shows that climatic changes may have profoundly influenced the diversification of Pterorana during the Miocene.

Species Persistence with Hybridization in Toad-Headed Lizards Driven by Divergent Selection and Low Recombination.

Speciation plays a central role in evolutionary studies, and particularly how reproductive isolation (RI) evolves. The origins and persistence of RI are distinct processes that require separate evaluations. Treating them separately clarifies the drivers of speciation and then it is possible to link the processes to understand large-scale patterns of diversity. Recent genomic studies have focused predominantly on how species or RI originate. However, we know little about how species persist in face of gene flow. Here, we evaluate a contact zone of two closely related toad-headed lizards (Phrynocephalus) using a chromosome-level genome assembly and population genomics. To some extent, recent asymmetric introgression from Phrynocephalus putjatai to P. vlangalii reduces their genomic differences. However, their highly divergent regions (HDRs) have heterogeneous distributions across the genomes. Functional gene annotation indicates that many genes within HDRs are involved in reproduction and RI. Compared with allopatric populations, contact areas exhibit recent divergent selection on the HDRs and a lower population recombination rate. Taken together, this implies that divergent selection and low genetic recombination help maintain RI. This study provides insights into the genomic mechanisms that drive RI and two species persistence in the face of gene flow during the late stage of speciation.

Impacts of climate change on herpetofauna diversity in the Qinghai-Tibetan Plateau.

Although numerous studies on the impacts of climate change on biodiversity have been published, only a handful are focused on the intraspecific level or consider population-level models (separate models per population). We endeavored to fill this knowledge gap relative to the Qinghai-Tibetan plateau (QTP) by combining species distribution modeling (SDMs) with population genetics (i.e., population-level models) and phylogenetic methods (i.e., phylogenetic tree reconstruction and phylogenetic diversity analyses). We applied our models to 11 endemic and widely distributed herpetofauna species inhabiting high elevations in the QTP. We aimed to determine the influence of environmental heterogeneity on species' responses to climate change, the magnitude of climate-change impacts on intraspecific diversity, and the relationship between species range loss and intraspecific diversity losses under 2 shared socioeconomic pathways (SSP245 and SSP585) and 3 future periods (2050s, 2070s, and 2090s). The effects of global climatic change were more pronounced at the intraspecific level (22% of haplotypes lost and 36% of populations lost) than the morphospecies level in the SSP585 climate change scenario. Maintenance of genetic diversity was in general determined by a combination of factors including range changes, species genetic structure, and the part of the range predicted to be lost. This is owing to the fact that the loss and survival of populations were observed in species irrespective of the predicted range changes (contraction or expansion). In the southeast (mountainous regions), climate change had less of an effect on range size (>100% in 3 species) than in central and northern QTP plateau regions (range size <100% in all species). This may be attributed to environmental heterogeneity, which provided pockets of suitable climate in the southeast, whereas ecosystems in the north and central regions were homogeneous. Generally, our results imply that mountainous regions with high environmental heterogeneity and high genetic diversity may buffer the adverse impacts of climate change on species distribution and intraspecific diversity. Therefore, genetic structure and characteristics of the ecosystem may be crucial for conservation under climate change.

Impactos del cambio climático sobre la diversidad de herpetofauna en la meseta Qinghai-Tíbet Región Aunque se han publicado numerosos estudios sobre los impactos del cambio climática en la biodiversidad, son muy pocos los que se enfocan en el nivel intraespecífico o que consideran modelos a nivel poblacional (modelos separados por población). Intentamos cerrar este vacío de conocimiento en relación con la meseta Qinghai-Tíbet (MQT) con la combinación entre modelos de distribución de especies (MDE) y genética poblacional (modelos a nivel poblacional) y métodos filogenéticos (reconstrucción de árboles filogenéticos y análisis de diversidad filogenética). Aplicamos nuestros modelos a once especies endémicas de herpetofauna con distribución amplia en las elevaciones más altas de la MQT. Nos planteamos determinar la influencia de la heterogeneidad de las especies sobre la respuesta de las especies al cambio climático, la magnitud de los impactos del cambio climático sobre la diversidad intraespecífica y la relación entre la pérdida de distribución de la especie y las pérdidas de diversidad intraespecífica bajo dos vías socioeconómicas (SSP245 y SSP585) y tres periodos del futuro (2050s, 2070s y 2090s). Los efectos del cambio climático global fueron más pronunciados a nivel intraespecífico (22% de pérdida en los haplotipos y 36% en las poblaciones) que al nivel morfoespecie en el escenario de cambio climático SSP585. El mantenimiento de la diversidad genética casi siempre estuvo determinado por una combinación de factores que incluyen cambios en la distribución, estructura genética de las especies y la parte de la distribución que se pronosticó se perdería. Esto se debe a que observamos la pérdida y supervivencia de las poblaciones sin importar los cambios pronosticados en la distribución (contracción o expansión). En las regiones montañosas del sureste, el cambio climático tuvo un efecto menor sobre la distribución (>100% en tres especies) comparado con las regiones de la meseta central y del norte de la MQT (distribución <100% en todas las especies). Esto puede atribuirse a la heterogeneidad ambiental, la cual proporciona recovecos de clima adecuado en el sureste, mientras que los ecosistemas en las regiones central y norte fueron homogéneos. De manera general, nuestros resultados implican que las regiones montañosas con una elevada heterogeneidad ambiental y una gran diversidad genética podrían reducir los impactos adversos del cambio climático sobre la distribución de las especies y la diversidad intraespecífica. Por lo tanto, la estructura genética y las características del ecosistema pueden ser cruciales para conservar bajo el cambio climático.

Genomic and transcriptomic investigations of the evolutionary transition from oviparity to viviparity.

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.

Multi-locus genetic analyses of Quasipaa from throughout its distribution.

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.

Species groups distributed across elevational gradients reveal convergent and continuous genetic adaptation to high elevations.

Although many cases of genetic adaptations to high elevations have been reported, the processes driving these modifications and the pace of their evolution remain unclear. Many high-elevation adaptations (HEAs) are thought to have arisen in situ as populations rose with growing mountains. In contrast, most high-elevation lineages of the Qinghai-Tibetan Plateau appear to have colonized from low-elevation areas. These lineages provide an opportunity for studying recent HEAs and comparing them with ancestral low-elevation alternatives. Herein, we compare four frogs (three species of Nanorana and a close lowland relative) and four lizards (Phrynocephalus) that inhabit a range of elevations on or along the slopes of the Qinghai-Tibetan Plateau. The sequential cladogenesis of these species across an elevational gradient allows us to examine the gradual accumulation of HEA at increasing elevations. Many adaptations to high elevations appear to arise gradually and evolve continuously with increasing elevational distributions. Numerous related functions, especially DNA repair and energy metabolism pathways, exhibit rapid change and continuous positive selection with increasing elevations. Although the two studied genera are distantly related, they exhibit numerous convergent evolutionary changes, especially at the functional level. This functional convergence appears to be more extensive than convergence at the individual gene level, although we found 32 homologous genes undergoing positive selection for change in both high-elevation groups. We argue that species groups distributed along a broad elevational gradient provide a more powerful system for testing adaptations to high-elevation environments compared with studies that compare only pairs of high-elevation versus low-elevation species.

Selection and environmental adaptation along a path to speciation in the Tibetan frog Nanorana parkeri.

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.

An integrative phylogenomic approach illuminates the evolutionary history of Old World tree frogs (Anura: Rhacophoridae).

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.

Resistin Regulates Fatty Acid Β Oxidation by Suppressing Expression of Peroxisome Proliferator Activator Receptor Gamma-Coactivator 1α (PGC-1α).

BACKGROUND/AIMS: Abnormal fatty acid ß oxidation has been associated with obesity and type 2 diabetes. Resistin is an adipokine that has been considered as a potential factor in obesity-mediated insulin resistance and type 2 diabetes. However, the effect of resistin on fatty acid ß oxidation needs to be elucidated. METHODS: We detected the effects of resistin on the expression of fatty acid oxidation (FAO) transcriptional regulatory genes, the fatty acid transport gene, and mitochondrial ß-oxidation genes using real-time PCR. The rate of FAO was measured using 14C-palmitate. Immunofluorescence assay and western blot analysis were used to explore the underlying molecular mechanisms. RESULTS: Resistin leads to a reduction in expression of the FAO transcriptional regulatory genes ERRα and NOR1, the fatty acid transport gene CD36, and the mitochondrial ß-oxidation genes CPT1, MCAD, and ACO. Importantly, treatment with resistin led to a reduction in the rate of cellular fatty acid oxidation. In addition, treatment with resistin reduced phosphorylation of acetyl CoA carboxylase (ACC) (inhibitory). Mechanistically, resistin inhibited the activation of CREB, resulting in suppression of PGC-1α. Importantly, overexpressing PGC-1α can rescue the inhibitory effects of resistin on fatty acid ß oxidation. CONCLUSIONS: Activating the transcriptional activity of CREB using small molecular chemicals is a potential pharmacological strategy for preventing the inhibitory effects of resistin on fatty acid ß oxidation.

Large-scale phylogenetic analyses provide insights into unrecognized diversity and historical biogeography of Asian leaf-litter frogs, genus Leptolalax (Anura: Megophryidae).

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.

DNA barcoding of Vietnamese bent-toed geckos (Squamata: Gekkonidae: Cyrtodactylus) and the description of a new species.

Species of bent-toed gecko (Cyrtodactylus) in Vietnam have been described at a rate of nearly four species per year since 2007 mostly based on morphological data. A tool that guides species delimitation will accelerate the rate of documentation, and at a time when the recognition of species greatly benefits conservation. We use DNA barcoding using COI (550 bp) to re-examine the levels of genetic divergence and taxonomic status of 21 described species of Vietnamese bent-toed geckos. Tree-based analyses resolve all sampled species and identify potential undescribed taxa. Kimura 2-parameter genetic distances between the described species average 21.0±4.2% and range from 4.3% to 28.7%. Further, our analyses discover two potentially new species from Vietnam, two from Laos and one from China. Herein we describe the new species Cyrtodactylus puhuensis sp. nov. from Vietnam on the basis of both genetics and morphology. Genetically, it differs from the remaining species by an average K2P distance of 24.0±1.8%. Morphologically, the new species is diagnosed by its medium-size (snout-vent length 79.24 mm and tail length 82.59 mm, for the single known individual), in having a series of moderately enlarged transverse subcaudals and a series of moderately enlarged femoral scales that extend from precloacal scales, in possessing femoral scales without pores, with males having five precloacal pores, and in exhibiting 8 supralabials, 10 infralabials, 23 narrow subdigital lamellae on its fourth toe, and 36 transverse ventrals.

Phylogeny of the Cyrtodactylus irregularis species complex (Squamata: Gekkonidae) from Vietnam with the description of two new species.

The number of described species of bent-toed geckos of the Cyrtodactylus irregularis species complex in Vietnam has increased from one to eight in the last six years. We combined morphological and molecular analyses to explore phylogenetic relationships among all described species in the group. The phylogeny required the description of two new species, Cyrtodactylus phuocbinhensis sp. nov. and Cyrtodactylus taynguyenensis sp. nov. Further, the tree resolved two additional undescribed clades that may also be new species. The species C. bugiamapensis and C. ziegleri were found to require redefinition. Cyrtodactylus phuocbinhensis sp. nov. is characterized by a series of enlarged femoral scales separated from preanal scales while Cyrtodactylus taynguyenensis sp. nov. does not possess enlarged femoral scales. Both new species are distinguished from other congeners by a combination of the following characters: small subcaudal scales, not transversely enlarged; presence (C. phuocbinhensis sp. nov.) or absence (C. taynguyenensis sp. nov.) of enlarged femoral scales; number of preanal pores; and dorsal pattern. Genetic distances between described species and new species were 16.5% and 2.0% in COI and RPL35, respectively, for C. phuocbinhensis sp. nov., and these distances were 18.8% and 2.2% for C. taynguyenensis sp. nov., respectively.

First national record of Microhylahmongorum Hoang, Nguyen, Phan, Pham, Ninh, Wang, Jiang, Ziegler and Nguyen, 2022 (Anura, Microhylidae, Microhyla) in China.

Background: To date, 10 species of the genus Microhyla have been recorded in China, of which six were distributed in Yunnan Province. Microhylahmongorum Hoang, Nguyen, Phan, Pham, Ninh, Wang, Jiang, Ziegler, and Nguyen, 2022 was also speculated to be distributed in Xishuangbana, Yunnan Province, China. However, there is no evidence of documentation of M.hmongorum. New information: We report the first country record of Microhylahmongorum, based on specimens collected from Yunnan border region. Morphologically, the specimen was consistent with the original descriptions of M.hmongorum. Phylogenetically, the sequences of the specimens from China clustered with the sequence of type specimens of M.hmongorum from Vietnam, with uncorrected pairwise distances of 0.9% at the 16S gene fragment analysed. Therefore, we report M.hmongorum as a new record species in China.

DNA barcoding of Chinese snakes reveals hidden diversity and conservation needs.

DNA barcoding has greatly facilitated studies of taxonomy, biodiversity, biological conservation, and ecology. Here, we establish a reliable DNA barcoding library for Chinese snakes, unveiling hidden diversity with implications for taxonomy, and provide a standardized tool for conservation management. Our comprehensive study includes 1638 cytochrome c oxidase subunit I (COI) sequences from Chinese snakes that correspond to 17 families, 65 genera, 228 named species (80.6% of named species) and 36 candidate species. A barcode gap analysis reveals gaps, where all nearest neighbour distances exceed maximum intraspecific distances, in 217 named species and all candidate species. Three species-delimitation methods (ABGD, sGMYC, and sPTP) recover 320 operational taxonomic units (OTUs), of which 192 OTUs correspond to named and candidate species. Twenty-eight other named species share OTUs, such as Azemiops feae and A. kharini, Gloydius halys, G. shedaoensis, and G. intermedius, and Bungarus multicinctus and B. candidus, representing inconsistencies most probably caused by imperfect taxonomy, recent and rapid speciation, weak taxonomic signal, introgressive hybridization, and/or inadequate phylogenetic signal. In contrast, 43 species and candidate species assign to two or more OTUs due to having large intraspecific distances. If most OTUs detected in this study reflect valid species, including the 36 candidate species, then 30% more species would exist than are currently recognized. Several OTU divergences associate with known biogeographic barriers, such as the Taiwan Strait. In addition to facilitating future studies, this reliable and relatively comprehensive reference database will play an important role in the future monitoring, conservation, and management of Chinese snakes.

Large-scale mtDNA screening reveals a surprising matrilineal complexity in east Asia and its implications to the peopling of the region.

In order to achieve a thorough coverage of the basal lineages in the Chinese matrilineal pool, we have sequenced the mitochondrial DNA (mtDNA) control region and partial coding region segments of 6,093 mtDNAs sampled from 84 populations across China. By comparing with the available complete mtDNA sequences, 194 of those mtDNAs could not be firmly assigned into the available haplogroups. Completely sequencing 51 representatives selected from these unclassified mtDNAs identified a number of novel lineages, including five novel basal haplogroups that directly emanate from the Eurasian founder nodes (M and N). No matrilineal contribution from the archaic hominid was observed. Subsequent analyses suggested that these newly identified basal lineages likely represent the genetic relics of modern humans initially peopling East Asia instead of being the results of gene flow from the neighboring regions. The observation that most of the newly recognized mtDNA lineages have already differentiated and show the highest genetic diversity in southern China provided additional evidence in support of the Southern Route peopling hypothesis of East Asians. Specifically, the enrichment of most of the basal lineages in southern China and their rather ancient ages in Late Pleistocene further suggested that this region was likely the genetic reservoir of modern humans after they entered East Asia.

Lactase persistence may have an independent origin in Tibetan populations from Tibet, China.

Milk consumption is prevalent in daily diets of Tibetans. To digest the milk sugar lactose, lactase persistence (LP) should be required. However, little is known about the genetic basis of LP in Tibetans. We screened 495 Tibetan individuals for five previously reported single-nucleotide polymorphisms (SNPs): -13907C/G (rs41525747), -13910C/T (rs4988235), -13915T/G (rs41380347), -14010G/C and -22018G/A (rs182549), which are associated with the LP in populations from a vast region surrounding Tibet. The five SNPs were nearly absent in Tibetan populations, suggesting LP likely to have an independent origin in Tibetans rather than to be introduced via gene flow from neighboring populations. We identified three novel SNPs (-13838G/A, -13906T/A and -13908C/T) in Tibetans. In particular, -13838G/A might be functional as it is located in the binding motif for HNF4α that acts as a transcription factor for intestinal gene expression. To investigate the potential association of this variant with LP, further detailed studies are required in the future.

Speciation in the Rana chensinensis species complex and its relationship to the uplift of the Qinghai-Tibetan Plateau.

Speciation remains a fundamental issue in biology. Herein, we report an investigation into speciation in the Rana chensinensis species complex using DNA sequence data from one mitochondrial and five nuclear genes. A phylogenetic analysis of the data revealed four major clades in the complex, and each of them was found to likely represent a species, including one cryptic species. Ecological niche models were generated from 19 climatic variables for three of the four major clades, which were represented by widespread sampling, including R. chensinensis, Rana kukunoris and the potential cryptic species. Each clade is associated with a unique ecological unit, and this indicates that ecological divergence probably drove speciation. Ecological divergence is likely related to the late Cenozoic orogenesis of the Qinghai-Tibetan Plateau. In addition, gene flow between species was detected but only in peripheral portions of the ranges of the four major clades, thus likely had little influence on the speciation processes. Discordances between mitochondrial and nuclear genes were also found; the nominal species, R. chensinensis, contains multiple maternal clades, suggesting potential mitochondrial introgression between R. chensinensis and R. kukunoris.

[Application of SNPstream for medical genetics research].

OBJECTIVE: To introduce the principle, procedure, efficacy and application of SNPstream genotyping technology. METHODS: Genotyping results of 152 SNPs were used to analyze the feasibility, call rate and accuracy of SNPstream technology. RESULTS: For the 152 selected SNPs, 122 SNPs can be genotyped with SNPstream, for which 116 SNPs were successfully genotyped. Replication study showed that the repeatability of genotyping is 99%. When the allele cluster was clear, the accuracy can reach 100%. But when the allele cluster was obscure, the accuracy was only 93.8%. CONCLUSION: SNPstream technology has the advantages of high accuracy, flexible throughput, and high cost performance, and may have a wide application for medical genetics research.