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.

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.

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.

Confirmation of Leptobrachellaventripunctata (Fei, Ye, and Li, 1990), based on molecular and morphological evidence in Thailand.

BACKGROUND: Thailand is considered a global biodiversity hotspot that is known to harbour a striking diversity of endemic species. However, several research studies have determined that the level of amphibian diversity in the country has been significantly underestimated. The megophryid genus Leptobrachella Smith, 1925 is currently known to include 89 species that are primarily distributed throughout southern China and Southeast Asia; however, only seven species have been found in Thailand. NEW INFORMATION: Based on an integrative approach encompassing genetic and morphological analyses, we have concluded that the population identified from Chiang Rai Province of Thailand is conspecific with Leptobrachellaventripunctata (Fei, Ye, and Li, 1990). Importantly, this is the first confirmation record of this species, based on molecular and morphological evidence in Thailand. The discovery of this species reaffirms that the diversity within the genus has been underestimated with many species yet to be discovered. In addition, the findings of our study further highlight the lack of existing knowledge on amphibian taxonomy and an underestimation of the biodiversity that exists along these national border areas.

Integrative taxonomy reveals a new species of Leptobrachella (Anura: Megophryidae) from the mountains of northern Thailand.

The genus Leptobrachella is one of the most speciose and taxonomically troubling groups of Asian anurans. Herein, we describe a new species of Leptobrachella from Doi Inthanon, Chiang Mai Province, Thailand based on the integration of morphological, acoustic and molecular data. The new species, Leptobrachella murphyi sp. nov., is morphologically distinguished from its congeners on the basis of body size, dorsal skin texture and coloration, ventral coloration and pattern, degree of webbing and fringing on the fingers and toes, iris coloration in life, pattern of markings on flanks and pattern of body macroglands. It also differs from its phylogenetically close congeners by an uncorrected p-distance of >9.6% for a fragment of 16S rRNA. The advertisement call of the new species consists of 4.54.7 kHz (at 15 C) and without a distinct introductory note. Leptobrachella murphyi sp. nov. likely occurs across the Thanon Thong Chai Range and analyses provide evidence of unknown biodiversity and species composition on Doi (mountain) Inthanon. In addition, the congeneric species L. minima was also confirmed in Doi Inthanon. The coexistence pattern of Leptobrachella in Doi Inthanon deserves further study. As Thailands highest mountain and biodiversity reservoir, the need for further biological exploration is urgent given ongoing habitat loss and degradation.

Herpetological phylogeographic analyses support a Miocene focal point of Himalayan uplift and biological diversification.

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.

First national record of Quasipaaverrucospinosa (Bourret, 1937) (Amphibia: Anura: Dicroglossidae) from Thailand with further comment on its taxonomic status.

BACKGROUND: Spiny Frog Quasipaa is a genus of frogs that belongs to a relatively poorly known group. Most of the species distribution has been recorded in China; however, a few incidences of identification have occurred in the eastern part of Indochina. To date, only one species (Quasipaafasciculispina) of Quasipaa has been recorded from Chanthaburi and Trat Provinces in south-eastern Thailand. NEW INFORMATION: Based on recent fieldwork conducted in northern Thailand, we report a new record of Quasipaaverrucospinosa from Doi Phu Kha National Park, Nan Province at an altitude of 900-1000 m a.s.l. Our study has demonstrated that populations of this species are paraphyletic and has revealed deep genetic differences. Therefore, it is recommended that a comprehensive study be undertaken to clarify the taxonomic and geographic distribution of this species for its suitable protection and conservation.

Molecular phylogeny and morphological comparisons of the genus Hebius Thompson, 1913 (Reptilia: Squamata: Colubridae) uncover a new taxon from Yunnan Province, China, and support revalidation of Hebius septemlineatus (Schmidt, 1925).

We describe a new species of the genus Hebius and provide evidence for the validity of H. septemlineatuscomb. nov. Morphological and molecular analyses of Hebius specimens collected in Yunnan Province, China, revealed three distinct lineages, namely the newly described Hebius weixiensissp. nov., as well as H. octolineatus (Boulenger, 1904), and H. septemlineatuscomb. nov. (Schmidt 1925), which is removed from synonymy with H. octolineatus. Based on mitochondrial genealogy, Hebius weixiensissp. nov. is sister to H. septemlineatuscomb. nov., while H. octolineatus is sister to H. bitaeniatus. The new species and H. septemlineatuscomb. nov. showed considerable genetic divergence from their recognized congeners (uncorrected P-distance ≥3.9%). Furthermore, the new species and H. septemlineatuscomb. nov. can be diagnosed from closely related congeners by a combination of pholidosis characters.

First records of the fanged frogs Limnonectes bannaensis Ye, Fei & Jiang, 2007 and L. utara Matsui, Belabut & Ahmad, 2014 (Amphibia: Anura: Dicroglossidae) in Thailand.

BACKGROUND: The taxonomic status of the Thai populations belonging to the Limnonectes kuhlii species complex is controversial, due to phenotypic similarity in the cryptic species complex. Recently, some studies on this group in Thailand have discovered four new species: L. taylori, L. megastomias, L. jarujini and L. isanensis. Even so, the diversity of this group is still incomplete. NEW INFORMATION: Based on an integrative approach encompassing genetic and morphological analyses, we conclude that the Limnonectes populations from Nan Province (northern) and Yala Province (southern) of Thailand are conspecific with L. bannaensis Ye, Fei & Jiang, 2007 and L. utara Matsui, Belabut & Ahmad, 2014, respectively. These are the first records of these species in Thailand. Our study highlights the importance of using DNA sequence data in combination with morphological data to accurately document species identity and diversity. This is especially important for morphologically cryptic species complexes and sympatrically occurring congeners.

First national record of Gracixalus quangi Rowley, Dau, Nguyen, Cao & Nguyen, 2011 and G. yunnanensis Yu, Li, Wang, Rao, Wu &Yang, 2019 (Amphibia: Anura: Rhacophoridae) from Thailand.

BACKGROUND: The bushfrog genus Gracixalus Delorme, Dubois, Grosjean & Ohler, 2005 is found in southern and south-western China, Vietnam, Laos, Thailand and Myanmar. It is presently comprised of 17 species. In Thailand, only two species have been recorded, namely G. carinensis (Boulenger) and G. seesom (Massui, Khonsue, Panha & Eto). The latter of these two species is currently known to be endemic to the country. NEW INFORMATION: Based on recent field work conducted in 2019 in Doi Phu Kha National Park, Nan Province of northern Thailand, we are reporting two new records of the genus Gracixalus, G. quangi and G. yunnanensis, from Thailand, based on morphological and molecular evidence. In addition, this is the first study to report on the identification of a female specimen of G. yunnanensis. Furthermore, morphological data and natural history notes of the aforementioned species in Thailand have been provided, along with updated locations for the distribution of both species.

Description of a new species of Bush frog (Anura: Rhacophoridae: Raorchestes) from northwestern Yunnan, China.

Approximately half of the species in speciose genus Raorchestes were described during the past 10 years, yet only 11 species are known from Southeast Asia and southern China (SEA-SC), adjacent Himalayas, and northeastern India. Field work in northwestern Yunnan province, China resulted in the discovery of one new species in the genus based on morphological and molecular analyses. The new species is diagnosed by small size with 15.0-19.0 mm SVL in adult males (n=3); tongue pyriform, notched posteriorly; rudimentary webbing between toes; fingers and toes with narrow lateral dermal fringes; tibiotarsal articulation reaching anterior of the eye when hindlimb is stretched along the side of the body; relative finger lengths: I < II < IV < III, relative toe lengths: I < II < V < III < IV; inner metatarsal tubercle oval, outer metatarsal tubercle absent; finger discs and toe discs greyish or orange; flank near the crotch with a distinct black region between two creamy white patches, and the thigh having a similar black patch near the groin, proximal to another creamy white patch; a distinct ") ("-shaped dark marking on the back; male with external single subgular vocal sac; nuptial pad absent. A phylogenetic tree was reconstructed based on the mitochondrial genes for 16S rRNA and ND1. The results indicated that these individuals form a monophyletic group, and show high genetic divergence to their closest relatives within the genus (uncorrected p-distances > 3.2%) by distance of 16S comparable to the divergence between recognized Raorchestes species. This study further enriches the diversity of rhacophorids, especially in northwestern Yunnan.

New country records for three species of frog from Myanmar including two genera (Nasutixalus and Oreolalax).

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.

A combined approach of mitochondrial DNA and anchored nuclear phylogenomics sheds light on unrecognized diversity, phylogeny, and historical biogeography of the torrent frogs, genus Amolops (Anura: Ranidae).

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.

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.

A new species of the genus Raorchestes (Anura: Rhacophoridae) from Yunnan Province, China.

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.

A new species of the genus Xenophrys Anura Megophryidae from northern Thailand.

Species of Xenophrys are conserved morphologically and live primarily in forests. In Thailand, the genus harbors many cryptic species. Herein we report the collection of specimens from Doi Inthanon, Chiang Mai Province, northern Thailand, which were identified previously as X. minor. Molecular and morphological analyses find that these specimens differ significantly from other known congeners, and therefore we describe a new species. Further, our phylogenetic analyses indicate that X. latidactyla is a junior synonym of X. palpebralespinosa.

[Physiology and genetics research progress of teat traits in pigs].

The pig teat traits are important indices of genetic improvement in pig breeding, which belong to reproductive traits and can directly affect the sows lactation rate and piglet survival rate. Understanding the genetic mechanism underlying the variation of teat traits is of immense value for the improvement of pig reproductive performance. However, the genetic mechanism underlying teat traits (including teat number, type, location distribution, and fluctuating asymmetry) remains elusive. In this review, we summarize the studies on physiology and genetics of teat traits in pigs, including the development process of the mammary gland, the QTL mapping, and candidate gene researches. This review aims to provide a new perspective for the identification of causal mutations and major genes affecting the teat traits and revealing the complex genetic mechanism of the differences in teat number, type and location distribution during embryonic development in pigs.

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.