A new species of the genus Hebius (Squamata, Natricidae), previously confused with H. boulengeri (Gressitt, 1937)

263 A new species of the genus Hebius (Squamata, Natricidae), previously confused with H. boulengeri (Gressitt, 1937) Shuo Liu1,2* , JiShan Wang3* , Mian Hou4, Liang Zhang5, Qiaoyan Wang6, Chunmiao Zong6, Jiang Zhou7, Dingqi Rao8, Patrick David9, Gernot Vogel10 1 Kunming Natural History Museum of Zoology, Kunming Institute of Zoology, Chinese Academy of Sciences, Kunming, Yunnan 650223, China 2 Yunnan Key Laboratory of Biodiversity Information, Kunming Institute of Zoology, Chinese Academy of Sciences, Kunming, Yunnan 650201, China 3 Southwest Survey and Planning Institute of National Forestry and Grassland Administration, Kunming, Yunnan 650031, China 4 College of Continuing (Online) Education, Sichuan Normal University, Chengdu, Sichuan 610066, China 5 Guangdong Key Laboratory of Animal Conservation and Resource Utilization, Guangdong Public Laboratory of Wild Animal Conservation and Utilization, Institute of Zoology, Guangdong Academy of Sciences, Guangzhou, Guangdong 510260, China 6 Research Institute of Yunnan Xishuangbanna National Nature Reserve, Jinghong, Yunnan 666100, China 7 Management and Protection Bureau of Mojiang Xiqi Alsophila Provincial Nature Reserve, Mojiang, Yunnan 654800, China 8 Kunming Institute of Zoology, Chinese Academy of Sciences, Kunming, Yunnan 650201, China 9 Reptiles & Amphibiens, Institut de Systématique, Évolution et Biodiversité (ISYEB), Muséum National d’Histoire Naturelle, CP 30, 57 rue Cuvier, F-75005 Paris, France 10 Society for South East Asian Herpetology, Im Sand-3, D-69115 Heidelberg, Germany Corresponding authors: Gernot Vogel (Gernot.V[email protected]); Patrick David (patrick.da[email protected]); Dingqi Rao ([email protected]) Copyright: © Shuo Liu et al. This is an open access article distributed under terms of the Creative Commons Attribution License (Attribution 4.0 International – CC BY 4.0). Research Article Abstract A new species of the genus Hebius is described from Yunnan Province, China. The new species was previously identified as H. boulengeri, but it can be separated from H. boulengeri and other species of this genus by a combination of the following characters: 19-19-17 dorsal scale rows, 148–155 ventrals, 90–105 subcaudals, one preocular, three postoculars, 23–25 maxillary teeth, a distinct continuous white streak from posteroventral margin of eye extending backward and upward to nape to connect with dorsolateral stripe on each side, and white venter. The genetic divergence between the new species and H. boulengeri was 6.6% and the genetic divergences between the new species and other congeners ranged from 6.8% to 15.9% in the mitochondrial cytochrome b gene. Key words: Keelback snake, mitochondrial gene, morphological comparison, phylogenetic analysis, Pu’er City, taxonomy, Xishuangbanna Prefecture Introduction The genus Hebius Thompson, 1913 is the most speciose genus in the family Natricidae (Bohra et al. 2025; Uetz et al. 2025). It is a group of small to medium-sized, non-venomous, semi-aquatic snakes that are widely distributed throughout southern and southeastern Asia (Guo et al. 2014; Liu et al. 2018; Purkayastha and David 2019; Deepak et al. 2022; Hauser et al. 2022; Ma et al. 2023; Xu et al. 2023, 2024; Bohra et al. 2025). Due to interspecific morphological similarities, the species of this genus often cannot be accurately identified by morphology, which leads to the species diversity of this genus being far underestimated (Liu et al. 2018; Ren et al. 2018; Xu et al. 2023). Academic editor: Minh Duc Le Received: 4 June 2025 Accepted: 19 August 2025 Published: 3 October 2025 ZooBank: https://zoobank. org/01F3164F-DDBE-41E3-B50AB7AF025DC2BC Citation: Liu S, Wang J, Hou M, Zhang L, Wang Q, Zong C, Zhou J, Rao D, David P, Vogel G (2025) A new species of the genus Hebius (Squamata, Natricidae), previously confused with H. boulengeri (Gressitt, 1937). ZooKeys 1254: 263–282. https://doi. org/10.3897/zookeys.1254.161130 ZooKeys 1254: 263–282 (2025) DOI: 10.3897/zookeys.1254.161130 * These authors contributed equally to this work. 264 ZooKeys 1254: 263–282 (2025), DOI: 10.3897/zookeys.1254.161130 Shuo Liu et al.: A new species of Hebius from China Boulenger’s keelback, H. boulengeri (Gressitt, 1937), was originally described as Natrix boulengeri by Gressitt (1937) from Tai-yong, Eastern Kwangtung, China (Jieyang City, Guangdong Province, China). Subsequently, this species was transferred to the genus Amphiesma and later to the genus Hebius (Guo et al. 2014). Previously, this species was considered to be widely distributed in southern China, including Guangdong, Fujian, Jiangxi, Hunan, Hainan, Guizhou, and Yunnan provinces, Chongqing Municipality, Guangxi Autonomous Region, and Hong Kong Special Administrative Region, as well as Vietnam and Cambodia (Huang 2021; Wang 2021; Uetz et al. 2025). In Yunnan Province, H. boulengeri was recorded from Honghe, Xishuangbanna, Dehong, Nujiang, and Diqing prefectures, as well as Pu’er, Lincang, and Baoshan cities (Zhao et al. 1998; Yang and Rao 2008). During our fieldwork in Yunnan Province, China, from 2018 to 2023, four specimens superficially resembling H. boulengeri were collected from Xishuangbanna Prefecture and Pu’er City. Morphological comparison and molecular data analysis showed that these specimens are distinct from H. boulengeri and other species of the genus. Herein, we describe a new species of the genus Hebius based on these four specimens. Material and methods The specimens were collected by hand and stored in 75% ethanol after being photographed, and then deposited at Kunming Natural History Museum of Zoology, Kunming Institute of Zoology, Chinese Academy of Sciences (KIZ). Measurements were taken using digital calipers to the nearest 0.1 mm. Bilateral scale counts were given as left/right. The abbreviations of measurements and scale counts are as follows: ATem number of anterior temporals DLip distance between the lower edge of the lip and the lower margin of the eye DS numbers of dorsal scale rows behind the neck, at the midbody, and before the vent, respectively EDH horizontal diameter of the eye EDV vertical diameter of the eye HL head length IL number of infralabials MXT number of maxillary teeth PosOc number of postoculars PreOc number of preoculars Sc number of subcaudals SL number of supralabials SnL snout length, between the eye and the anterior margin of the rostral SVL snout–vent length TaL tail length TL total length Ven number of ventrals 265 ZooKeys 1254: 263–282 (2025), DOI: 10.3897/zookeys.1254.161130 Shuo Liu et al.: A new species of Hebius from China We examined 81 specimens of H. boulengeri from various museums, including the holotype and paratype. The morphological data of other species of this genus were obtained from literature (Gressitt 1937; Zhao et al. 1998; Yang and Rao 2008; Liu et al. 2018; Ren et al. 2018; Purkayastha and David 2019; Huang 2021; Hauser et al. 2022; Ma et al. 2023; Xu et al. 2023, 2024; Bohra et al. 2025). Total Genomic DNA for the four newly collected specimens was extracted from the liver tissue. A fragment of the mitochondrial cytochrome b gene (cytb) was amplified using the primer pairs L14910: 5′-GACCTGTGATMTGAAAAACCAYCGTTGT-3′ and H16064: 5′-CTTTGGTTTACAAGAACAATGCTTTA-3′ (Burbrink et al. 2000). The amplification products were purified and sequenced by Sangon Biotech (Shanghai) Co., Ltd. The generated sequences were assembled using SeqMan in Lasergene 7.1 (Burland 2000). Amphiesma stolatum (Linnaeus, 1758) was used as outgroup according to Bohra et al. (2025). The sequences of 34 species of the genus Hebius and the outgroup were obtained from GenBank (Table 1). Sequences were aligned using MAFFT 7.471 (Katoh and Standley 2013) with default parameters. The genetic divergence (uncorrected p-distance) between species was calculated in MEGA 12.0.10 (Kumar et al. 2024). Bayesian-inference (BI) and maximum-likelihood analysis (ML) were used to construct the phylogenetic tree. The best-fit substitution models for BI and ML, respectively, were chosen using the Akaike information criterion in ModelFinder (Kalyaanamoorthy et al. 2017). The BI was performed in MrBayes 3.2.6 (Ronquist et al. 2012) using the model GTR+F+I+G4, and the Markov chains were run for one million generations with sampled every 100 generations. The ML was performed in IQ-TREE 1.6.12 (Nguyen et al. 2015) using the model TIM2+F+I+G4, and nodal support was estimated by 1,000 ultrafast bootstrap replicates. Results The phylogenetic trees resulted in very similar topologies by BI and ML. The sequences of the newly collected specimens from Yunnan clustered together to form a distinct lineage between the clade of H. boulengeri and the clade consisting of H. gilhodesi (Wall, 1925) and H. khasiensis (Boulenger, 1890), but the exact phylogenetic position of the novel lineage was not resolved (Fig. 1). The genetic divergence between the novel lineage and H. boulengeri was 6.6%, the genetic divergence between the novel lineage and H. khasiensis was 6.8%, and the genetic divergence between the novel lineage and H. gilhodesi was 7.6% (Table 2). Morphologically, the newly collected specimens from Yunnan can be distinguished from all named species of the genus. The newly collected specimens differ from all other species of the genus, except H. boulengeri, in having a distinct continuous white streak from the posteroventral margin of the eye extending backward and upward to the nape on each side. The newly collected specimens differ from H. boulengeri in having fewer maxillary teeth (Suppl. material 1). Based on the results of molecular and morphological analysis, the newly collected specimens from Yunnan should be regarded as a distinct new species, which we formally describe here. 266 ZooKeys 1254: 263–282 (2025), DOI: 10.3897/zookeys.1254.161130 Shuo Liu et al.: A new species of Hebius from China Table 1. Sequences of Hebius species and outgroup used in this study. Species name locality Voucher Accession Reference H. shantianfangi sp. nov. Mengla, Yunnan, China KIZ 064970 PX206432 This study H. shantianfangi sp. nov. Pu’er, Yunnan, China KIZ 064971 PX206433 This study H. shantianfangi sp. nov. Mojiang, Yunnan, China KIZ 064972 PX206434 This study H. shantianfangi sp. nov. Mojiang, Yunnan, China KIZ 064973 PX206435 This study H. andreae Khammouane, Laos VNUF R.2017.25 MK253674 Ziegler et al. 2019 H. annamensis Xe Kong, Laos FMNH 258637 OK315812 Deepak et al. 2021 H. atemporalis Pu Hoat, Nge An, Vietnam ZMMU NAP-07877 OK315813 Deepak et al. 2021 H. bitaeniatus Chiang Mai, Thailand AUP-00062 OK315816 Deepak et al. 2021 H. boulengeri Ha Giang, Vietnam AMNH 148562 KJ685664 Guo et al. 2014 H. boulengeri Tam Dao, Vinh Phuc, Vietnam ZMMU NAP-08675 OK315817 Deepak et al. 2021 H. boulengeri Hechi, Guagnxi, China SYS r001552 MK201520 Li et al. 2020 H. boulengeri Fujian, China GP2433 KJ685699 Guo et al. 2014 H. boulengeri Guangdong, China GP1789 KJ685684 Guo et al. 2014 H. boulengeri Chebaling, Guangdong, China RE55 MK201380 Li et al. 2020 H. boulengeri Jinpenshan, Jiangxi, China SYS r001371 MK201483 Li et al. 2020 H. boulengeri Hainan, China GP2134 KJ685691 Guo et al. 2014 H. boulengeri Hainan, China MVZ 236752 KJ685710 Guo et al. 2014 H. boulengeri Hainan, China HS11107 MK201295 Li et al. 2020 H. boulengeri Dawuling, Guangdong, China SYS r001506 MK201509 Li et al. 2020 H. chapaensis Lao Cai, Vietnam VNMN 06103 MH778700 Ren et al. 2018 H. citrinoventer Yingjiang, Yunnan, China ANU 20230016 PP472750 Xu et al. 2024 H. clerki Nujiang, Yunnan, China CAS 215036 KJ685666 Guo et al. 2014 H. concelarus Miyakojima, Okinawa, Japan KUZ:R20253 AB989268 Kaito and Mamoru 2016 H. craspedogaster Guizhou, China GP1240 KJ685672 Guo et al. 2014 H. deschauenseei Chiang Mai, Thailand AUP-00182 OK315827 Deepak et al. 2021 H. gilhodesi Kachin state, Myanmar CAS 221504 KJ685668 Guo et al. 2014 H. gilhodesi Kachin state, Myanmar CAS 221525 KJ685669 Guo et al. 2014 H. igneus Ha Giang, Vietnam AMNH 148575 KJ685665 Guo et al. 2014 H. ishigakiensis Iriomotejima, Okinawa, Japan KUZ:R33043 AB989292 Kaito and Mamoru 2016 H. jingdongensis Jingdong, Yunnan, China CIB 119044 OR285310 Ma et al. 2023 H. johannis Yunnan, China KIZ 014484 MZ570479 Hou et al. 2021 H. khasiensis Mairang, Meghalaya, India MZMU 3526 PQ288047 Bohra et al. 2025 H. khasiensis Sailam, Aizawl, Mizoram, India MZMU 2540 PQ288048 Bohra et al. 2025 H. maximus Sichuan, China GP864 KJ685706 Guo et al. 2014 H. metusia Sichuan, China GP871 KJ685707 Guo et al. 2014 H. modestus Yunnan, China CAS 234262 KJ685671 Guo et al. 2014 H. octolineatus Yunnan, China GP1569 KJ685678 Guo et al. 2014 H. optatus Guizhou, China GP1885 KJ685687 Guo et al. 2014 H. popei Hainan, China GP2169 KJ685692 Guo et al. 2014 H. pryeri Tokunoshima, Kagoshima, Japan KUZ:R34062 AB989126 Kaito and Mamoru 2016 H. sangzhiensis Hunan, China SYNU 08070350 MK340763 Zhou et al. 2019 H. sauteri Taiwan, China GP2549 KJ685701 Guo et al. 2014 H. septemlineatus Tengchong, Yunnan, China KIZ 037720 MZ570486 Hou et al. 2021 H. taronensis Kachin State, Myanmar CAS 224426 OK315828 Deepak et al. 2021 H. venningi Kachin, Myanmar CAS 233206 KJ685670 Guo et al. 2014 H. vibakari Kyoto, Japan KUZ:R21587 AB989302 Kaito and Mamoru 2016 H. weixiensis Weixi, Yunnan, China KIZ 035740 MZ570488 Hou et al. 2021 H. yanbianensis Yanbian, Sichuan, China GP4006 MH532291 Guo et al. 2014 H. youjiangensis Baise, Guangxi, China HSR22184 OQ085073 Xu et al. 2023 H. sp. Quang Nam, Vietnam AMNH 148552 KJ685663 Guo et al. 2014 H. sp. Kon Ka Kinh, Gia Lai, Vietnam ZMMU R-16531 OK315818 Deepak et al. 2021 H. sp. Vietnam HT0679 LC325327 Takeuchi et al. 2018 Amphiesma stolatum Ha Giang, Vietnam CAS 215037 KJ685667 Guo et al. 2014 267 ZooKeys 1254: 263–282 (2025), DOI: 10.3897/zookeys.1254.161130 Shuo Liu et al.: A new species of Hebius from China Figure 1. Bayesian phylogenetic tree based on cytb sequences. The numbers after and behind “/” indicate the Bayesian posterior probabilities (above 0.90 retained) and maximum likelihood ultrafast bootstrap values (above 90 retained), respectively. Amphiesma stolatum KJ685667 Hebius andreae MK253674 Hebius youjiangensis OQ085073 Hebius modestus KJ685671 Hebius deschauenseei OK315827 Hebius chapaensis MH778700 Hebius igneus KJ685665 Hebius annamensis OK315812 Hebius clerki KJ685666 Hebius sangzhiensis MK340763 Hebius vibakari AB989302 Hebius maximus KJ685706 Hebius ishigakiensis AB989292 Hebius concelarus AB989268 Hebius pryeri AB989126 Hebius optatus KJ685687 Hebius sauteri KJ685701 Hebius popei KJ685692 Hebius atemporalis OK315813 Hebius yanbianensis MH532291 Hebius craspedogaster KJ685672 Hebius octolineatus KJ685678 Hebius johannis MZ570479 Hebius metusia KJ685707 Hebius bitaeniatus OK315816 Hebius septemlineatus MZ570486 Hebius weixiensis MZ570488 Hebius citrinoventer PP472750 Hebius taronensis OK315828 Hebius venningi KJ685670 Hebius jingdongensis OR285310 LC325327 KJ685663 OK315818 Hebius sp. KJ685684 MK201380 MK201483 KJ685699 MK201509 KJ685691 KJ685710 MK201295 MK201520 KJ685664 OK315817 Hebius boulengeri KIZ 064972 KIZ 064971 KIZ 064973 KIZ 064970 Hebius shantianfangi sp. nov. KJ685668 KJ685669 Hebius gilhodesi PQ288047 PQ288048 Hebius khasiensis 0.10 1/100 1/96 1/98 1/100 0.90/91 -/90 1/97 0.99/92 1/99 1/100 1/99 1/100 -/- -/- 1/100 -/- -/- -/- -/- -/- -/- 0.94/- -/- -/- 1/100 1/99 1/99 1/100 0.98/91 0.98/- 1/100 1/100 1/100 1/100 1/100 -/- 1/100 1/91 1/98 1/93 1/94 268 ZooKeys 1254: 263–282 (2025), DOI: 10.3897/zookeys.1254.161130 Shuo Liu et al.: A new species of Hebius from China Table 2. Percentage genetic divergence (uncorrected p-distance) based on cytb sequences between species of Hebius. Values between the new species and closely related species are highlighted in bold. 1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 1 Hebius shantianfangi sp. nov. 2 Hebius andreae 15.9 3 Hebius annamensis 14.0 14.8 4 Hebius atemporalis 11.8 16.3 14.5 5 Hebius bitaeniatus 11.9 15.4 13.9 10.9 6 Hebius boulengeri 6.6 16.2 13.0 12.0 10.8 7 Hebius chapaensis 12.9 14.2 6.3 13.1 13.6 12.4 8 Hebius citrinoventer 13.3 16.2 14.2 12.9 10.5 13.1 14.1 9 Hebius clerki 11.2 15.7 13.5 12.3 12.3 10.7 12.3 12.9 10 Hebius concelarus 13.6 18.0 16.1 14.4 13.2 13.7 15.4 13.8 14.0 11 Hebius craspedogaster 11.3 16.2 13.2 11.4 11.8 10.8 13.0 12.0 12.0 14.0 12 Hebius deschauenseei 13.2 15.4 6.9 14.3 13.6 13.2 6.1 14.5 13.3 15.4 13.2 13 Hebius gilhodesi 7.6 16.6 14.4 12.4 12.3 7.8 13.3 14.3 11.7 13.4 12.2 13.7 14 Hebius igneus 13.9 15.3 6.0 13.4 13.3 13.4 1.0 14.4 13.1 15.2 13.0 5.2 14.0 15 Hebius ishigakiensis 12.4 17.0 14.2 12.6 11.7 12.2 14.1 12.9 11.9 12.1 12.4 14.4 13.1 14.1 16 Hebius jingdongensis 10.5 14.8 12.6 11.0 11.2 9.6 12.5 11.7 12.3 14.0 10.4 11.9 11.2 12.4 12.7 17 Hebius johannis 10.0 14.2 12.0 11.0 10.7 10.0 11.9 11.1 11.3 12.5 5.0 12.4 11.5 12.3 12.3 10.4 18 Hebius khasiensis 6.8 16.0 13.3 12.5 12.7 7.5 12.0 13.3 11.8 14.0 11.9 12.7 6.2 12.6 13.1 10.1 10.9 19 Hebius maximus 13.1 14.5 13.7 13.1 10.8 13.4 13.2 13.6 12.3 13.5 11.8 12.5 13.8 13.2 11.2 12.8 11.7 20 Hebius metusia 10.9 15.2 13.1 10.6 10.4 10.6 12.5 11.8 12.1 13.3 4.6 12.5 12.0 12.7 12.2 9.8 4.7 21 Hebius modestus 14.7 15.6 7.0 15.0 13.5 14.1 5.6 14.5 13.1 16.0 14.4 6.1 15.0 5.1 13.7 12.8 13.2 22 Hebius octolineatus 11.5 14.7 12.9 11.0 11.6 10.6 12.2 11.8 11.8 12.8 5.1 12.7 12.2 12.8 12.1 10.1 5.3 23 Hebius optatus 13.2 17.1 15.5 13.1 12.0 13.0 14.1 13.7 13.4 12.8 12.4 15.1 13.7 14.8 11.8 13.2 12.0 24 Hebius popei 12.3 16.8 14.8 13.1 12.3 13.3 14.5 14.4 12.7 14.4 13.4 14.5 13.6 15.3 12.4 12.0 12.4 25 Hebius pryeri 12.8 17.3 15.7 12.8 12.8 12.8 15.5 13.0 12.7 9.1 13.1 15.8 14.1 15.5 10.4 13.3 12.3 26 Hebius sangzhiensis 12.9 15.3 14.5 13.6 10.8 12.6 14.0 13.4 13.0 12.0 11.8 13.3 12.8 14.1 11.1 12.8 10.9 27 Hebius sauteri 11.9 16.7 14.4 12.1 11.2 12.4 13.8 12.0 12.0 12.9 10.7 14.2 12.2 13.9 10.7 10.2 11.0 28 Hebius septemlineatus 11.3 15.3 14.0 11.6 3.5 10.9 13.8 10.5 12.1 12.7 11.2 13.2 12.0 13.7 11.5 10.6 10.5 29 Hebius taronensis 11.4 15.4 14.7 13.0 10.1 13.0 13.5 10.8 12.7 13.8 11.9 13.5 13.6 14.3 12.4 12.0 11.0 30 Hebius venningi 11.6 14.1 14.2 11.8 8.8 12.3 12.2 9.7 12.0 13.0 11.0 13.5 12.8 13.1 12.1 11.1 9.8 31 Hebius vibakari 13.0 14.2 13.9 12.3 10.5 12.3 13.7 13.4 13.3 13.2 12.1 13.8 13.3 13.5 12.0 11.6 11.5 32 Hebius weixiensis 11.6 15.3 13.3 10.9 6.4 10.6 12.7 10.8 11.0 12.7 10.1 13.5 12.0 12.9 12.1 10.2 10.5 33 Hebius yanbianensis 11.5 15.4 13.7 11.2 11.2 10.8 13.1 11.8 11.9 13.3 5.1 13.2 12.1 13.3 12.1 11.0 6.1 34 Hebius youjiangensis 14.4 15.8 7.3 14.3 14.2 14.0 5.6 14.6 12.4 15.5 13.9 5.9 14.7 5.4 14.2 12.7 13.0 35 Hebius sp. 9.3 15.3 14.4 13.0 12.6 9.0 13.3 13.4 11.6 13.8 11.9 13.8 10.6 13.8 12.4 11.1 11.0 269 ZooKeys 1254: 263–282 (2025), DOI: 10.3897/zookeys.1254.161130 Shuo Liu et al.: A new species of Hebius from China 18 19 20 21 22 23 24 25 26 27 28 29 30 31 32 33 34 19 Hebius maximus 14.0 20 Hebius metusia 11.2 11.8 21 Hebius modestus 13.2 13.0 14.2 22 Hebius octolineatus 11.1 12.0 4.9 13.9 23 Hebius optatus 14.7 11.6 12.3 14.9 12.8 24 Hebius popei 13.0 12.2 12.8 14.3 13.0 11.6 25 Hebius pryeri 14.0 12.1 12.0 15.3 11.8 12.9 13.7 26 Hebius sangzhiensis 13.3 7.6 12.3 13.7 11.5 12.0 12.8 11.1 27 Hebius sauteri 12.9 11.2 10.8 14.2 11.5 11.4 11.6 11.8 11.5 28 Hebius septemlineatus 12.3 10.4 10.2 13.8 10.6 11.8 11.5 12.3 10.4 10.7 29 Hebius taronensis 12.9 11.4 11.4 14.1 12.0 13.2 12.2 13.0 12.3 12.2 9.5 30 Hebius venningi 12.7 11.6 10.1 13.4 10.9 12.3 11.8 12.1 12.1 11.8 9.2 5.6 31 Hebius vibakari 13.4 8.2 11.6 13.5 11.6 11.1 12.8 12.4 6.1 11.2 10.0 11.5 11.2 32 Hebius weixiensis 11.9 11.1 9.8 13.5 10.4 11.5 12.8 12.5 10.4 10.7 5.7 10.3 9.2 10.3 33 Hebius yanbianensis 11.7 11.5 5.4 14.2 6.4 12.4 13.0 12.4 11.4 11.5 10.6 11.7 10.5 11.7 9.3 34 Hebius youjiangensis 13.5 13.4 13.9 4.1 13.5 15.2 14.7 15.7 13.9 13.9 14.2 14.6 13.8 13.7 13.5 14.5 35 Hebius sp. 10.2 12.9 11.8 14.5 11.1 13.9 14.7 13.1 13.4 13.1 12.9 13.2 12.6 13.1 12.7 12.4 14.3 Taxonomic accounts Hebius shantianfangi sp. nov. https://zoobank.org/B7CE0216-9A4B-42B6-9AFA-40BDB92CBA73 Figs 2–4, 5A, 6A Type materials. Holotype • KIZ 064970; adult female; Longlin Village, Mengla Town, Mengla County, Xishuangbanna Prefecture, Yunnan Province, China; 21°38′15″N, 101°27′0″E, 1,040 m; 27 April 2019; JiShan Wang leg. Paratypes • KIZ 064971; adult male; Yutang Village, Yixiang Town, Simao District, Pu’er City, Yunnan Province, China; 22°36′58″N, 101°7′32″E, 1,300 m; 18 September 2018; Mian Hou leg. • KIZ 064972; adult female; Guangfeng Village, Sinanjiang Town, Mojiang County, Pu’er City, Yunnan Province, China; 22°56′49″N, 101°49′51″E, 1810 m; 31 May 2023; Mo Wang leg. • KIZ 064973; adult female; Bingli Village, Yutang Town, Mojiang County, Pu’er City, Yunnan Province, China; 23°7′10″N, 101°31′46″E, 1850 m; 26 June 2023; Mo Wang leg. Diagnosis. Body slender, SVL 332–406 mm, TL 506–610 mm; tail relatively long, TaL/TL 0.31–0.34; dorsal scales in 19-19-17 rows, all strongly keeled except for 1st row on each side; ventrals 148–155; cloacal plate divided; subcaudals 90–105, paired; loreal one, not entering orbit; preocular one, postoculars three; supralabials mostly nine, rarely ten, mostly 4th–6th entering orbit, rarely 5th–7th entering orbit; infralabials 9–11; maxillary teeth 23–25, last one or two distinctly enlarged, no diastema separating enlarged posterior teeth with anterior ones; a distinct continuous white streak from posteroventral margin of eye extending backward and upward to nape on each side; dorsolateral stripes 270 ZooKeys 1254: 263–282 (2025), DOI: 10.3897/zookeys.1254.161130 Shuo Liu et al.: A new species of Hebius from China Figure 2. The type specimens of Hebius shantianfangi sp. nov. in preservative. A. Dorsal view; B. Ventral view. 271 ZooKeys 1254: 263–282 (2025), DOI: 10.3897/zookeys.1254.161130 Shuo Liu et al.: A new species of Hebius from China Figure 3. The holotype (KIZ 064970) of Hebius shantianfangi sp. nov. in life. A. Dorsal view; B. Ventral view. present; venter white, ventrolateral blotches present, composed of a dark grey spot on edge of each ventral on each side. Etymology. Named after the late renowned Chinese storytelling artist, Tianfang Shan (17.12.1934–11.09.2018). This species was previously confused 278 ZooKeys 1254: 263–282 (2025), DOI: 10.3897/zookeys.1254.161130 Shuo Liu et al.: A new species of Hebius from China species is unknown. Other species of amphibians and reptiles found at the type locality include Amolops vitreus (Bain, Stuart & Orlov, 2006), Hylarana menglaensis Fei, Ye & Xie, 2008, Kurixalus odontotarsus (Ye & Fei, 1993), Limnonectes bannaensis Ye, Fei & Jiang, 2007, L. limborgi (Sclater, 1892), Leptobrachella eos (Ohler, Wollenberg, Grosjean, Hendrix, Vences, Ziegler & Dubois, 2011), L. ventripunctata (Fei, Ye & Li, 1990), Microhyla mukhlesuri Hasan, Islam, Kuramoto, Kurabayashi & Sumida, 2014, Polypedates impresus Yang, 2008, Zhangixalus pachyproctus Yu, Hui, Hou, Wu, Rao & Yang, 2019, Acanthosaura rubrilabris Liu, Rao, Hou, Orlov, Ananjeva & Zhang, 2022, Calotes emma Gray, 1845, Pareas berdmorei Theobald, 1868, and Trimeresurus lanna Idiiatullina, Nguyen, Pawangkhanant, Suwannapoom, Chanhome, Mirza, David, Vogel & Poyarkov, 2024. Discussion Through morphological and molecular data, this study demonstrated that the populations from Mengla County of Xishuangbanna Prefecture and Mojiang County of Pu’er City, Yunnan Province, which were previously considered as H. boulengeri, represent a distinct species, which we described herein as new. In addition, H. boulengeri was also considered to be distributed in other areas Figure 7. Map showing the distributions of Hebius shantianfangi sp. nov. (red), H. boulengeri sensu stricto (green), and Hebius cf. boulengeri (blue). Star indicates type locality. 279 ZooKeys 1254: 263–282 (2025), DOI: 10.3897/zookeys.1254.161130 Shuo Liu et al.: A new species of Hebius from China of Yunnan Province, such as Hekou, Lvchun, and Pingbian counties of Honghe Prefecture, Menglian County of Pu’er City, Cangyuan County of Lincang City, Longling County of Baoshan City, Gongshan County of Nujiang Prefecture, and Weixi County of Diqing Prefecture (Yang and Rao 2008). For the populations that were identified as H. boulengeri in these areas of Yunnan Province, we speculate that they also do not belong to H. boulengeri sensu stricto. Due to the relatively conservative morphology and the lack of available molecular data for these populations, their taxonomic status remains to be determined. Phylogenetic analysis revealed relatively significant genetic differentiation within the populations currently referred to as Hebius boulengeri (Fig. 1). The sequences of specimens from Guangdong (KJ685684, MK201380, and MK201509), Fujian (KJ685699), Jiangxi (MK201483), and Hainan (KJ685691, KJ685710, and MK201295) provinces of China clustered together to form a less divergence clade, with 1.8% genetic distance within it. The sequences of specimens from northwestern Guangxi Autonomous Region of China (MK201520) and northern Vietnam (KJ685664 and OK315817) formed two distinct clades, respectively, and there were relatively large genetic distances (3.3–4.4%) between them and the clade from Guangdong, Fujian, Jiangxi, and Hainan. In addition, the other two specimens from Guangdong we examined are in good agreement with the type specimens in morphology, while the specimens from Guangxi Autonomous Region and Guizhou Province of China, Vietnam, Cambodia, Laos, and Thailand, show some morphological differences (e.g. number of ventrals, subcaudals, and postoculars; Suppl. material 1) from the type specimens and from each other. Therefore, it is likely that H. boulengeri sensu stricto is restricted to Guangdong, Fujian, Jiangxi, and Hainan provinces, and Hong Kong Special Administrative Region, in China. The populations of H. boulengeri sensu lato from Guangxi and Guizhou of China, Vietnam, Cambodia, Laos, and Thailand, may represent multiple cryptic species, which are currently best referred to as Hebius cf. boulengeri. Further studies combining morphological and molecular data will reveal their taxonomic status. Acknowledgements We thank the staff and forest rangers of Yunnan Xishuangbanna National Nature Reserve and Mojiang Xiqi Alsophila Provincial Nature Reserve for their assistance in the field survey. Thanks to Mo Wang (Southwest Forestry University) for collecting and providing specimens. We thank Mark-Oliver Rödel and Frank Tillack (ZMB), Alain Dubois, Ivan Ineich, and Nicolas Vidal (MNHN), Jimmy A. McGuire and Carol Spencer (MVZ), Jiafan Ma and Jianqiang Cen (SMNH), Yuezhao Wang and Xiaomao Zeng (CIB), David Kizirian and Lauren Vonnahme (AMNH), Truong Quang Nguyen, Canh Xuan Le, and Dang Xuan Nguyen (IEBR), Lauren Scheinberg, Erica Ely, Jens Vindum, and Alan Leviton (CAS), Alan Resetar (FMNH), Patrick Campbell (BMNH), Amy Lathrop and Robert W. Murphy (ROM), and Lee Grismer (LSUHC) for permission to examine specimens under their care. We also thank the editors and reviewers for their working on the manuscript. Additional information Conflict of interest The authors have declared that no competing interests exist. 280 ZooKeys 1254: 263–282 (2025), DOI: 10.3897/zookeys.1254.161130 Shuo Liu et al.: A new species of Hebius from China Ethical statement No ethical statement was reported. Use of AI No use of AI was reported. Funding This work was supported by the project of the second comprehensive scientific investigation of Yunnan Xishuangbanna National Nature Reserve, Biological Resources Programme, Chinese Academy of Sciences, the Position of Bioclassonomist of Chinese Academy of Sciences (No. CAS-TAX-24), Foundation of Yunnan Key Laboratory of Biodiversity Information, Kunming Institute of Zoology, Chinese Academy of Sciences, “Support Plan for Revitalizing Yunnan talents” Youth Talent Special Project (No. 220621198312300734, 152301198408141020), the Natural Science Foundation of Yunnan Province of China (No. 202201AT070046), and the project of Ministry of Ecology and Environment of China: investigation and assessment of amphibians and reptiles in Jinghong City, Menghai County, and Mengla County. Author contributions Conceptualization: SL, MH, GV, PD, DR. Formal analysis: SL, JW. Investigation: SL, JW, MH, LZ, QW, CZ, JZ. Writing – original draft: SL. 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Science Press, Beijing, 522 pp. Appendix 1 Specimens examined Hebius boulengeri (n = 81): MVZ 23623, holotype, Jieyang, Guangdong, China; MVZ 23622, paratype, Ganzhou, Jiangxi, China; ZMB 27694, northern Guangdong, China; SMNH 1238, Guangdong, China; CIB 602339, Laibin, Guangxi, China; CIB 63II5098, Leishan, Guizhou, China; AMNH 148552, 148562, Ha Giang, Vietnam; IEBR 1290, MNHN 1997.3307, 1999.9093–1999.9094, 1935.0061– 1935.0063, 1935.0451–1935.0452, 1935.0453, 1958.0459–1958.0460, PSGV 0002 S, MVZ 224141–224143, 224145–224148, 224153, 226513, MVZ 226515, Tam Dao, Vinh Phu, Vietnam; CAS 73737, FMNH 71709, southern Vietnam; BMNH 1921.4.1.3–1921.4.1.5, 1969.1716–1969.1718, Lang Bian, Vietnam; FMNH 252117, 252120–252123, Ankhe, Gia Lai, Vietnam; IEBR 353, 1650–1652, 1654–1656, 1658–1662, 2540–2541, 2537, Kon Tum, Vietnam; IEBR 2977, ROM 03848, 21434, 21438, Tay Gian, Quang Nam, Vietnam; BMNH 1928.6.29.9, 1969.1710–1969.1715, 1969.1720, Bokor Mts., Cambodia; LSUHC 07442– 07444, 07464–07465, 07484, Phnom Aural, Cambodia; MNHN 1928.0056– 1928.0057, Xiengkhuang, Laos; FMNH 180153–180154, Khao Yai, Thailand. Supplementary material 1 Detailed measurements and scale counts for each examined specimen of Hebius boulengeri Authors: Patrick David, Gernot Vogel Data type: xlsx Explanation note: All measurements are in the unit of mm. “N/A” indicates not available. For measurement and count methods and abbreviations, see the Materials and methods. Copyright notice: This dataset is made available under the Open Database License (http://opendatacommons.org/licenses/odbl/1.0/). The Open Database License (ODbL) is a license agreement intended to allow users to freely share, modify, and use this Dataset while maintaining this same freedom for others, provided that the original source and author(s) are credited. Link: https://doi.org/10.3897/zookeys.1254.161130.suppl1