Integrated evidence reveals a new subspecies of Amolops Cope, 1865 (Anura, Ranidae) from northeastern Yunnan, China

39 Integrated evidence reveals a new subspecies of Amolops Cope, 1865 (Anura, Ranidae) from northeastern Yunnan, China Pengying Li1, Jiangyu Li2,3 , Wei Zhang1, Xiangbo Yi1, Zhiyong Yuan2,3 , Junkai Huang4,Xiaolong Liu2,3 1 Management and Conservation Bureau of Yunnan Wumeng Mountain National Nature Reserve, Zhaotong 657000, China 2 Integrative Science Center of Germplasm Creation in Western China (Chongqing) Science City, Biological Science Research Center, Southwest University, Chongqing 400715, China 3 Key Laboratory of Freshwater Fish Reproduction and Development (Ministry of Education), School of Life Sciences, Southwest University, Chongqing 400715, China 4 Fuzhou Shuiyuzan Agriculture and Forestry Technology Company, Fuzhou 350100, China Corresponding authors: Junkai Huang ([email protected]); Xiaolong Liu ([email protected]) Copyright: © Pengying Li 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 subspecies of Amolops dafangensis Li, Liu, Ke, Cheng, & Wang, 2024, designated as Amolops dafangensis wumengmontis ssp. nov., is described from Mt. Wumeng, Zhaotong City, northeastern Yunnan, China, based on phylogenetic analysis of the 16S rRNA and CO1 genes, as well as morphological examinations. Phylogenetic analysis indicated that specimens from the Wumeng Mountains clustered within Amolops dafangensis, forming two main clades within this species. Amolops dafangensis wumengmontis ssp. nov. demonstrates a close phylogenetic relationship with A. d. dafangensis and is differentiated from the nominal subspecies by specific characteristics: 1) tympanum indistinct; 2) tibiotarsal articulation just reaching anterior corner of the eye; 3) flat and indistinct inner and outer metacarpal tubercles; and 4) webbing formula: I1–1II1–1½III1–2IV2–1V. Key words: Amolops dafangensis wumengmontis ssp. nov., cascade frogs, species group, subspecies, taxonomy Introduction The genus Amolops Cope, 1865, also known as cascade frogs, comprises 89 recognized species and is widely distributed across southern Asia, from the southern and eastern Himalayas to southeastern mainland China, extending southward to Peninsular Malaysia. China harbors the highest species diversity within this genus, with 60 documented species (Wu et al. 2020; Zeng et al. 2020; AmphibiaChina 2025; Frost 2025). Recent taxonomic revisions, integrating morphological and molecular phylogenetic analyses, have subdivided Amolops into eight distinct species groups: the A. mantzorum group, A. monticola group, A. hainanensis group, A. ricketti group, A. marmoratus group, A. larutensis group, A. daiyunensis group, and the A. viridimaculatus group, based on the amalgamation of morphological and molecular phylogenetic evidence (Cai et al. 2007; Lyu et al. 2019a, 2019b; Wu et al. 2020; Zeng et al. 2020; Jiang et al. 2021; Patel et al. 2021; Saikia et al. 2022a, 2022b). Academic editor: Anthony Herrel Received: 19 July 2025 Accepted: 22 September 2025 Published: 24 October 2025 ZooBank: https://zoobank. org/3E6089B6-7624-4162-971063EFCBFD3AC3 Citation: Li P, Li J, Zhang W, Yi X, Yuan Z, Huang J, Liu X (2025) Integrated evidence reveals a new subspecies of Amolops Cope, 1865 (Anura, Ranidae) from northeastern Yunnan, China. ZooKeys 1257: 39–54. https://doi. org/10.3897/zookeys.1257.165617 ZooKeys 1257: 39–54 (2025) DOI: 10.3897/zookeys.1257.165617 40 ZooKeys 1257: 39–54 (2025), DOI: 10.3897/zookeys.1257.165617 Pengying Li et al.: One new subspecies of the Amolops mantzorum Group from China Amolops mantzorum group, established by Fei et al. (1999) based on morphological characteristics, is distinguished from other species groups by a set of unique morphological characteristics, including medium body size, finger I with an expanded tip lacking a circummarginal groove, absence of vocal sac, absence of dorsolateral fold (or presence of glandular fold in some individuals), indistinct but visible tympanum, presence of a nuptial pad on finger I in males, and overlapping heels when hindlimbs are flexed at a right angle (Lyu et al. 2019b; Wu et al. 2020; Tang et al. 2023; Qian et al. 2023; Li et al. 2024). During herpetological surveys in Wumeng Mountain National Nature Reserve, Zhaotong City, northeastern Yunnan, China, a series of specimens belonging to the A. mantzorum group were collected, exhibiting similarities in appearance to A. dafangensis Li, Liu, Ke, Cheng & Wang, 2024 specimens from Guizhou Province. Subsequent molecular phylogenetic and morphological analyses revealed their close phylogenetic relationship to A. dafangensis, albeit with discernible genetic distinctions. Through integrated morphological comparisons, we propose recognizing these specimens as a new subspecies of A. dafangensis, designated as A. d. wumengmontis ssp. nov. Materials and methods Fieldwork was conducted in Wumeng Mountain National Nature Reserve, Zhaotong City, northeastern Yunnan, China (Fig. 3). Six specimens were collected in August 2022 and November 2023. Four specimens were euthanized using a low concentration of clove oil solution, and two specimens were released after toe V was clipped to minimize depletion of the wild-sampled population while ensuring individual survival. Liver and muscle tissues were extracted and preserved in 95% ethanol, and the specimens were subsequently fixed in 75% ethanol. Voucher specimens (SWU 0009561, SWU 0009562, SWU 007379, SWU 007381) were deposited at Southwest University (SWU). All sample collections were performed according to the ethical guidelines approved by the Institutional Animal Care and Use Committee (IACUC) of Southwest University (Approval No. LAC2025-2-0113). Morphology and morphometrics All the measurements were taken using digital slide calipers (accuracy: ± 0.1 mm). Morphological terminology follows Fei et al. (2009) and Lyu et al. (2019b), with detailed definitions provided in Table 1. The webbing formula adheres to the standards of Savage (1975). Complete morphological measurements for all specimens are listed in Table 2. Sex was determined by the presence/absence of nuptial pads on the finger. DNA sequencing and molecular analyses To construct a phylogeny for the A. mantzorum group, we extracted total DNA from liver or muscle tissue using the Animal Tissue DNA Isolation Kit provided by Thermo Fisher Scientific. In this study, we sequenced two mitochondrial genes: 16S rRNA and CO1. The primers used for polymerase chain reaction (PCR) amplification are detailed in Suppl. material 1. The PCR amplification 41 ZooKeys 1257: 39–54 (2025), DOI: 10.3897/zookeys.1257.165617 Pengying Li et al.: One new subspecies of the Amolops mantzorum Group from China Table 1. Morphological characters used for adult individuals. Abbreviation Morphology SVL Snout-vent length HDL Head length HDW Head width SL Snout length IND Internarial distance IOD Interorbital distance UEW Width of upper eyelid NED Nasal to eye distance NSD Nasal to snout distance ED Diameter of eye TD Diameter of tympanum HL Hand length FAHL Length of forearm and hand LAL Length of lower arm LAW Width of lower arm HLL Hindlimb length or leg length FML Femur length TFL Length of foot and tarsus FL Foot length TBL Tibia length TBW Tibia width Table 2. Measurements (mm) of adult specimens in the type series of A. d. wumengmontis ssp. nov. No. SWU 0009561 SWU 0009562 SWU 007379 SWU 007381 Sex Male Male Male (holotype) Female SVL 43.0 43.3 45.5 62.1 HDL 14.7 14.8 15.7 22.3 HDW 12.4 13.7 12.9 19.0 SL 5.9 5.7 6.7 8.4 IND 5.2 5.0 5.0 6.9 IOD 3.6 4.1 4.4 5.9 UEW 3.2 2.8 4.0 4.5 NED 2.8 1.9 2.6 4.0 NSD 3.0 3.2 3.4 5.0 ED 5.4 5.5 5.6 7.1 TD 1.6 1.9 1.5 2.3 HL 13.1 13.5 13.3 18.2 FAHL 20.8 21.3 22.3 30.1 LAL 7.7 7.8 9.0 11.9 LAW 4.7 4.7 4.8 5.3 HLL 77.0 78.0 80.7 110.3 FML 24.1 26.5 25.5 33.8 TFL 34.8 35.6 36.2 47.5 FL 22.5 22.9 22.3 32.9 TBL 25.9 26.7 26.2 33.1 TBW 5.0 4.9 5.8 7.3 42 ZooKeys 1257: 39–54 (2025), DOI: 10.3897/zookeys.1257.165617 Pengying Li et al.: One new subspecies of the Amolops mantzorum Group from China process was carried out in a 20 μL reaction volume, following the reaction cycling settings below: an initial denaturation step at 95 °C for 4 min; followed by 36 cycles of denaturation at 95 °C for 30 s, annealing at 54 °C (for 16S rRNA)/ 49 °C (for CO1) for 40 s, and extension at 72 °C for 70 s, and all sequences have been uploaded to GenBank (Table 3). Based on Tang et al. (2023), we selected species from other species groups of Amolops, Amolops chaochin Jiang, Ren, Lyu & Li, 2021 and A. ricketti (Boulenger, 1899) as outgroups to reconstruct the phylogenetic relationships of the A. mantzorum group; all GenBank accession numbers are listed in Table 3. Sequences were aligned using the MUSCLE option in MEGA v. 7 (Kumar et al. 2016). Before phylogenetic reconstruction, the best substitution model was selected using the Akaike Information Criterion (AIC) in jMODELTEST v. 2.1.10 (Darriba et al. 2012). Bayesian inference (BI) was performed using MrBayes v. 3.2.6 (Ronquist et al. 2012), with two runs performed simultaneously, each consisting of four Markov chains starting from a random tree. The chain was run for 6,000,000 generations, with sampling every 1000 generations. When the average standard deviation of the split frequency was less than 0.01, the first 25% of the sampled trees were discarded as burn-in, and the remaining trees were used to create a consensus tree and estimate the Bayesian posterior probability. Maximum likelihood (ML) analyses were performed using RAxML v. 7.0.3 (Stamatakis 2014) under the GTRGAMMA model with 1000 bootstrap replicates. We used Assemble Species by Automatic Partitioning (ASAP) for species delimitation. For this method, we used the simple distance (p-distance) model, and the partition with the lowest ASAP p-score was deemed the best (Puillandre et al. 2021). Additionally, due to the excessively small genetic distances in 16S rRNA among species within the A. mantzorum group, the ASAP could not accurately delimit species; we only selected the CO1 gene for species delimitation. Results Phylogenetic analyses The length of the sequence alignment was 1094 base pairs. Phylogenetic trees generated using BI and ML methods exhibit overall congruence (Fig. 1). In contrast, the species delimitation results based on ASAP analysis of CO1 sequences differed from the phylogenetic tree topology (Fig. 2). Mean p-distances for the 16S rRNA and CO1 genes among the Amolops species used in this study are provided in Suppl. material 2. Phylogenetic analysis indicated that specimens from the Wumeng Mountains clustered within Amolops dafangensis, forming two main clades within this species. The species delimitation results based on ASAP analysis of CO1 sequences indicate that the specimens from the Wumeng Mountains form a sister group with A. sangzhiensis Qian, Xiang, Jiang, Yang & Gui, 2023 and A. loloensis (Liu, 1950), suggesting that they should be recognized as a distinct species. The p-distance between specimens from the Wumeng Mountains and A. dafangensis is 0.5% on the 16S rRNA gene and 2.8% on the CO1 gene. 43 ZooKeys 1257: 39–54 (2025), DOI: 10.3897/zookeys.1257.165617 Pengying Li et al.: One new subspecies of the Amolops mantzorum Group from China Table 3. Amolops species used in phylogenetic analyses of this study; “/” means unknown. ID Species Voucher ID Locality GenBank accession number 16S rRNA CO1 1A. d. wumengmontis ssp. nov. SWU 0009561 Mt. Wumeng, Zhaotong, Yunnan, China PX411479 PX410826 2A. d. wumengmontis ssp. nov. SWU 0009562 Mt. Wumeng, Zhaotong, Yunnan, China PX411478 PX410827 3A. d. wumengmontis ssp. nov. SWU 007379 Yiliang, Zhaotong, Yunnan, China PX411483 PX410822 4A. d. wumengmontis ssp. nov. SWU 007381 Yiliang, Zhaotong, Yunnan, China PX411482 PX410823 5A. d. wumengmontis ssp. nov. Tissue ID: Yuan 30620 Yiliang, Zhaotong, Yunnan, China PX411481 PX410824 6A. d. wumengmontis ssp. nov. Tissue ID: Yuan 30621 Yiliang, Zhaotong, Yunnan, China PX411480 PX410825 7A. sangzhiensis CSUFT 901 Mt. Doupeng, Sangzhi, Hunan, China OQ079538 OQ078903 8A. sangzhiensis CSUFT 905 Mt. Doupeng, Sangzhi, Hunan, China OQ079539 OQ078904 9A. sangzhiensis CSUFT 907 Mt. Doupeng, Sangzhi, Hunan, China OQ079540 OQ078905 10 A. sangzhiensis CSUFT 912 Mt. Doupeng, Sangzhi, Hunan, China OQ079541 OQ078906 11 A. sangzhiensis CSUFT 916 Mt. Doupeng, Sangzhi, Hunan, China OQ079542 OQ078907 12 A. sangzhiensis CSUFT 927 Mt. Doupeng, Sangzhi, Hunan, China OQ079543 OQ078908 13 A. granulosus SYS a005399 Mt. Guangwu, Sichuan, China MK573811 MK568326 14 A. granulosus SYS a005400 Mt. Guangwu, Sichuan, China MK573812 MK568327 15 A. granulosus SYS a005318 Mt. Wawu, Sichuan, China MK573802 MK568317 16 A. granulosus SYS a005319 Mt. Wawu, Sichuan, China MK573803 MK568318 17 A. granulosus SCUM 045823HX Dayi, Sichuan, China MN953680 JN700804 18 A. jinjiangensis SYS a004571 Mt. Gaoligong, Yunnan, China MK573801 MK568316 19 A. jinjiangensis SCUM 050434CHX Deqing, Yunnan, China MN953700 MN961402 20 A. jinjiangensis SCUM 050435CHX Deqing, Yunnan, China EF453741 MN961403 21 A. lifanensis SYS a005374 Lixian, Sichuan, China MK573809 MK568324 22 A. lifanensis SYS a005375 Lixian, Sichuan, China MK573810 MK568325 23 A. lifanensis SYS a005376 Lixian, Sichuan, China MK604868 MK605626 24 A. lifanensis SYS a005377 Lixian, Sichuan, China MK604869 MK605627 25 A. lifanensis SYS a005378 Lixian, Sichuan, China MK604870 MK605628 26 A. loloensis SYS a005346 Zhaojue, Sichuan, China MK604854 MK605612 27 A. loloensis SYS a005347 Zhaojue, Sichuan, China MK604855 MK605613 28 A. loloensis SYS a005348 Zhaojue, Sichuan, China MK604856 MK605614 29 A. loloensis SYS a005349 Zhaojue, Sichuan, China MK604857 MK605615 30 A. loloensis SCUM 045806HX Xichang, Sichuan, China MN953704 MN961407 31 A. loloensis SCUM 045807HX Xichang, Sichuan, China EF453743 MN961456 32 A. mantzorum SYS a005365 Fengtongzhai, Sichuan, China MK573808 MK568323 33 A. mantzorum SYS a005366 Fengtongzhai, Sichuan, China MK604862 MK605620 34 A. mantzorum SYS a005367 Fengtongzhai, Sichuan, China MK604863 MK605621 35 A. mantzorum SYS a005368 Fengtongzhai, Sichuan, China MK604864 MK605622 36 A. mantzorum SYS a005370 Fengtongzhai, Sichuan, China MK604865 MK605623 37 A. mantzorum SYS a005371 Fengtongzhai, Sichuan, China MK604866 MK605624 38 A. mantzorum SYS a005372 Fengtongzhai, Sichuan, China MK604867 MK605625 39 A. mantzorum SYS a005356 Kangding, Sichuan, China MK604858 MK605616 40 A. mantzorum SYS a005357 Kangding, Sichuan, China MK604859 MK605617 41 A. mantzorum SYS a005358 Kangding, Sichuan, China MK604860 MK605618 44 ZooKeys 1257: 39–54 (2025), DOI: 10.3897/zookeys.1257.165617 Pengying Li et al.: One new subspecies of the Amolops mantzorum Group from China Taxonomic account Amolops dafangensis wumengmontis Huang, Yuan & Liu, ssp. nov. https://zoobank.org/AA78B643-6A58-4A8D-9298-0CC7E169572C Figs 3, 5; Table 2 Holotype. • SWU 007379, adult male (Fig. 3), collected in July 2021 by Xiaolong Liu and Rui Chen from Wumeng Mountain National Nature Reserve, Zhaotong City, northeastern Yunnan, China (27.810588°N, 104.266768°E; 1915 m a.s.l.) (Fig. 4). Paratypes. • Two adult males (SWU 0009561, SWU 0009562) were collected at Wumeng Mountain National Nature Reserve, Zhaotong City, northeastern Yunnan, China (28.213406°N, 103.935201°E; elevation 1797 m a.s.l.) in November 2023 by Xiaolong Liu and Rui Chen. • One adult female (SWU 007381) was collected at the same locality as the holotype in July 2021 by Xiaolong Liu and Rui Chen. ID Species Voucher ID Locality GenBank accession number 16S rRNA CO1 42 A. mantzorum SYS a005336 Mt. Wawu, Sichuan, China MK573804 MK568319 43 A. mantzorum SYS a005337 Mt. Wawu, Sichuan, China MK604853 MK605611 44 A. mantzorum SCUM 045817HX Wolong, Sichuan, China MN953706 MN961408 45 A. mantzorum SCUM 045825HX Dayi, Sichuan, China MN953707 MN961409 46 A. mantzorum xinduqiao KIZ 041127 Kangding, Sichuan, China MN953764 MN961465 47 A. mantzorum xinduqiao KIZ 041129 Kangding, Sichuan, China MN953765 MN961466 48 A. minutus KIZ 2023068 Yuanyang, Yunnan, China PQ346031 / 49 A. minutus KIZ 2023069 Yuanyang, Yunnan, China PQ346032 / 50 A. minutus KIZ 2023070 Yuanyang, Yunnan, China PQ346033 / 51 A. shuichengicus SYS a004956 Shuicheng, Guizhou, China MK604845 MK605603 52 A. shuichengicus SYS a004957 Shuicheng, Guizhou, China MK604846 MK605604 53 A. shuichengicus SYS a004958 Shuicheng, Guizhou, China MK604847 MK605605 54 A. shuichengicus SYS a004971 Shuicheng, Guizhou, China MK604848 MK605606 55 A. tuberodepressus SCUM 050430CHX Jingdong, Yunnan, China MN953730 MN961433 56 A. tuberodepressus SCUM 050433CHX Jingdong, Yunnan, China MN953729 MN961432 57 A. tuberodepressus SYS a003900 Mt. Ailao, Yunnan, China MK573797 MK568314 58 A. tuberodepressus SYS a003901 Mt. Ailao, Yunnan, China MK573798 MK568315 59 A. tuberodepressus SYS a003902 Mt. Ailao, Yunnan, China MK604844 MK605602 60 A. tuberodepressus SYS a003931 Mt. Wuliang, Yunnan, China MK573799 MG991933 61 A. ailao GXNU YU000001 Mt. Ailao, Yunnan, China MN650751 MN650737 62 A. ailao GXNU YU000002 Mt. Ailao, Yunnan, China MN650752 MN650738 63 A. ailao GXNU YU000003 Mt. Ailao, Yunnan, China MN650753 MN650739 64 A. ailao GXNU YU000004 Mt. Ailao, Yunnan, China MN650754 MN650740 65 A. d. dafangensis MT DF20230601002 Dafang, Guizhou, China OR936315 OR924345 66 A. d. dafangensis MT DF20230601003 Dafang, Guizhou, China OR936316 OR924346 67 A. d. dafangensis MT DF20230601004 Dafang, Guizhou, China OR936317 OR924347 68 A. chaochin XM5526 Wenxian, Gansu, China KX645666 KX645666 69 A. ricketti SYS a004141 Mt. Wuyi, Fujian, China MK263259 MG991927 45 ZooKeys 1257: 39–54 (2025), DOI: 10.3897/zookeys.1257.165617 Pengying Li et al.: One new subspecies of the Amolops mantzorum Group from China Figure 1. Bayesian phylogram of Amolops mantzorum group inferred from the combination of 16S rRNA and CO1 genes. The numbers above and below the branches are Bayesian posterior probabilities (BPP) and maximum likelihood bootstrap values (BS); “-” denotes a BPP < 0.95 and BS < 70. Figure 2. ASAP species delimitation based on CO1 sequences. The best partition with the lowest score is highlighted with a red frame. 46 ZooKeys 1257: 39–54 (2025), DOI: 10.3897/zookeys.1257.165617 Pengying Li et al.: One new subspecies of the Amolops mantzorum Group from China Etymology. The specific epithet “wumengmontis” is derived from the type locality, Wumeng Mountain National Nature Reserve, Zhaotong City, northeastern Yunnan, China. We suggest ‘Wumeng Cascade Frog’ as its English common name, and ‘Dà Fāng Tuān Wā Wū Mēng Yà Zhŏng’ (大方湍蛙乌蒙亚种) as its Chinese common name. Diagnosis. Amolops dafangensis wumengmontis ssp. nov. was classified within the genus Amolops and further assigned to the A. mantzorum group due to the combination of following features: 1) moderate body size, with a SVL of 43.0–45.5 mm in adult males (N = 3) and 62.1 mm in adult female (N = 1); 2) presence of vomerine teeth; 3) slightly longer head length compared to head width; 4) indistinct tympanum; 5) lack of webbing or lateral fringes on fingers, as well as absence of lateral fringes on toes; 6) maxillary glands extremely indistinct, appearing as three small clusters; 7) presence of supratympanic folds and dorsolateral folds formed by a series of glands; 8) tibiotarsal articulation reaching the anterior corner of the eye; 9) heels overlapping when hindlimbs flexed at right angles to the body axis; and 10) absence of vocal sacs in males, with nuptial pads featuring velvety white nuptial spines at the base of finger I. Description of holotype. Adult male, body size moderate (SVL = 45.5 mm) with a head length slightly surpassing head width (HDL/HDW = 1.2). The snout is moderately rounded, the canthus rostralis is well-defined and curved, while the loreal region slopes concavely. The nostrils are circular, positioned between the snout and eyes, and slightly protruding. The eyes are large (ED/SL = 0.8), the internarial distance exceeds the interorbital distance (IND/IOD = 1.1), and the width of the upper eyelid is slightly narrower than the interorbital distance (UEW/IOD = 0.9). A discernible pineal spot is present, the pupil is round and Figure 3. Holotype SWU 007379 of A. d. wumengmontis ssp. nov. in situ. 47 ZooKeys 1257: 39–54 (2025), DOI: 10.3897/zookeys.1257.165617 Pengying Li et al.: One new subspecies of the Amolops mantzorum Group from China horizontally oriented, and the mouth corner is smooth. The tympanum is indistinct, with maxillary glands appearing extremely indistinct, resembling three small clusters. There is an indistinct supratympanic fold, vomerine teeth were observed, choanae are round, the tongue is anteriorly attached and cordiform, with a notched posterior margin, and a vocal sac is notably absent. The forelimbs are robust, with the relative length order of fingers being I < II < IV < III; all fingers are expanded into discs, with circummarginal grooves present on all except finger I. There is no webbing or lateral fringes on the fingers. Nuptial pads featuring velvety white nuptial spines are located at the base of finger I. Webbing between the fingers is absent. The subarticular tubercles are distinct and rounded, supernumerary tubercles are present, and both the inner and outer metacarpal tubercles are flat and indistinct. The hindlimbs are elongated, nearly twice the snout-vent length (HLL/SVL = 1.8), with the tibiotarsal articulation reaching the anterior corner of the eye. When the hindlimbs are flexed at right angles to the body axis, the heels overlap. The tibia length exceeds the combined lengths of the forearm and hand (TBL/FAHL = 1.2). The relative length order of the toes is I < II < III < V < IV, with Figure 4. Distribution localities of A. d. wumengmontis ssp. nov. in Wumeng Mountains, northeastern Yunnan, China. 54 ZooKeys 1257: 39–54 (2025), DOI: 10.3897/zookeys.1257.165617 Pengying Li et al.: One new subspecies of the Amolops mantzorum Group from China Savage JM (1975) Systematics and distribution of the Mexican and Central American stream frogs related to Eleutherodactylus rugulosus. Copeia 2(2): 254–306. https:// doi.org/10.2307/1442883 Stamatakis A (2014) RAxML version 8: A tool for phylogenetic analysis and post-analysis of large phylogenies. Bioinformatics (Oxford, England) 30(9): 1312–1313. https:// doi.org/10.1093/bioinformatics/btu033 Tang SJ, Sun T, Liu S, Luo SD, Yu GH, Du LN (2023) A new species of cascade frog (Anura: Ranidae: Amolops) from central Yunnan, China. Zoological Letters 9(15): 1–19. https://doi.org/10.1186/s40851-023-00214-9 Wu YH, Yan F, Stuart BL, Prendini E, Suwannapoom C, Dahn HA, Zhang BL, Cai HX, Xu YB, Jiang K, Chen HM, Lemmon EM, Raxworthy CJ, Orlov NL, Murphy RW, Che J (2020) 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). Molecular Phylogenetics and Evolution 144(106701): 106789. https://doi.org/10.1016/j.ympev.2020.106789 Zeng ZC, Liang D, Li JX, Lyu ZT, Wang YY, Zhang P (2020) Phylogenetic relationships of the Chinese torrent frogs (Ranidae: Amolops) revealed by phylogenomic analyses of AFLP-Capture data. Molecular Phylogenetics and Evolution 146: 106753. https://doi. org/10.1016/j.ympev.2020.106753 Supplementary material 1 Primer pairs for PCR amplification used in this study Authors: Pengying Li, Jiangyu Li, Wei Zhang, Xiangbo Yi, Zhiyong Yuan, Junkai Huang, Xiaolong Liu Data type: xlsx 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.1257.165617.suppl1 Supplementary material 2 Mean uncorrected pairwise distances of 16S rRNA and CO1 (%) between clades of A. mantzorum group Authors: Pengying Li, Jiangyu Li, Wei Zhang, Xiangbo Yi, Zhiyong Yuan, Junkai Huang, Xiaolong Li Data type: xlsx 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.1257.165617.suppl2