Research Article |
Corresponding author: Bin Wang ( wangbin@cib.ac.cn ) Corresponding author: Jun Wu ( wujun@nies.org ) Academic editor: Anthony Herrel
© 2020 Bin Wang, Yan-Qing Wu, Jun-Wei Peng, Sheng-Chao Shi, Ning-Ning Lu, Jun Wu.
This is an open access article distributed under the terms of the Creative Commons Attribution License (CC BY 4.0), which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited.
Citation:
Wang B, Wu Y-Q, Peng J-W, Shi S-C, Lu N-N, Wu J (2020) A new Megophrys Kuhl & Van Hasselt (Amphibia, Megophryidae) from southeastern China. ZooKeys 904: 35-62. https://doi.org/10.3897/zookeys.904.47354
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A new species of the genus Megophrys from Zhejiang Province, China is described. Molecular phylogenetic analyses supported the new taxon as an independent clade nested into the Megophrys clade and sister to M. lishuiensis. The new species could be distinguished from its congeners by a combination of the following morphological characteristics: (1) small size (SVL 31.0–36.3 mm in male and 41.6 mm in female); (2) vomerine ridge present and vomerine teeth absent; (3) tongue not notched behind; (4) a small horn-like tubercle at the edge of each upper eyelid; (5) tympanum distinctly visible, rounded; (6) two metacarpal tubercles in hand; (7) relative finger lengths: II < I < IV < III; (8) toes with rudimentary webbing at bases; (9) heels overlapping when thighs are positioned at right angles to the body; (10) tibiotarsal articulation reaching tympanum to eye when leg stretched forward; (11) an internal single subgular vocal sac in male; (12) in breeding male, the nuptial pads with black nuptial spines on the dorsal bases of the first and second fingers.
Taxonomy, new species, molecular phylogenetic analysis, morphology, Zhejiang Province, China
Megophryidae Bonaparte, 1850 (Amphibia: Anura) is a large Asian toad family which was divided into three subfamilies, i.e., Leptobrachiinae Dubois, 1983, Leptolalaginae Delorme, Dubois, Grosjean & Ohler, 2006 and Megophryinae Bonaparte, 1850 (
The genus Megophrys is widely distributed from eastern China through the eastern and southern Himalayas, throughout mainland Indochina and the islands of the Sunda shelf in Indonesia and parts of the Philippines (
During our field surveys in the Xianju County, Zhejiang Province, China, some Megophrys specimens were collected from the montane forests. Molecular phylogenetic analyses and morphological comparisons support that it is distinctly different from its Megophrys congeners. Therefore, we describe it herein as a new species.
Seven adult males, two adult females, and six tadpoles of the new taxon (for voucher information see Suppl. material
Localities for specimens used in this study. 1, the type locality of Megophrys xianjuensis sp. nov., Xianju County, Zhejiang Province, China; 2. the common type locality of M. boettgeri (black) and M. kuatunensis (purple), Wuyi Mountain, Nanping County, Fujian Province, China; 3, the type locality of M. lishuiensis, Lishui City, Zhejiang Province, China.
Four male, two female, and three tadpole specimens of the new taxon were included in the molecular analyses (for voucher information see Table
Localities, voucher information, and GenBank accession numbers for molecular samples used in this study.
Species | Voucher number | Locality | GenBank accession number | ||
---|---|---|---|---|---|
16S | COI | ||||
Megophrys aceras | KIZ025467 | Khao Nan National Park, Nakhon Si Thammarat, Thailand | KX811925 | KX812159 | |
M. acuta | SYS a001957 | Heishiding Nature Reserve, Guangdong Prov., China | KJ579118 | – | |
M. auralensis | NCSM 79599 | Aural, Kampong Speu, Cambodia | KX811807 | – | |
M. baluensis | ZMH A13125 | Gunung Kinabalu National Park, Kogopan Trail, Malaysia | KJ831310 | – | |
M. baolongensis | KIZ019216 | Baolong, Chongqing City, China | KX811813 | KX812093 | |
M. binchuanensis | KIZ019441 | Mt. Jizu, Yunnan Prov., China | KX811849 | KX812112 | |
M. binlingensis | KIZ025807 | Mt. Wawu, Sichuan Prov., China | KX811852 | KX812115 | |
M. boettgeri | Tissue ID: YPXJK033 | Mt. Wuyi, Fujian Prov., China | KX811814 | KX812104 | |
CIBWY18082307 | Mt. Wuyi, Fujian Prov., China | MN563762 | MN563778 | ||
CIBWY18082603 | Mt. Wuyi, Fujian Prov., China | MN563763 | MN563779 | ||
M. brachykolos | ROM 16634 | Hong Kong, China | KX811897 | KX812150 | |
M. carinense | Tissue ID: YPX20455 | Dayao Shan, Guangxi Prov., China | KX811811 | KX812057 | |
M. cheni | SYS a001427 | Jinggang Shan, Jiangxi Prov., China | KJ560391 | – | |
M. chuannanensis | CIB20050081 | Hejiang, Sichuan Prov., China | KM504261 | – | |
M. daweimontis | KIZ048997 | Dawei Shan, Yunnan Prov., China | KX811867 | KX812125 | |
M. dongguanensis | SYS a001972 | Mt. Yinping, Guangdong Prov., China | MK524098 | MK524129 | |
M. dringi | UNIMAS 8943 | Gunung Mulu National Park, Sarawak, Malaysia | KJ831317 | – | |
M. edwardinae | FMNH 273694 | Bintulu, Sarawak, Malaysia | KX811918 | KX812050 | |
M. elfina | ZMMU ABV-00454 | Bidoup Mountain, Lam Dong, Vietnam | KY425379 | – | |
M. fansipanensis | VNMN 2018.01 | Lao Cai, Sa Pa, Vietnam | MH514886 | – | |
M. feae | KIZ046706 | Huangcaoling, Yunnan Prov., China | KX811810 | KX812056 | |
M. flavipunctata | SDBDU2009.297 | East Khasi Hills dist., Meghalaya | KY022307 | MH647536 | |
M. gerti | ITBCZ 1108 | Nui Chua National Park, Ninh Thuan, Vietnam | KX811917 | KX812161 | |
M. gigantica | SYSa003933 | Wuliang shan, Yunnan Prov., China | MH406775 | MH406235 | |
M. glandulosa | KIZ048439 | Husa, Yunnan Prov., China | KX811762 | KX812075 | |
M. hansi | KIZ010360 | Phong Dien Nature Reserve, Thua Thien Hue, Vietnam | KX811913 | KX812155 | |
M. himalayana | SDBDU2009.75 | East Siang dist., Arunachal Pradesh, India | KY022311 | – | |
M. hoanglienensis | VNMN 2018.02 | Lao Cai, Sa Pa, Vietnam | MH514889 | – | |
M. huangshanensis | KIZ022004 | Mt. Huang, Anhui Prov., China | KX811821 | KX812107 | |
M. intermedia | ZFMK 87596 | U Bo, Phong Nha-Ke Bang NP, Vietnam | HQ588950 | – | |
M. jingdongensis | KIZ-LC0805067 | Huanglianshan, Yunnan Prov., China | KX811872 | KX812131 | |
M. jinggangensis | KIZ07132 | Chashan Forest Farm, Jiangxi Prov., China | KX811840 | KX812108 | |
M. jiulianensis | SYS a002107 | Mt. Jiulian, Jiangxi Prov., China | MK524099 | MK524130 | |
M. kalimantanensis | MZB. Amph 21482 | Tanah Bumbu, Kalimantan Selatan, Borneo, Indonesia | MG993554 | – | |
M. kobayashii | UNIMAS 8148 | Gunung Kinabalu National Park, Sabah, Malaysia | KJ831313 | – | |
M. kuatunensis | CIBWY18082407 | Mt. Wuyi, Fujian Prov., China | MN563764 | MN563780 | |
CIBWY18082408 | Mt. Wuyi, Fujian Prov., China | MN563765 | MN563781 | ||
SYS a001579 | Mt. Wuyi, Fujian Prov., China | KJ560376 | – | ||
M. lancip | MZB:Amp:22233 | – | KY679891 | – | |
M. leishanensis | CIBLS20171101001 | Mt. Leigong, Guizhou Prov., China | MK005310 | MK005306 | |
M. ligayae | ZMMU NAP-05015 | Palawan, Philippines | KX811919 | KX812051 | |
M. lini | SYS a002370 | Suichuan Co., Jiangxi Prov., China | KJ560412 | – | |
M. lishuiensis | CIBWYF00169 | Lishui City, Zhejiang Prov., China | KY021418 | – | |
CIBWYF00170 | Lishui City, Zhejiang Prov., China | KY113084 | – | ||
CIBWYF11011 | Lishui City, Zhejiang Prov., China | KY113085 | – | ||
M. major | SYSa002961 | Zhushihe, Yunnan Prov., China | MH406728 | MH406180 | |
M. mangshanensis | KIZ021786 | Nanling National Forest Park, Guangdong Prov., China | KX811790 | KX812079 | |
M. maosonensis | KIZ016045 | Xiaoqiaogou Nature Reserve, Yunnan Prov., China | KX811780 | KX812080 | |
M. medogensis | KIZ06621 | Beibeng, Xizang Prov., China | KX811767 | KX812082 | |
M. microstoma | KIZ048799 | Xiaoqiaogou Nature Reserve, Yunnan Prov., China | KX811914 | KX812156 | |
M. minor | KIZ01939 | Qingcheng Shan, Sichuan Prov., China | KX811896 | KX812145 | |
M. montana | LSUMZ 81916 | Sukabumi, Java, Indonesia | KX811927 | KX812163 | |
M. monticola | SDBDU 2011.1047 [h] | Darjeeling dist., West Bengal, India | KX894679 | – | |
M. mufumontana | SYS a006391 | Mt. Mufu, Hunan Prov., China | MK524105 | MK524136 | |
M. nankiangensis | CIB ZYC517 | Nanjiang, Sichuan Prov., China | KX811900 | – | |
M. nankunensis | SYS a004498 | Mt. Nankun, Guangdong Prov., China | MK524108 | MK524139 | |
M. nanlingensis | SYS a001959 | Nanling Nature Reserve, Guangdong Prov., China | MK524111 | MK524142 | |
M. nasuta | KIZ019419 | Malaysia | KX811921 | KX812054 | |
M. obesa | SYS a002272 | Heishiding Nature Reserve, Guangdong Prov., China | KJ579122 | – | |
M. ombrophila | KRM18 | Mt. Wuyi, Fujian Prov., China | KX856404 | – | |
M. omeimontis | KIZ025765 | Mt. E’mei, Sichuan Prov., China | KX811884 | KX812136 | |
M. oreocrypta | BNHS 6046 | West Garo Hills dist., Meghalaya | KY022306 | – | |
M. pachyproctus | KIZ010978 | Beibeng, Xizang Prov., China | KX811908 | KX812153 | |
M. palpebralespinosa | KIZ011603 | Pu Hu Nature Reserve, Thanh Hoa, Vietnam | KX811888 | KX812137 | |
M. parva | SYSa003042 | Zhushihe, Yunnan Prov., China | MH406737 | MH406189 | |
M. periosa | BNHS 6061 | West Kameng dist., Arunachal Pradesh, India | KY022309 | MH647528 | |
M. popei | SYS a000589 | Naling Nature Reserve, Guangdong Prov., China | KM504251 | – | |
M. sangzhiensis | Tissue ID: YPX11006 | Badagongshan Nature Reserve, Hunan Prov., China | KX811856 | KX812117 | |
M. shapingensis | KIZ014512 | Liziping Nature Reserve, Sichuan Prov., China | KX811904 | KX812060 | |
M. spinata | KIZ016100 | Mt. Leigong, Guizhou Prov., China | KX811864 | KX812119 | |
M. stejnegeri | KU 314303 | Pasonanca Natural Park, Zamboanga, Philippines | KX811922 | KX812052 | |
M. synoria | FMNH 262778 | O’Reang, Mondolkiri, Cambodia | KY022198 | – | |
M. tuberogranulata | Tissue ID: YPX10987 | Badagongshan Nature Reserve, Hunan Prov., China | KX811823 | KX812095 | |
M. wawuensis | KIZ025799 | Wawu Shan, Sichuan Prov., China | KX811902 | KX812062 | |
M. wugongensis | SYS a002610 | Wugongshan Scenic Area, Jiangxi Prov., China | MK524114 | MK524145 | |
M. wuliangshanensis | KIZ046812 | Huangcaoling, Yunnan Prov., China | KX811881 | KX812129 | |
M. wushanensis | KIZ045469 | Guangwu Mountain, Sichuan Prov., China | KX811838 | KX812094 | |
M. xianjuensis sp. nov. | CIBXJ190505 | Xianju Co., Zhejiang Prov., China | MN563753 | MN563769 | |
CIBXJ20190801 | Xianju Co., Zhejiang Prov., China | MN563754 | MN563770 | ||
CIBXJ20190802 | Xianju Co., Zhejiang Prov., China | MN563755 | MN563771 | ||
CIBXJ20190803 | Xianju Co., Zhejiang Prov., China | MN563756 | MN563772 | ||
CIB20180514008 | Xianju Co., Zhejiang Prov., China | MN563757 | MN563773 | ||
CIBXJ190503 | Xianju Co., Zhejiang Prov., China | MN563758 | MN563774 | ||
CIBXJT19050702 | Xianju Co., Zhejiang Prov., China | MN563759 | MN563775 | ||
CIBXJT19050703 | Xianju Co., Zhejiang Prov., China | MN563760 | MN563776 | ||
CIBXJT19050704 | Xianju Co., Zhejiang Prov., China | MN563761 | MN563777 | ||
M. zhangi | KIZ014278 | Zhangmu, Xizang Prov., China | KX811765 | KX812084 | |
Leptobrachium boringii | Tissue ID: YPX37539 | Emei Shan, Sichuan Prov., China | KX811930 | KX812164 | |
L. oshanensis | KIZ025778 | Emei Shan, Sichuan Prov., China | KX811928 | KX812166 |
Total DNA was extracted using a standard phenol-chloroform extraction protocol (
For molecular analyses, the available sequence data for all related species of the genus Megophrys were downloaded from GenBank, mainly from previous studies (
Sequences were assembled and aligned using the Clustalw module in BioEdit v. 7.0.9.0 (
Phylogenetic trees were reconstructed for the concatenated data of the mitochondrial genes. Phylogenetic analyses were conducted using maximum likelihood (ML) and Bayesian Inference (BI) methods, implemented in PhyML v. 3.0 (
Nine adult specimens of the new taxon and the holotype and seven paratypes of M. lishuiensis were measured (for voucher information see Suppl. material
ED eye diameter (distance from the anterior corner to the posterior corner of the eye);
FIIIL third finger length (distance from base to tip of finger III);
FIIL second finger length (distance from base to tip of finger II);
FIL first finger length (distance from base to tip of finger I);
FIVL fourth finger length (distance from base to tip of finger IV);
FL foot length (distance from tarsus to the tip of fourth toe);
HAL hand length (distance from the posterior end of the inner metacarpal tubercle to the distal tip of finger III);
HDL head length (distance from the tip of the snout to the articulation of jaw);
HDW maximum head width (greatest width between the left and right articulations of jaw);
IND internasal distance (minimum distance between the inner margins of the external nares);
IOD interorbital distance (minimum distance between the inner edges of the upper eyelids);
LAL length of lower arm and hand (distance from the elbow to the distal end of the finger III);
LW lower arm width (maximum width of the lower arm);
SL snout length (distance from the tip of the snout to the anterior corner of the eye);
SNT distance from the tip of the snout to the naris;
SVL snout-vent length (distance from the tip of the snout to the posterior edge of the vent);
TFL length of foot and tarsus (distance from the tibiotarsal articulation to the distal end of the Toe IV);
THL thigh length (distance from vent to knee);
TL tibia length (distance from knee to tarsus);
TW maximal tibia width;
TYD maximal tympanum diameter;
UEW upper eyelid width (greatest width of the upper eyelid margins measured perpendicular to the anterior-posterior axis).
For seven tadpoles of the new taxon (for voucher information see Suppl. material
BH maximum body height;
BW maximum body width;
IOD interocular distance (minimum distance between eye);
MW mouth width (distance between two corners of mouth);
SL snout length (distance from the tip of the snout to the anterior corner of the eye);
SS snout to spiraculum (distance from spiraculum to the tip of the snout);
SVL snout-vent length;
TAH tail height (maximum height between upper and lower edges of tail);
TAL tail length (distance from base of vent to the tip of tail);
TBW maximum width of tail base;
TOL total length (distance from the tip of the snout to the tip of tail).
In order to reduce the impact of allometry, a size-corrected value from the ratio of each character to SVL was calculated and then log-transformed for the following morphometric analyses. One-way analysis of variance (ANOVA) was used to test the significance of differences on morphometric characters between different sexes as well as between different species. The significance level was set at 0.05. Furthermore, to show the spatial distribution of each species on the morphometric characters, principal component analyses (PCA) were performed. These analyses were carried out in R (
We compared morphological characters of the new taxon with other Megophrys species. Comparative data were obtained from the literature for 92 species of the genus (Table
References for morphological characters for congeners of the genus Megophrys.
Species | Literature obtained |
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M. aceras Boulenger, 1903 |
|
M. acuta Wang, Li & Jin, 2014 |
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M. ancrae Mahony, Teeling & Biju, 2013 |
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M. auralensis Ohler, Swan & Daltry, 2002 |
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M. baluensis (Boulenger, 1899) |
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M. baolongensis Ye, Fei & Xie, 2007 |
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M. binchuanensis Ye & Fei, 1995 |
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M. binlingensis Jiang, Fei & Ye, 2009 |
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M. boettgeri (Boulenger, 1899) |
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M. brachykolos Inger & Romer, 1961 |
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M. carinense (Boulenger, 1889) |
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M. caudoprocta Shen, 1994 |
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M. cheni (Wang & Liu, 2014) |
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M. chuannanensis (Fei, Ye & Huang, 2001) |
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M. damrei Mahony, 2011 |
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M. daweimontis Rao & Yang, 1997 |
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M. dongguanensis Wang & Wang, 2019 |
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M. dringi Inger, Stuebing & Tan, 1995 |
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M. edwardinae Inger, 1989 |
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M. elfina Poyarkov, Duong, Orlov, Gogoleva, Vassilieva, Nguyen, Nguyen, Nguyen, Che & Mahony, 2017 |
|
M. fansipanensis Tapley, Cutajar, Mahony, Nguyen, Dau, Luong, Le, Nguyen, Nguyen, Portway, Luong & Rowley, 2018 |
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M. feae Boulenger, 1887 |
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M. feii Yang, Wang & Wang, 2018 |
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M. flavipunctata Mahony, Kamei, Teeling & Biju, 2018 |
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M. gerti (Ohler, 2003) |
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M. gigantica Liu, Hu & Yang, 1960 |
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M. glandulosa Fei, Ye & Huang, 1990 |
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M. hansi (Ohler, 2003) |
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M. himalayana Mahony, Kamei, Teeling & Biju, 2018 |
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M. hoanglienensis Tapley, Cutajar, Mahony, Nguyen, Dau, Luong, Le, Nguyen, Nguyen, Portway, Luong & Rowley, 2018 |
|
M. huangshanensis Fei & Ye, 2005 |
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M. insularis (Wang, Liu, Lyu, Zeng & Wang, 2017) |
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M. intermedia Smith, 1921 |
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M. jingdongensis Fei & Ye, 1983 |
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M. jinggangensis (Wang, 2012) |
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M. jiulianensis Wang, Zeng, Lyu & Wang, 2019 |
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M. kalimantanensis Munir, Hamidy, Matsui, Iskandar, Sidik & Shimada, 2019 |
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M. kobayashii Malkmus & Matsui, 1997 |
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M. koui Mahony, Foley, Biju & Teeling, 2017 |
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M. kuatunensis Pope, 1929 |
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M. lancip Munir, Hamidy, Farajallah & Smith, 2018 |
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M. leishanensis Li, Xu, Liu, Jiang, Wei & Wang, 2018 |
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M. lekaguli Stuart, Chuaynkern, Chan-ard & Inger, 2006 |
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M. liboensis (Zhang, Li, Xiao, Li, Pan, Wang, Zhang & Zhou, 2017) |
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M. ligayae Taylor, 1920 |
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M. lini (Wang & Yang, 2014) |
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M. lishuiensis (Wang, Liu & Jiang, 2017) |
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M. longipes Boulenger, 1886 |
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M. major Boulenger, 1908 |
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M. mangshanensis Fei & Ye, 1990 |
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M. maosonensis Bourret, 1937 |
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M. medogensis Fei, Ye & Huang, 1983 |
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M. megacephala Mahony, Sengupta, Kamei & Biju, 2011 |
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M. microstoma (Boulenger, 1903) |
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M. minor Stejneger, 1926 |
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M. montana Kuhl & Van Hasselt, 1822 | Kuhl & Van Hasselt 1822 |
M. monticola (Günther, 1864) |
|
M. mufumontana Wang, Lyu & Wang, 2019 |
|
M. nankiangensis Liu & Hu, 1966 |
|
M. nankunensis Wang, Zeng &. Wang, 2019 |
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M. nanlingensis Lyu, Wang, Liu & Wang, 2019 |
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M. nasuta (Schlegel, 1858) |
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M. obesa Wang, Li & Zhao, 2014 |
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M. ombrophila Messenger & Dahn, 2019 |
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M. omeimontis Liu, 1950 |
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M. oreocrypta Mahony, Kamei, Teeling & Biju, 2018 |
|
M. oropedion Mahony, Teeling & Biju, 2013 |
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M. pachyproctus Huang, 1981 |
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M. palpebralespinosa Bourret, 1937 |
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M. parallela Inger & Iskandar, 2005 |
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M. parva (Boulenger, 1893) |
|
M. periosa Mahony, Kamei, Teeling & Biju, 2018 |
|
M. popei (Zhao, Yang, Chen, Chen & Wang, 2014) |
|
M. robusta Boulenger, 1908 |
|
M. rubrimera Tapley, Cutajar, Mahony, Chung, Dau, Nguyen, Luong & Rowley, 2017 |
|
M. sangzhiensis Jiang, Ye & Fei, 2008 |
|
M. serchhipii (Mathew & Sen, 2007) |
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M. shapingensis Liu, 1950 |
|
M. shuichengensis Tian & Sun, 1995 |
|
M. shunhuangensis Wang, Deng, Liu, Wu & Liu, 2019 |
|
M. spinata Liu & Hu, 1973 |
|
M. stejnegeri Taylor, 1920 |
|
M. synoria (Stuart, Sok & Neang, 2006) |
|
M. takensis Mahony, 2011 |
|
M. tuberogranulata Shen, Mo & Li, 2010 |
|
M. vegrandis Mahony, Teeling, Biju, 2013 |
|
M. wawuensis Fei, Jiang & Zheng, 2001 |
|
M. wugongensis Wang, Lyu & Wang, 2019 |
|
M. wuliangshanensis Ye & Fei, 1995 |
|
M. wushanensis Ye & Fei, 1995 |
|
M. zhangi Ye & Fei, 1992 |
|
M. zunhebotoensis (Mathew & Sen, 2007) |
|
The advertisement calls of the new taxon from Xianju County, Zhejiang Province, China were recorded in the field. SONY PCM-D50 digital sound recorder was used to record within 20 cm of the calling individuals. The sound files in wave format were resampled at 48 kHz with sampling depth 24 bits. The sonograms and waveforms were generated by WaveSurfer software (
Aligned sequence matrix of 16S+COI contained 1104 bps. ML and BI trees of the mitochondrial DNA dataset presented almost consistent topology, though relationships of some lineages were unresolved (Fig.
The results of one-way ANOVA showed that in the new taxon, the male group was significantly different from the female group on SVL and the ratio of ED and IND to SVL (p-value < 0.05). Therefore, morphometric analyses between the new taxon and M. lishuiensis were conducted separately for male and female. In PCA, the total variation of the first two principal components was 56.2% in male and 74.9% in female respectively. In both male and female, the new taxon could be distinctly separated from its phylogenetically sister species M. lishuiensis on the two-dimensional plots of PC1 vs. PC2 (Fig.
Morphometric comparisons between the adult specimens of Megophrys xianjuensis sp. nov. and M. lishuiensis. Units are in mm. See abbreviations for the morphological characters in Materials and methods section.
Character | Megophrys xianjuensis sp. nov. (MX) | M. lishuiensis (ML) | P-value from ANOVA | ||||||||
Males (N = 7) | Female (N = 2) | Males (N = 6) | Female (N = 2) | ||||||||
Range | Mean ± SD | Range | Mean ± SD | Range | Mean ± SD | Range | Mean ± SD | Male vs. female in MX | MX vs. ML in male | MX vs. ML in female | |
SVL | 31.0–36.3 | 33.1 ± 1.8 | 32.8–41.6 | 37.2 ± 6.2 | 30.5–34.7 | 32.5 ± 1.6 | 37.6–40.4 | 39.0 ± 1.9 | 0.119 | 0.633 | 0.728 |
HDL | 9.4–10.5 | 9.9 ± 0.4 | 10.1–10.8 | 10.4 ± 0.5 | 9.4–10.3 | 9.8 ± 0.4 | 9.9–11.4 | 10.6 ± 1.1 | 0.365 | 0.916 | 0.705 |
HDW | 10.6–11.5 | 11.1 ± 0.3 | 10.7–12.9 | 11.8 ± 1.5 | 10.6–11.6 | 10.9 ± 0.3 | 12.1–12.1 | 12.1 ± 0.02 | 0.361 | 0.813 | 0.561 |
SL | 3.8–4.7 | 4.2 ± 0.3 | 3.6–5.2 | 4.4 ± 1.1 | 4.1–4.5 | 4.3 ± 0.2 | 4.3–4.4 | 4.3 ± 0.1 | 0.318 | 0.569 | 0.510 |
SNT | 1.9–2.5 | 2.2 ± 0.2 | 1.8–2.5 | 2.1 ± 0.5 | 2.0–2.5 | 2.3 ± 0.2 | 2.4–2.7 | 2.6 ± 0.2 | 0.165 | 0.177 | 0.177 |
IND | 3.2–3.7 | 3.4 ± 0.2 | 3.2–3.6 | 3.4 ± 0.3 | 3.1–4.0 | 3.5 ± 0.4 | 4.2–4.2 | 4.2 ± 0.01 | 0.045* | 0.470 | 0.121 |
IOD | 2.3–3.3 | 2.8 ± 0.4 | 2.8–2.9 | 2.8 ± 0.1 | 2.4–2.8 | 2.5 ± 0.1 | 2.8–3.1 | 2.9 ± 0.2 | 0.345 | 0.140 | 0.959 |
UEW | 2.3–2.9 | 2.7 ± 0.2 | 2.7–3.4 | 3.1 ± 0.5 | 2.6–3.6 | 3.2 ± 0.3 | 3.7–3.7 | 3.7 ± 0.02 | 0.807 | 0.004** | 0.047* |
ED | 3.3–4.0 | 3.6 ± 0.3 | 2.9–4.0 | 3.4 ± 0.8 | 3.8–4.6 | 4.1 ± 0.2 | 4.1–4.6 | 4.3 ± 0.3 | 0.002** | 0.002** | 0.046* |
TYD | 1.8–2.2 | 2.0 ± 0.1 | 1.7–2.7 | 2.2 ± 0.6 | 1.9–2.5 | 2.1 ± 0.2 | 2.2–2.3 | 2.3 ± 0.1 | 0.571 | 0.044* | 0.962 |
LAL | 14.2–17.2 | 15.3 ± 1.0 | 13.7–19.2 | 16.5 ± 3.9 | 14.6–15.7 | 15.2 ± 0.4 | 17.5–17.6 | 17.5 ± 0.04 | 0.279 | 0.792 | 0.767 |
HAL | 7.6–9.2 | 8.2 ± 0.6 | 7.8–10.0 | 8.9 ± 1.5 | 8.0–8.8 | 8.4 ± 0.3 | 8.7–9.3 | 9.0 ± 0.4 | 0.489 | 0.220 | 0.021* |
LW | 2.1–2.7 | 2.5 ± 0.2 | 2.0–2.6 | 2.3 ± 0.4 | 2.4–3.1 | 2.8 ± 0.2 | 2.4–2.4 | 2.4 ± 0.01 | 0.062 | 0.042* | 0.879 |
FIL | 2.5–3.1 | 2.9 ± 0.2 | 2.8–3.9 | 3.4 ± 0.78 | 2.4–3.0 | 2.7 ± 0.2 | 2.8–3.6 | 3.2 ± 0.5 | 0.488 | 0.644 | 0.602 |
FIIL | 2.7–3.5 | 3.0 ± 0.3 | 3.0–4.0 | 3.5 ± 0.78 | 2.5–3.3 | 2.9 ± 0.3 | 3.0–3.2 | 3.1 ± 0.2 | 0.675 | 0.975 | 0.151 |
FIIIL | 4.4–6.2 | 4.9 ± 0.6 | 4.8–5.9 | 5.4 ± 0.8 | 4.9–5.7 | 5.2 ± 0.2 | 5.3–5.8 | 5.6 ± 0.3 | 0.622 | 0.111 | 0.949 |
FIVL | 2.9–4.1 | 3.3 ± 0.2 | 3.0–4.4 | 3.6 ± 1.0 | 3.2–3.9 | 3.5 ± 0.2 | 3.2–3.6 | 3.4 ± 0.3 | 0.726 | 0.110 | 0.471 |
THL | 13.8–16.0 | 14. ± 0.8 | 13.1–17.6 | 15.3 ± 3.2 | 13.7–14.7 | 14.2 ± 0.4 | 15.7–16.3 | 16. 0 ± 0.4 | 0.177 | 0.646 | 0.996 |
TL | 14.5–16.1 | 15.0 ± 0.5 | 13.0–17.8 | 15.4 ± 3.4 | 14.3–15.5 | 15.0 ± 0.3 | 16.4–16.7 | 16.5 ± 0.2 | 0.019* | 0.558 | 0.628 |
TW | 2.8–4.4 | 3.4 ± 0.6 | 2.9–3.3 | 3.1 ± 0.3 | 3.9–4.4 | 4.1 ± 0.2 | 4.0–4.7 | 4.3 ± 0.5 | 0.124 | 0.004** | 0.050 |
TFL | 18.0–21.5 | 19.5 ± 1.14 | 19.3–23.5 | 21.4 ± 3.0 | 18.6–20.4 | 19.7 ± 0.7 | 21.0–22.4 | 21.7 ± 1.0 | 0.483 | 0.444 | 0.697 |
FL | 11.2–14.8 | 12.9 ± 1.1 | 12.4–15.6 | 14.0 ± 2.2 | 12.3–13.5 | 12.840 ± 0.5 | 13.7–14.8 | 14.3 ± 0.8 | 0.562 | 0.946 | 0.734 |
Holotype. CIBXJ190503 (Fig.
Holotype CIBXJ190503 of Megophrys xianjuensis sp. nov. and holotype CIBWYF00164 of M. lishuiensis A, B dorsal view and ventral of Megophrys xianjuensis sp. nov., respectively C, D dorsal view and ventral of M. lishuiensis, respectively E oral of Megophrys xianjuensis sp. nov. showing vomerine ridges (red arrows) F oral of M. lishuiensis showing no vomerine ridge G ventral view of hand of Megophrys xianjuensis sp. nov. H ventral view of hand of M. lishuiensis I ventral view of foot of Megophrys xianjuensis sp. nov. J ventral view of foot of M. lishuiensis.
Paratype. One adult male CIB20180514007 and one adult female CIB20180514008 collected from the type locality of the new species by Bin Wang on 14 May 2018; one adult male CIBXJ190501 and one adult female CIBXJ190505 collected from the type locality by Bin Wang on 7 May 2019; five adult males CIBXJ20190801, CIBXJ20190802, CIBXJ20190803, CIBXJ20190804 and CIBXJ20190805 collected from the type locality by Bin Wang on 28 August 2019.
Megophrys xianjuensis sp. nov. is assigned to the genus Megophrys based on molecular phylogenetic analyses and the following generic diagnostic characters: snout shield-like; projecting beyond the lower jaw; canthus rostralis distinct; chest gland small and round, closer to the axilla than to midventral line; femoral gland on rear of thigh; vertical pupils.
The new species could be identified from its congeners by a combination of the following morphological characters: (1) small size (SVL 31.0–36.3 mm in males and 41.6 mm in female); (2) vomerine ridge present and vomerine teeth absent; (3) tongue not notched behind; (4) a small horn-like tubercle at the edge of each upper eyelid; (5) tympanum distinctly visible, rounded; (6) two metacarpal tubercles in hand; (7) relative finger lengths: II < I < IV < III; (8) toes with rudimentary webbing at bases; (9) heels overlapping when thighs are positioned at right angles to the body; (10) tibiotarsal articulation reaching tympanum to eye when leg stretched forward; (11) an internal single subgular vocal sac in male; (12) in breeding male, the nuptial pads with black nuptial spines on the dorsal bases of the first and second fingers.
SVL 34.4 mm; head wider than long (HDW/HDL ratio 1.2); snout obtusely pointed, protruding well beyond the margin of the lower jaw in ventral view; loreal region vertical and concave; canthus rostralis well developed; top of head flat on in dorsal view; an small horn-like tubercle at the edge of the upper eyelid; eye large and convex, eye diameter 40.4% of head length; pupils vertical; nostril orientated laterally, closer to snout than eye; tympanum distinct, TYP/EYE ratio 0.56; vomerine ridges present and vomerine teeth absent; margin of tongue smooth, not notched behind (Fig.
Forelimbs slender, the length of lower arm and hand 44.9% of SVL; fingers slender, relative finger lengths: II < I < IV < III; tips of digits globular, without lateral fringes; subarticular tubercle distinct at the base of each fingers; two metacarpal tubercles, prominent, oval-shaped, the inner one bigger than the outer (Fig.
Hindlimbs slender, heels overlapping when thighs are positioned at right angles to the body, tibiotarsal articulation reaching tympanum to eye when leg stretched forward; tibia length slightly longer than thigh length; relative toe lengths I < II < V < III < IV; tips of toes round, slightly dilated; subarticular tubercle absent; toes with rudimentary webbing at bases; lateral fringe narrow; inner metatarsal tubercle oval-shaped; outer metatarsal tubercle absent (Fig.
Dorsal rough, with numerous granules; several large warts scattered on flanks; an small horn-like tubercle at the edge of each upper eyelid; a dark brown inverted triangular pattern between anterior corner of eyes, tubercles on the dorsum forming a weak X-shaped ridge and two discontinuous dorsolateral parallel ridges on either side of the X-shaped ridge; several tubercles on the flanks and dorsal surface of thighs and tibias, and limbs barred with dark brown forming four transverse rows; supratympanic fold distinct (Fig.
Ventral surface with numerous white granules; chest gland distinct and round, closer to the axilla than to midventral line; femoral gland on rear of thigh; posterior end of the body protrudes distinct and appears as an arc-shaped swelling, upper the anal region (Fig.
An inverted triangular brown speckle between the eyes; an X-shaped ridges on the dorsum of body, four transverse bands on the dorsal surface of the hindlimb; several dark brown and white vertical bars on the lower and upper lip; the venter purple and the colour of throat is deeper than belly, flank and middle of throat with black brown spots, numerous white granules on the ventral surface and limbs; palms and soles purple and the metacarpal tubercles are orange, tip of digits greyish white; pectoral and femoral glands white (Fig.
Dorsal surface fade to olive; the inverted triangular brown speckle between the eyes, X-shaped ridges on dorsum and transverse bands on limbs and digits and brown spots on flank and middle throat are more distinct; ventral surface greyish white; creamy-white substitutes the orange in metacarpal tubercles; the posterior of ventral surface of body, inner of thigh and upper of tibia creamy-white (Fig.
In some adult individuals a brown Y-shaped marking on the dorsum of head and disconnected with a X-shaped marking on back (Fig.
Ten advertisement calls from two individuals of the new species were recorded in the Xianju County, Zhejiang Province, China between 21:00–23:00 on 7 May 2019. The call description is based on recordings of the holotype CIBXJ090503 (Fig.
Adult females with SVL 41.6 mm, larger than adult males with 31.0–36.3 mm. Adult males have a single subgular vocal sac (Fig.
The tadpole was confirmed as Megophrys xianjuensis sp. nov. by molecular phylogenetic analyses. The following tadpole description is based on the specimen CIBXJT19050704 at Stage 31 (Fig.
By having small size body, Megophrys xianjuensis sp. nov. differs from M. aceras, M. auralensis, M. binlingensis, M. carinense, M. caudoprocta, M. chuannanensis, M. damrei, M. edwardinae (in female), M. feae, M. flavipunctata, M. gigantica, M. glandulosa, M. himalayana, M. intermedia, M. jingdongensis, M. kobayashii, M. kalimantanensis, M. lekaguli, M. liboensis, M. ligayae, M. longipes, M. major, M. mangshanensis, M. maosonensis, M. medogensis, M. omeimontis, M. oreocrypta, M. periosa, M. popei, M. robusta, M. spinata, M. sangzhiensis, M. shapingensis and M. shuichengensis and M. takensis (maximum SVL < 42.0 mm in the new species vs. minimum SVL > 45 mm in the latter).
By lacking vomerine teeth, Megophrys xianjuensis sp. nov. differs from M. aceras, M. ancrae, M. carinense, M. baluensis, M. caudoprocta, M. chuannanensis, M. damrei, M. daweimontis, M. dongguanensis, M. fansipanensis, M. flavipunctata, M. glandulosa, M. hoanglienensis, M. himalayana, M. insularis, M. intermedia, M. jingdongensis, M. jinggangensis, M. jiulianensis, M. kalimantanensis, M. kobayashii, M. lancip, M. lekaguli, M. liboensis, M. ligayae, M. major, M. mangshanensis, M. maosonensis, M. medogensis, M. megacephala, M. montana, M. nasuta, M. nankunensis, M. nanlingensis, M. omeimontis, M. oropedion, M. oreocrypta, M. palpebralespinosa, M. parallela, M. parva, M. periosa, M. popei, M. robusta, M. rubrimera, M. sangzhiensis, M. stejnegeri, M. takensis, M. zhangi and M. zunhebotoensis (vs. present in the latter).
By having a small horn-like tubercle at the edge of each upper eyelid, Megophrys xianjuensis sp. nov. differs from M. binchuanensis, M. binlingensis, M. damrei, M. gigantica, M. minor, M. nasuta, M. nankiangensis, M. oropedion, M. pachyproctus, M. spinata, M. stejnegeri, M. takensis, M. wuliangshanensis, M. wushanensis, M. zhangi, and M. zunhebotoensis (vs. tubercle lacking in the latter) and differs from M. carinense, M. feae, M. gerti, M. hansi, M. intermedia, M. kalimantanensis, M. koui, M. liboensis, M. microstoma, M. palpebralespinosa, M. popei, M. shuichengensis, and M. synoria (vs. having a prominent and elongated tubercle at the edge of each upper eyelid in the latter).
With its tongue not notched behind, Megophrys xianjuensis sp. nov. differs from M. ancrae, M. baolongensis, M. binlingensis, M. boettgeri, M. carinense, M. cheni, M. chuannanensis, M. damrei, M. dringi, M. fansipanensis, M. feae, M. feii, M. flavipunctata, M. gerti, M. glandulosa, M. hoanglienensis, M. huangshanensis, M. insularis, M. jiulianensis, M. jingdongensis, M. kalimantanensis, M. kuatunensis, M. liboensis, M. mangshanensis, M. maosonensis, M. medogensis, M. minor, M. nankiangensis, M. nanlingensis, M. omeimontis, M. oropedion, M. pachyproctus, M. parallela, M. popei, M. robusta, M. sangzhiensis, M. shapingensis, M. shuichengensis, M. spinata, M. vegrandis, M. wawuensis, M. zhangi, and M. zunhebotoensis (vs. tongue notched behind in the latter).
By toes rudimentary webbing at base, Megophrys xianjuensis sp. nov. differs from M. brachykolos, M. carinense, M. flavipunctata, M. jingdongensis, M. jinggangensis, M. lini, M. major, M. palpebralespinosa, M. popei, M. shuichengensis, and M. spinata (vs. at least one-fourth webbed in the latter).
By heels overlapping when thighs are positioned at right angles to the body, Megophrys xianjuensis sp. nov. differs from M. acuta, M. brachykolos, M. dongguanensis, M. huangshanensis, M. kuatunensis, M. nankunensis, M. obesa, M. ombrophila, and M. wugongensis (vs. heels not meeting when thighs are positioned at right angles to the body in the latter).
By tibiotarsal articulation reaching forward to the region between tympanum and eye when hindlimb is stretched along the side of the body, Megophrys xianjuensis sp. nov. differs from M. baolongensis, M. nankiangensis, M. pachyproctus, M. serchhipii, M. shuichengensis, and M. tuberogranulata (vs. reaching posterior corner of the eye in the latter); differs from M. daweimontis, M. glandulosa, M. lini, M. major, M. medogensis, M. obesa, and M. sangzhiensis (vs. reaching the anterior corner of the eye or beyond eye or nostril and tip of snout in the latter); differs from M. leishanensis (vs. reaching middle part of eye); differs from M. mufumontana (vs. reaching tympanum in males and to the eye in females).
By having an internal single subgular vocal sac in male, Megophrys xianjuensis sp. nov. differs from M. caudoprocta, M. shapingensis, and M. shuichengensis (vs. vocal sac absent in the latter).
By having nuptial pads and nuptial spines on the dorsal base of the first and second fingers in breeding male, Megophrys xianjuensis sp. nov. differs from M. acuta, M. feii, M. shapingensis, and M. shuichengensis (vs. nuptial pads and nuptial spines lacking in the latter); differs from M. boettgeri, and M. elfina (vs. nuptial pads and nuptial spines only on the first finger in the latter).
Megophrys boettgeri and M. kuatunensis were suggested to be distributed in Zhejiang Province, China and might be sympatric with Megophrys xianjuensis sp. nov. (Fei et al. 2016;
Molecular phylogenetic analyses revealed that the new species was genetically closer to M. lishuiensis. The new species can be distinguished from M. lishuiensis by a series of morphological characters as follows. Vomerine ridges present vs. vomerine ridges absent; heels overlapping when thighs are positioned at right angles to the body vs. heels just meeting or not meeting; light round patches on the shoulder absent vs. present; having significantly lower ratios of UEW, ED, TYD, LW and TW to SVL in males; and having significantly higher ratios of UEW, ED and HAL to SVL in females (all p-values < 0.05; Table
Megophrys xianjuensis sp. nov. is known from the type locality, Xianju County, Zhejiang Province, China at elevations between 320–480 m a.s.l. This new species is frequently found on stones in the streams in the subtropical montane forests (Fig.
The specific epithet xianjuensis refers to Xianju County, Zhejiang Province, China, where the type locality of the species is located. We propose the common name “Xianju horned toad” in English and Xian ju Jiao Chan in Chinese.
Although Megophrys xianjuensis sp. nov. superficially resembles M. lishuiensis, our integrative comparisons with morphological and molecular data can clearly identify the new species from the latter. This indicates that conserved morphology could hamper species delineation, requiring the incorporation of detailed morphological, genetic, and bioacoustic data to recognise cryptic species. The discovery of Megophrys xianjuensis sp. nov. brings the total number of species in the genus to 93, with 49 of them recorded in China (Fei et al. 2016;
According to records in Fei et al. (2016) and
Many Megophrys species have narrow distributions fitting the “micro-endemism” model of
We are grateful to the editors and reviewers for their working on the manuscript. Collections in field were permitted by Administration of Xianju National Park (No. AXJNP2018020801). This study was approved by the animal ethical committee of Chengdu Institute of Biology, Chinese Academy of Sciences, and animal experiments were carried out following the institutional guidelines (No. 2018CIBAEC0345). The work was supported by Demonstration Project on Biodiversity Conservation and Development of Xianju County Project Financed by AFD, Project supported by the biodiversity investigation, observation and assessment program (2019–2023) of Ministry of Ecology and Environment of China, and the Strategic Priority Research Program of the Chinese Academy of Sciences (Grant No. XDPB0202).
Table S1. Measurements of the adult specimens of Megophrys xianjuensis sp. nov. and M. lishuiensis
Data type: measurements
Table S2. Measurements of the tadpole specimens of Megophrys xianjuensis sp. nov.
Data type: measurements
Table S3. Uncorrected p-distance between Megophrys species based on the 16S gene sequences
Data type: molecular data