Print
Description of six new species of the subgenus Panophrys within the genus Megophrys (Anura, Megophryidae) from southeastern China based on molecular and morphological data
expand article infoJian Wang, Zhi-Tong Lyu, Zu-Yao Liu, Cheng-Kai Liao§, Zhao-Chi Zeng, Jian Zhao, Yu-Long Li, Ying-Yong Wang
‡ Sun Yat-sen University, Guangzhou, China
§ Jiulianshan National Nature Reserve, Jiangxi, China
Open Access

Abstract

The diversity of the subgenus Panophrys within the genus Megophrys has been revealed to be extremely underestimated from southeastern China. Herpetological surveys coupled with extensive sampling in a longitudinal mountain belt located in southeastern China resulted in the discoveries of six new species of the subgenus Panophrys. Furthermore, the new discoveries support the findings of “micro-endemism”, “sympatric phenomenon” and “sympatric but distant phylogenetically” which appear to be common among Panophrys species, and also indicates that the Asian horned toads would be good candidates for studies on speciation and biogeography, and additionally emphasizes the conservation difficulties of these toads.

Keywords

Conservation, Megophrys, southeastern China, species diversity, subgenus Panophrys, speciation, biogeography

Introduction

The Asian horned toads (Megophrys) comprise 85 recognized species which were previously classified in the subfamily Megophryinae (Frost 2019). They are widespread in montane forest area in tropical and subtropical Asia, including southern mainland China, southern and eastern Himalayas, across Indochina to Malay, to the islands of the Sunda Shelf and the Philippines (Chen et al. 2017; Deuti et al. 2017; Mahony et al. 2017; Mahony et al. 2018; Li et al. 2018; Liu et al. 2018; Munir et al. 2018; Messenger et al. 2019; Tapley et al. 2018; Frost 2019). As a consequence of both morphological similarity among species and the complex patterns of genetic divergence, the taxonomy of these toads always has been controversial. Although several researchers have proposed different taxonomic schemes in recent decades (Dubois 1987; Rao and Yang 1997; Dubois and Ohler 1998; Jiang et al. 2003; Zheng et al. 2004; Frost et al. 2006; Li and Wang 2008; Fei et al. 2009; Fei and Ye 2016; Chen et al. 2017; Mahony et al. 2017), the debate remains. Based on a large-scale molecular analysis, Chen et al. (2017) considered that subfamily Megophryinae is composed of five genera, namely Atympanophrys Tian & Hu, 1983, Brachytarsophrys Tian & Hu, 1983, Megophrys Kuhl & Van Hasselt, 1822, Ophryophryne Boulenger, 1903 and Xenophrys Günther, 1864. Almost at the same time, based on the integrative analysis with phylogeny and morphological examination, Mahony et al. (2017) treated the entire subfamily Megophryinae as a single genus Megophrys and divided it into seven subgenera, i.e. Atympanophrys, Brachytarsophrys, Megophrys, Ophryophryne, Panophrys Rao & Yang, 1997, Pelobatrachus Beddard, 1908 and Xenophrys, and 25 known species were placed in the subgenus Panophrys. Subsequently, Liu et al. (2018) partially agreed with this taxonomic system based on a substantial study on phylogenetic similarity, and revealed unusually high levels of species diversity in the subgenus Panophrys with a total number of 60 species, 2.4 times of previously known, including 41 unnamed cryptic species and 39 of which were from southeastern China. Therefore, Panophrys species diversity from southeastern China is extremely underestimated.

In the past years, we have carried out continual herpetological surveys coupled with extensive sampling in a longitudinal mountain belt with a west-east width of 100 km, north-south length of 800 km in the middle of southeastern China, from Hong Kong and Shenzhen in the Pearl River Delta, across the Jiulian Mountains and Luoxiao Mountains, north to the Yangtze River (Fig. 1). The surveys resulted in the discovery of 15 unnamed Panophrys species (Liu et al. 2018) and descriptions of 14 new species of amphibians and reptiles, namely Leptobrachella laui (Sung, Yang & Wang, 2014), Megophrys (Brachytarsophrys) popei (Zhao, Yang, Chen, Chen & Wang, 2014), M. (Panophrys) cheni (Wang & Liu, 2014), M. (Pa.) lini (Wang & Yang, 2014), Megophrys (Pa.) jinggangensis (Wang, 2012), Nidirana nankunensis Lyu, Zeng, Wang, Lin, Liu & Wang, 2017, Amolops albispinus Sung, Hu, Wang, Liu & Wang, 2016, Gracixalus jinggangensis Zeng, Zhao, Chen, Chen, Zhang & Wang, 2017 and Gr. guangdongensis Wang, Zeng, Lyu, Liu & Wang, 2018; Goniurosaurus yingdeensis Wang, Yang & Cui, 2010, Go. zhelongi Wang, Jin, Li & Grismer, 2014, Takydromus albomaculosus Wang, Gong, Liu & Wang, 2017, Rhabdophis guangdongensis Zhu, Wang, Takeuchi & Zhao, 2014 and Opisthotropis shenzhenensis Wang, Guo, Liu, Lyu, Wang, Luo, Sun & Zhang, 2017.

Figure 1. 

Collection localities of the six new Megophrys species in this study: 1 Mt. Yinping in Dongguan City of Guangdong Province, the type locality of M. dongguanensis sp. nov. 2 Mt. Nankun in Huizhou City of Guangdong Province, the type locality of M. nankunensis sp. nov., and one of the localities of its sympatric species, M. jiulianensis sp. nov. 3a Nanling Nature Reserve in Shaoguan City of Guangdong Province, the type locality of M. nanlingensis sp. nov. 3b Mt. Qiyun in Ganzhou City of Jiangxi Province, the other collection locality of M. nanlingensis sp. nov. 4 Mt. Jiulian in Ganzhou City of Jiangxi Province, the type locality of M. jiulianensis sp. nov. 5 Yangshimu Scenic Area in Pingxiang City and adjacent Wugongshan Scenic Area in Ji’an City of Jiangxi Province, the type locality of M. wugongensis sp. nov. 6 Mt. Mufu in Yueyang City of Hunan Province, the type locality of M. mufumontana sp.nov.

In the present study, we re-reviewed several species defined by Liu et al. (2018) from this mountain belt based on molecular and morphological data and formally described six new species of Megophrys.

Material and methods

Sampling

For molecular analysis, a total of 42 samples (17 were attained from GenBank and 25 were new materials in this study) from the collection of unnamed specimens of the subgenus Panophrys, together with 39 samples (37 from GenBank and two new materials) from 21 recognized species of Panophrys were used as in-groups in this study. In addition, four samples (all from GenBank) from two recognized species of the subgenus Atympanophrys, four samples (three from GenBank and one new materials) from two recognized species of the subgenus Brachytarsophrys, three samples (one from GenBank and two new materials) from two recognized species of the subgenus Ophryophryne, two samples (all from GenBank) from two recognized species of the subgenus Pelobatrachus, and six samples (five from GenBank and one new materials) of three recognized species of the subgenus Xenophrys were incorporated into our dataset and used as out-groups. Details see Table 1. All muscle samples were preserved in 95% ethanol and stored at -40 °C.

Localities, voucher information, and GenBank accession numbers for all specimens used in this study.

Subgenus of Megophrys s. l. ID Species name Locality Specimen voucher no. Genbank Accession No.
16S CO1
Panophrys 1 M. dongguanensis sp. nov. China: Mt. Yinping, Dongguan City, Guangdong SYS a001971/CIB110006 MK524097 MK524128
2 M. dongguanensis sp. nov. China: Mt. Yinping, Dongguan City, Guangdong SYS a001972 MK524098 MK524129
3 M. dongguanensis sp. nov. China: Mt. Yinping, Dongguan City, Guangdong SYS a001973 MH406647 MH406083
4 M. dongguanensis sp. nov. China: Mt. Yinping, Dongguan City, Guangdong SYS a001974 MH406648 MH406084
5 M. dongguanensis sp. nov. China: Mt. Yinping, Dongguan City, Guangdong SYS a001975 MH406649 MH406085
6 M. dongguanensis sp. nov. China: Mt. Yinping, Dongguan City, Guangdong SYS a002007 MH406654 MH406090
7 M. jiulianensis sp. nov. China: Mt. Jiulian, Ganzhou City, Jiangxi SYS a002107 MK524099 MK524130
8 M. jiulianensis sp. nov. China: Mt. Jiulian, Ganzhou City, Jiangxi SYS a002108 MK524100 MK524131
9 M. jiulianensis sp. nov. China: Mt. Jiulian, Ganzhou City, Jiangxi SYS a002109 MK524101 MK524132
10 M. jiulianensis sp. nov. China: Mt. Jiulian, Ganzhou City, Jiangxi SYS a004219 MH406791 MH406253
11 M. jiulianensis sp. nov. China: Mt. Nankun, Huizhou City, Guangdong SYS a003622 MK524102 MK524133
12 M. jiulianensis sp. nov. China: Mt. Nankun, Huizhou City, Guangdong SYS a003623 MK524103 MK524134
13 M. mufumontana sp. nov. China: Mt. Mufu, Pingjiang County, Hunan SYS a006390/CIB110012 MK524104 MK524135
14 M. mufumontana sp. nov. China: Mt. Mufu, Pingjiang County, Hunan SYS a006391 MK524105 MK524136
15 M. mufumontana sp. nov. China: Mt. Mufu, Pingjiang County, Hunan SYS a006392 MK524106 MK524137
16 M. mufumontana sp. nov. China: Mt. Mufu, Pingjiang County, Hunan SYS a006419 MK524107 MK524138
17 M. nankunensis sp. nov. China: Mt. Nankun, Huizhou City, Guangdong SYS a004498 MK524108 MK524139
18 M. nankunensis sp. nov. China: Mt. Nankun, Huizhou City, Guangdong SYS a004499 MK524109 MK524140
19 M. nankunensis sp. nov. China: Mt. Nankun, Huizhou City, Guangdong SYS a004500 MK524110 MK524141
20 M. nankunensis sp. nov. China: Mt. Nankun, Huizhou City, Guangdong SYS a004501 MH406822 MH406284
21 M. nankunensis sp. nov. China: Mt. Nankun, Huizhou City, Guangdong SYS a004502 MH406823 MH406285
22 M. nankunensis sp. nov. China: Mt. Nankun, Huizhou City, Guangdong SYS a004503 MH406824 MH406286
Panophrys 23 M. nanlingensis sp. nov. China: Nanling Nature Reserve, Shaoguan City, Guangdong SYS a001959 MK524111 MK524142
24 M. nanlingensis sp. nov. China: Nanling Nature Reserve, Shaoguan City, Guangdong SYS a001960 MK524112 MK524143
25 M. nanlingensis sp. nov. China: Nanling Nature Reserve, Shaoguan City, Guangdong SYS a001964 MH406646 MH406082
26 M. nanlingensis sp. nov. China: Mt. Qiyun, Chongyi County, Jiangxi SYS a002334 MH406686 MH406132
27 M. nanlingensis sp. nov. China: Mt. Qiyun, Chongyi County, Jiangxi SYS a002356 MK524113 MK524144
28 M. nanlingensis sp. nov. China: Mt. Qiyun, Chongyi County, Jiangxi SYS a002357 MH406687 MH406133
29 M. nanlingensis sp. nov. China: Mt. Qiyun, Chongyi County, Jiangxi SYS a002358 MH406688 MH406134
30 M. wugongensis sp. nov. China: Wugongshan Scenic Area, Anfu County, Jiangxi SYS a002610 MK524114 MK524145
31 M. wugongensis sp. nov. China: Wugongshan Scenic Area, Anfu County, Jiangxi SYS a002611 MK524115 MK524146
32 M. wugongensis sp. nov. China: Yangshimu Scenic Area, Pingxiang City, Jiangxi SYS a002625 MK524116 MK524147
33 M. wugongensis sp. nov. China: Wugongshan Scenic Area, Anfu County, Jiangxi SYS a004777/CIB110011 MK524117 MK524148
34 M. wugongensis sp. nov. China: Wugongshan Scenic Area, Anfu County, Jiangxi SYS a004796 MK524118 MK524149
35 M. wugongensis sp. nov. China: Wugongshan Scenic Area, Anfu County, Jiangxi SYS a004797 MK524119 MK524150
36 M. wugongensis sp. nov. China: Wugongshan Scenic Area, Anfu County, Jiangxi SYS a004798 MK524120 MK524151
37 M. wugongensis sp. nov. China: Wugongshan Scenic Area, Anfu County, Jiangxi SYS a004799 MH406852 MH406314
38 M. wugongensis sp. nov. China: Wugongshan Scenic Area, Anfu County, Jiangxi SYS a004800 MH406853 MH406315
39 M. wugongensis sp. nov. China: Wugongshan Scenic Area, Anfu County, Jiangxi SYS a004801 MH406854 MH406316
40 M. wugongensis sp. nov. China: Wugongshan Scenic Area, Anfu County, Jiangxi SYS a004802 MH406855 MH406317
41 M. wugongensis sp. nov. China: Wugongshan Scenic Area, Anfu County, Jiangxi SYS a004803 MH406856 MH406318
42 M. wugongensis sp. nov. China: Wugongshan Scenic Area, Anfu County, Jiangxi SYS a004804 MK524121 MK524152
43 M. acuta China: Heishiding Nature Reserve, Zhaoqing City, Guangdong SYS a001957 KJ579118 MF667898
44 M. acuta China: Heishiding Nature Reserve, Zhaoqing City, Guangdong SYS a002159 MF667869 MF667899
Panophrys 45 M. binlingensis China: Mt. Wawu, Meishan City, Sichuan SYS a005313 MH406892 MH406354
46 M. binlingensis China: Mt. Wawu, Meishan City, Sichuan SYS a005314 MH406893 MH406355
47 M. boettgeri China: Longhu Forest Station, Shaowu City, Fujian SYS a004126 MH406785 MH406245
48 M. boettgeri China: Mt. Wuyi, Fujian SYS a004150 MF667879 MF667914
49 M. brachykolos China: Hongkong SYS a005563 MK524122 MK524153
50 M. brachykolos China: Hongkong SYS a005564 MK524123 MK524154
51 M. caudoprocta China: Mt. Badagongshan, Zhangjiajie City, Hunan SYS a004281 MH406795 MH406257
52 M. caudoprocta China: Mt. Badagongshan, Zhangjiajie City, Hunan SYS a004293 MH406796 MH406258
53 M. cheni China: Taoyuandong Nature Reserve, Zhuzhou City, Hunan SYS a002123 KJ560396 MF667904
54 M. cheni China: Taoyuandong Nature Reserve, Zhuzhou City, Hunan SYS a002140 MF667872 MF667905
55 M. huangshanensis China: Mt. Huangshan, Anhui SYS a002702 MF667882 MF667919
56 M. huangshanensis China: Mt. Huangshan, Anhui SYS a002703 MF667883 MF667920
57 M. insularis China: Nan’ao Island, Guangdong SYS a002169 (Holotype) MF667887 MF667924
58 M. insularis China: Nan’ao Island, Guangdong SYS a002170 MF667888 MF667925
59 M. jingdongensis China: Mt. Wuliang, Yunnan SYS a003928 MH406773 MH406232
60 M. jingdongensis China: Mt. Wuliang, Yunnan SYS a003929 MH406774 MH406233
61 M. jinggangensis China: Mt. Jinggang, Jiangxi SYS a004028 MH406780 MH406239
62 M. jinggangensis China: Mt. Sifang, Hengdong County, Hunan SYS a004825 MH406858 MH406320
63 M. kuatunensis China: Mt. Wuyi, Jiangxi SYS a003449 MF667881 MF667916
64 M. lini China: Nanfengmian Nature Reserve, Jiangxi SYS a002128 KJ560416 MF667907
65 M. lini China: Nanfengmian Nature Reserve, Jiangxi SYS a002381 MF667874 MF667908
66 M. minor China: Dujiangyan City, Sichuan SYS a003209 MF667862 MF667891
67 M. minor China: Dujiangyan City, Sichuan SYS a003210 MF667863 MF667892
68 M. obesa China: Heishiding Nature Reserve, Guangdong SYS a002271 KJ579121 MH406123
69 M. obesa China: Heishiding Nature Reserve, Guangdong SYS a005025 MH406868 MH406330
70 M. ombrophila China: Mt. Wuyi, Fujian WUYI2015101 KX856397 /
71 M. omeimontis China: Mt. Laojunshan, Yibin City, Sichuan SYS a002741 MH406710 MH406162
Panophrys 72 M. omeimontis China: Hejiang County, Sichuan SYS a004916 MH406864 MH406326
73 M. sangzhiensis China: Mt. Badagongshan, Hunan SYS a004307 MH406798 MH406260
74 M. sangzhiensis China: Mt. Badagongshan, Hunan SYS a004313 MH406802 MH406264
75 M. spinata China: Mt. Leigong, Guizhou SYS a002226 MH406675 MH406115
76 M. spinata China: Mt. Leigong, Guizhou SYS a002227 MH406676 MH406116
77 M. tuberogranulatus China: Mt. Badagongshan, Hunan SYS a004310 MH406801 MH406263
78 M. wushanensis China: Shennongjia Forestry District, Hubei SYS a003008 MH406732 MH406184
79 M. wushanensis China: Shennongjia Forestry District, Hubei SYS a003009 MH406733 MH406185
80 M. wuliangshanensis China: Mt. Wuliang, Yunnan SYS a003924 MH406771 MH406230
81 M. wuliangshanensis China: Mt. Wuliang, Yunnan SYS a003925 MH406772 MH406231
Atympanophrys 82 M. gigantica China: Mt. Ailao, Yunnan SYS a003883 MH406766 MH406225
83 M. gigantica China: Mt. Wuliang, Yunnan SYS a003933 MH406775 MH406234
84 M. shapingensis China: Mt. Wawu, Sichuan SYS a005310 MH406890 MH406352
85 M. shapingensis China: Zhaojue County, Sichuan SYS a005339 MH406897 MH406359
Brachytarsophrys 86 M. chuannanensis China: Hejiang County, Sichuan SYS a004926 MH406901 MH406364
87 M. chuannanensis China: Hejiang County, Sichuan SYS a004927 MH406902 MH406365
88 M. popei China: Taoyuandong Nature Reserve, Hunan SYS a001864 MH406361 KM504256
89 M. popei China: Mt. Jinggang, Jiangxi SYS a004209 MK524124 MK524155
Ophryophryne 90 M. hansi Vietnam: Quang Nam, Tra My District AMNH 163680 KY022203 /
91 M. microstoma China: Mt. Wuhuang, Pubei County, Guangxi SYS a003492 MK524125 MK524156
92 M. microstoma China: Mt. Wuhuang, Pubei County, Guangxi SYS a003493 MK524126 MK524157
Pelobatrachus 93 M. nasuta Malaysia: Sabah, Lahad Datu District FMNH 231281 KY022186 /
94 M. stejnegeri Philippines: Mindanao, Bukidnon Province FMNH 250842 KY022190 /
Xenophrys 95 M. glandulosa China: Mt. Gaoligong, Yunnan SYS a003758 MH406755 MH406214
96 M. glandulosa China: Mt. Gaoligong, Yunnan SYS a003794 MH406759 MH406218
97 M. mangshanensis China: Mt. Longtou, Guangdong SYS a002750 MF667866 MF667895
98 M. mangshanensis China: Mt. Dayao, Guangxi SYS a004870 MH406861 MH406323
99 M. medogensis China: Medog County, Tibet SYS a002932 MH406725 MH406177
100 M. medogensis China: Medog County, Tibet SYS a002933 MK524127 MK524158

DNA Extraction, PCR and sequencing

Genomic DNA was extracted from muscular tissue using a DNA extraction kit from Tiangen Biotech (Beijing) Co., Ltd. All samples were sequenced for two mitochondrial genes, i.e., partial 16S ribosomal RNA gene (16S) and complete cytochrome C oxidase 1 gene (CO1). Primers used for 16S were L3975 (5’-CGCCTGTTTACCAAAAACAT-3’) and H4551 (5’-CCGGTCTGAACTCAGATCACGT-3’) following Simon et al. (1994), and for CO1 were Chmf4 (5’-TYTCWACWAAYCAYAAAGAYATCGG-3’) and Chmr4 (5’-ACYTCRGGRTGRCCRAARAATCA-3’) following Meyer et al. (2005). PCR amplifications were processed in a 20-reaction volume with the cycling conditions that initial denaturing step at 95 °C for 4 min, 35 cycles of denaturing at 94 °C for 40 s, annealing at 53 °C for 40 s and extending at 72 °C for 1 min, and final extending step of 72 °C for 10 min. PCR products were purified with spin columns. The purified products were sequenced with both forward and reverse primers using BigDye Terminator Cycle Sequencing Kit per the guidelines, on an ABI Prism 3730 automated DNA sequencer by Shanghai Majorbio Bio-pharm Technology Co., Ltd and Beijing Genomics Institute. All sequences have been deposited in GenBank (Table 1).

Phylogenetic analyses

DNA sequences were aligned in MEGA 6 (Tamura et al. 2013) by the Clustal W algorithm with default parameters (Thompson et al. 1997). Two gene segments, 535 base pairs (bp) of 16S and 645 bp of CO1, were concatenated seriatim into a 1180-bp single sequence. The dataset was partitioned according to the genes and codon positions, and then tested respectively in jmodeltest v2.1.2 with Akaike and Bayesian information criteria, all resulting in the best-fitting nucleotide substitution models of GTR + I + G. Sequenced data was analyzed using Bayesian inference (BI) in MrBayes 3.2.4 (Ronquist et al. 2012). Three independent runs were conducted in BI analysis, each of which was performed for 2,000,000 generations and sampled every 1000 generations with the first 25% samples were discarded as burn-in, resulting a potential scale reduction factor (PSRF) of < 0.01. Pairwise distances (p-distance) were calculated in MEGA 6 using the uncorrected p-distance model.

Morphometrics

All specimens were fixed in 10 % buffered formalin and later transferred to 70% ethanol for preservation, and deposited at the Museum of Biology, Sun Yat-sen University (SYS) and Chengdu Institute of Biology, the Chinese Academy of Sciences (CIB), China.

Measurements follow Fei et al. (2009), and were taken with digital calipers to the nearest 0.1 mm. These measurements were as follows:

SVL snout‒vent length (from tip of snout to vent);

HDL head length (from tip of snout to rear of jaws);

HDW head width (head width at commissure of jaws);

SNT snout length (from tip of snout to anterior corner of eye);

ED eye diameter (diameter of exposed portion of eyeball);

IOD interorbital distance (minimum distance between upper eyelids);

IND internasal distance (distance between nares);

TD tympanum diameter (horizontal diameter of tympanum);

TED tympanum–eye distance (distance from anterior edge of tympanum to posterior corner of eye);

HND hand length (distance from distal end of radioulna to tip of phalanx of finger III);

RAD radioulna length;

TIB tibia length (distance from knee to heel);

FTL foot length (distance from distal end of tibia to tip of distal phalanx of toe IV).

Sex was determined by direct observation of calls, the presence of internal vocal sac openings and the presence of testicles observed through dissection for males, as well as the presence of eggs and ovaries on the abdomen through anatomise for females. Presence or absence of nuptial pads/spines was examined with a microscope.

Comparative morphological data of Megophrys species allocated to the subgenus Panophrys (currently contains 32 species) (Mahony et al. 2017; Tapley et al. 2017; Wang et al. 2017a; Wang et al. 2017b; Zhang et al. 2017; Li et al. 2018; Tapley et al. 2018), and a small-sized species M. feii (incertae sedis), were obtained from examination of museum specimens (see Appendix 1) and from the literature (Table 2). The order of the new species accounts follows the distributions of the new species that located in the longitudinal mountain belt from the south to the north.

References for morphological characters for congeners of the subgenus Panophrys and Megophrys feii (incertae sedis).

ID Subgenus Panophrys Literature obtained
1 M. acuta Wang, Li & Jin, 2014 Li et al. 2014
2 M. baolongensis Ye, Fei & Xie, 2007 Ye et al. 2007
3 M. binchuanensis Ye & Fei, 1995 Ye and Fei 1995
4 M. binlingensis Jiang, Fei & Ye, 2009 Fei et al. 2009
5 M. boettgeri (Boulenger, 1899) Fei et al. 2012
6 M. brachykolos Inger & Romer, 1961 Inger and Romer 1961
7 M. caudoprocta Shen, 1994 Fei et al. 2012
8 M. cheni (Wang & Liu, 2014) Wang et al. 2014
9 M. daweimontis Rao & Yang, 1997 Fei et al. 2012
10 M. fansipanensis Tapley, Cutajar, Mahony, Nguyen, Dau, Luong, Le, Nguyen, Nguyen, Portway, Luong & Rowley, 2018 Tapley et al. 2018
11 M. huangshanensis Fei & Ye, 2005 Fei et al. 2012
12 M. hoanglienensis Tapley, Cutajar, Mahony, Nguyen, Dau, Luong, Le, Nguyen, Nguyen, Portway, Luong &, 2018 Tapley et al. 2018
13 M. insularis (Wang, Liu, Lyu, Zeng & Wang, 2017) Wang et al. 2017b
14 M. jingdongensis Fei & Ye, 1983 Fei et al. 2012
15 M. jinggangensis (Wang, 2012) Wang et al. 2012
16 M. kuatunensis Pope, 1929 Fei et al. 2012
17 M. latidactyla Orlov, Poyarkov & Nguyen, 2015 Orlov et al. 2015
18 M. leishanensis Li, Xu, Liu, Jiang, Wei & Wang, 2018 Li et al. 2018
19 M. liboensis (Zhang, Li, Xiao, Li, Pan, Wang, Zhang & Zhou, 2017) Zhang et al. 2017
20 M. lini (Wang &Yang, 2014) Wang et al. 2014
21 M. lishuiensis (Wang, Liu & Jiang, 2017) Wang et al. 2017a
22 M. minor Stejneger, 1926 Wang et al. 2017b
23 M. obesa Wang, Li & Zhao, 2014 Li et al. 2014
24 M. ombrophila Messenger & Dahn, 2019 Messenger et al. 2019
25 M. omeimontis Liu, 1950 Fei et al. 2009
26 M. palpebralespinosa Bourret, 1937 Fei et al. 2012
27 M. rubrimera Tapley, Cutajar, Mahony, Chung, Dau, Nguyen, Luong & Rowley, 2017 Tapley et al. 2017
28 M. sangzhiensis Jiang, Ye & Fei, 2008 Jiang et al. 2008
29 M. shuichengensis Tian & Sun, 1995 Tian et al. 2000
30 M. spinata Liu & Hu, 1973 Fei et al. 2012
31 M. tuberogranulatus Shen, Mo & Li, 2010 Mo et al. 2010
32 M. wuliangshanensis Ye & Fei, 1995 Ye and Fei 1995
33 M. wushanensis Ye & Fei, 1995 Ye and Fei 1995
Incertae sedis
1 M. feii Yang, Wang & Wang, 2018 Yang et al. 2018

Results

Phylogenetics

The Bayesian inference (BI) phylogenetic tree was integrated in Figure 2; the p-distances at the mitochondrial 16S rRNA gene fragment among all samples of the subgenus Panophrys were given in Table 3.

Figure 2. 

Bayesian inference tree derived from partial DNA sequences of the mitochondrial 16S rRNA + CO1 genes.

Uncorrected p-distances among Megophrys species of the subgenus Panophrys in this study, based on mitochondrial 16S r RNA genes.

Species & ID No. (1)–(6) (7)–(12) (13)–(16) (17)–(22) (23)–(29) (30)–(42) (43)–(44) (45)–(46) (47)–(48) (49)–(50) (51)–(52) (53)–(54) (55)–(56) (57)–(58)
M. dongguanensis sp. nov. (1)–(6) 0–0.2
M. jiulianensis sp. nov. (7)–(12) 5.3–5.8 0–0.7
M. mufumontana sp. nov. (13)–(16) 6.3 3.7–4 0
M. nankunensis sp. nov. (17)–(22) 2.6–2.8 4.7–4.9 4.9–5.1 0–0.7
M. nanlingensis sp. nov. (23)–(29) 5.3–6.1 5.3–6.3 5.8–6.5 4.9–5.8 0–0.7
M. wugongensis sp. nov. (30)–(42) 5.3–5.4 4–4.2 6–6.1 4.7–4.9 4.9–5.4 0
M. acuta (43)–(44) 7.4–7.5 7–7.4 7.9 7.9–8.1 6.7–7.2 8.1–8.2 0
M. binlingensis (45)–(46) 6.3 4.9–5.4 5.1 5.8–6.1 5.6–6.1 5.4 8.2 0
M. boettgeri (47)–(48) 5.1–5.4 2.3–2.8 2.8–3 4–4.4 4.4–5.4 4.2–4.4 6.5–6.8 4.2–4.4 0–0.2
M. brachykolos (49)–(50) 6.8 5.8–6.3 6.8 5.8–6.1 6.1–7.5 7.7 7.2 7.7 6.1 0–0.1
M. caudoprocta (51)–(52) 6.3 3.5–3.7 4 4.9–5.1 6.8–7.5 5.8 6.8 4.4 3.3 6.5 0
M. cheni (53)–(54) 3–3.3 3.5–4 4.2–4.4 5.8–6 3.5–4.4 5.7–6.6 7–7.2 4.7–4.9 2.6 5.1–5.4 4.4–4.7 0–0.2
M. huangshanensis (55)–(56) 5.6 3–3.3 3.7 4.7–4.9 5.1–5.8 4.7 6.5 4.9 0.9–1.2 6.5 3.5 3.3–3.5 0
M. insularis (57)–(58) 4.4–4.9 5.1–5.8 5.1–6.5 3.5–4.2 4.4–5.8 4.9–5.1 7.7–8.4 5.8 4.4–5.1 6.3–6.5 6.1–6.5 3.3–3.5 4.7–5.4 0
M. jingdongensis (59)–(60) 7.4 6–6.3 5.8 5.8–6 6.7–7.5 5.3–5.4 7.9 6.2–6.7 4.9–5.1 7.9 4.7 4.7–4.9 5.6 6.7 0
M. jinggangensis (61)–(62) 5.8–6.3 4.4–4.9 4.2–4.4 4.7–5.1 4.9–5.8 4.4 6.7 4.9 3.7–4 6.1 5.4 3–3.3 4.4 4.7 5.3 0–0.5
M. kuatunensis (63) 3.3 4–4.2 4.2 2.8–3 4.7–5.4 5.1 7.4 5.4 3.3–3.5 4.7 4.4 2.6–2.8 3.7 3.5 5.8 4.4 0
M. lini (64)–(65) 5.1 5.1–5.6 6–6.1 4.7–4.9 3.7–4.4 5.6 6.5 6.3 4.2–4.4 4.9 5.8 3.3–3.5 4.7 4.9 6.3 4.4 3.7 0
M. minor (66)–(67) 7.9–8.2 7–7.7 6.8–7 7.7–8.2 7–7.2 6.5–6.8 8.9–9.1 6.3–6.5 5.6–5.8 8.7–8.9 7–7.2 6.5–6.8 6.3–6.5 8.4–8.6 7.2–7.5 6.8–7 6.5–6.8 7.7–7.9 0–0.2
M. obesa (68)–(69) 4.7–5.8 5.6–6 6.3 4.2 4.4–4.9 5.1 7.4 7.2 4.2–4.4 6.3 6.8 2.8–3 4.9–5.1 4.9 6.3 4.9–5.3 4.2 4.2 7.2–7.5 0
M. ombrophila (70) 5.1 5.6–5.8 6–6.1 4.4–4.7 6–6.8 5.3–5.4 8.8 6.8 4.7–4.9 7 6.8 3–3.3 5.1–5.4 4.7 6.7 5.3–5.8 4.9 5.8 8.2–8.4 3 0
M. omeimontis (71)–(72) 5.8 4.2–4.4 4.9 4.7–4.9 5.1–5.8 4.3–4.4 7.9 5.5 4–4.2 7 4.2 3.7–4 4.7 5.4 3 4.4–4.9 4.7 5.1 5.8–6.1 5.6 5.6 0
M. sangzhiensis (73)–(74) 6.3 5.3–5.8 5.8 6.3 5.6–6 5.1 8.6 6.9 4.7–4.9 8.2 4.9 4.7–4.9 5.4 5.6 3.9 5.6 5.8 6.3 7.2–7.5 7.2 6.5 4 0
M. spinata (75)–(76) 6.5 5.1–5.6 5.8 6.3–6.5 5.3–6.1 4.9 8.4 4.5 4.4–4.7 8.4 5.1 4.7–4.9 4.7 6.3 3.9 5.8 6 5.6 6.5–6.8 7 6.7 3 2 0
M. tuberogranulatus (77) 4.7 2.6–2.8 3 3.3–3.5 4.7–5.1 4 6.5 7.5 3.4 5.4 3 2.1–2.3 2.3 4.2 4 3 3 4.7 5.8–6.1 4.7–5.8 4.4 3.7 3.7 3.7 0
M. wushanensis (78)–(79) 4.9–5.4 3.5–4 5.8 4.2–4.7 5.6–6.3 4.9–5.1 7.2–7.5 7.5–7.6 3–3.5 5.8 4.2–4.4 3.3–3.7 3.7–4 3.7–4 5.4–5.6 3.7–4 3.7–4 5.6–5.8 7.2–7.5 5.4–5.8 4.7–4.9 3.7–4 5.1–5.4 5.1–5.4 2.1 0
M. wuliangshanensis (80)–(81) 7.2 5.3–5.8 4 5.8–6 7–7.2 6–6.1 8.4 6.5–6.7 4.9–5.1 7.5 5.6 5.1–5.4 5.4–5.6 6.7 3.7 5.8 5.1 6.7 6.5–6.8 5.8 6.5 3.5 4.9 4.4 4.2 4.7–5.1 0–0.5

In our phylogenic tree, all sequences of the genus Megophrys grouped into six clades with strong node support values, which were consistent with the results from Mahony et al. (2017) and Liu et al. (2018), and corresponded to the six subgenera: Panophrys, Ophryophryne, Xenophrys, Atympanophrys, Brachytarsophrys and Pelobatrachus. The subgenus Panophrys is further divided into three subclades, named western subclade A, western subclade B and eastern subclade.

The western subclade A is composed of Megophrys omeimontis, M. binglingensis, M. sangzhiensis, M. spinata, M. wuliangshanensis and M. jingdongensis, and the western subclade B is composed of M. minor, all of which are distributed in southwestern China.

The eastern subclade contains 14 known species from southeastern China, i.e. M. boettgeri, M. huangshanensis, M. kuatunensis, M. brachykolos, M. insularis, M. cheni, M. lini, M. jinggangensis, M. obesa, M. ombrophila, M. acuta, M. sangzhiensis, M. caudoprocta, M. tuberogranulatus and wushanensis, and other six lineages made up of samples from the aforementioned longitudinal mountain belt in the middle of southeastern China with significant genetic differences (Table 3).

Among them, all samples from Mt. Mufu, Hunan (samples 13–16 in Table 1) clustered into a basal lineage of an eastern subclade with strong node supports and almost have no molecular differences; further, this population can be distinguished from all known species and other undescribed lineages by distinctive morphological characters and significant molecular differences with a lowest p-distance of 2.8%. Therefore, the population from Mt. Mufu represented a separately evolving lineage, and is described as a new species, Megophrys (Panophrys) mufumontana sp. nov., below.

All samples from Mt. Wugong, Jiangxi (samples 30–42 from Yangshimu Scenic Area and Wugongshan Scenic Area) clustered into a lineage with strong node supporting values and almost no genetic differences, which was defined as a species and recognized as M. sp12 by Liu et al. (2018); further, the population from Mt. Wugong can be distinguished from all known species and other undescribed lineages by distinctive morphological differences and significant molecular differences with a lowest p-distance of 4%. Therefore, the population from Mt. Wugong represented a separately evolving lineage and is described as a new species, Megophrys (Panophrys) wugongensis sp. nov., below.

All samples from Mt. Yinping, Guangdong (samples 1–6) clustered into a lineage with strong node supportg values and small genetic differences (highest p-distance 0.2%), which was defined as a species and recognized as M. sp11 by Liu et al. (2018); samples 17–22 from Mt. Nankun, Guangdong clustered into a lineage with strong node support values and small genetic differences (highest p-distance 0.7%), which was defined as a species and recognized as M. sp10 by Liu et al. (2018); these two populations are sister taxa to each other with significant genetic differences (p-distances 2.6–2.8%), and can be further distinguished from all known species and other undescribed lineages by distinctive morphological differences and significant molecular differences. Therefore, the populations from Mt. Yinping and Mt. Nankun represented two separately evolving lineages, and are described as new species, Megophrys (Panophrys) dongguanensis sp. nov. and Megophrys (Panophrys) nankunensis sp. nov., below.

Samples 7–10 from Mt. Jiulian, Jiangxi and samples 11–12 from Mt. Nankun, Guangdong clustered into a lineage with small genetic differences (highest p-distance 0.7%), which is a sister subclade to M. boettgeri and M. huangshanensis with large genetic differences (lowest p-distance 2.3%); therefore, these samples represented a separately evolving lineage, which was defined as a species and recognized as M. sp30 by Liu et al. (2018), and is described as a new species, Megophrys (Panophrys) jiulianensis sp. nov., below.

Samples 26–29 from Mt. Qiyun, Jiangxi were defined as a species and recognized as M. sp6 by Liu et al. (2018) and the samples 23–25 from Nanling Nature Reserve, Guangdong were defined as a species and recognized as M. sp7 by Liu et al. (2018). Although the populations from two locations are divided into two branches, the highest p-distance is only 0.7%. Moreover, there are no distinct morphological characters that can distinguish them from each other. Herein, we considered these two populations as one taxon, which is the sister taxon to M. lini with large genetic differences (p-distances 3.7–4.4%), representing a new species and described as, Megophrys (Panophrys) nanlingensis sp. nov., below.

Taxonomic accounts

Megophrys (Panophrys) dongguanensis J. Wang & Y.Y. Wang, sp. nov.

Fig. 3, Table 4

Holotype

SYS a001973, adult male, collected by Run-Lin Li on 13 December 2012 from Mt. Yinping, Xiegang County (22°54'17.20"N, 114°13'23.88"E; 132 m a.s.l.), Dongguan City, Guangdong Province, China.

Paratypes (10 males)

SYS a002007, adult male, collected on 17 March 2013 by Run-Lin Li from Mt. Yinping, Qingxi County (22°53'26.21"N, 114°10'14.82"E; 277 m a.s.l.), Dongguan City, China; adult males, SYS a001971/CIB110006, SYS a001972, 1974–1975, collected on 12–13 December 2012, SYS a001492–1495, collected on 23 December 2012 by Run-Lin Li from the same locality as the holotype (100–300 m a.s.l.).

Diagnosis

(1) Body size small to moderate, SVL 30.2–39.3 mm in 11 adult male specimens; (2) head width slightly larger than head length, HDW/HDL ratio 1.04–1.09; (3) snout pointed in dorsal view; (4) tympanum distinct, moderate-sized, TD/ED ratio 0.42–0.60; (5) strong vomerine ridge bearing vomerine teeth; (6) margin of tongue not notched behind; (7) hindlimbs short, heels not meeting, tibio-tarsal articulation reaching the region between tympanum and eye; (8) presence of subarticular tubercles and absence of lateral fringes on fingers, relative finger lengths II < I ≤ IV < III; (9) toes with rudiment of webbing at their bases and without lateral fringes, subarticular tubercles only present at the base of each toe; (10) numerous granules present on dorsal surface of body, several large tubercles present on surface of flanks; (11) presence of a barely visible reddish horn-like tubercle at the edge of the upper eyelid; (12) supratympanic fold distinct, whitish; (13) yellowish brown dorsally, with an incomplete dark triangular marking between eyes and usually an X-shaped marking on back of trunk; (14) ventral surface black brown, with white spots on posterior surface of abandon; (15) males with a single subgular vocal sac; (16) presence of nuptial pads with darker nuptial spines on dorsal surface of the first and second fingers in adult males during breeding season, respectively.

Comparisons

Comparative data of Megophrys dongguanensis sp. nov. with M. feii and the 33 recognized members of Megophrys s.l. allocated to the subgenus Panophrys are listed in Table 5.

With significantly smaller body size, SVL 30.2–39.3 mm in males, Megophrys dongguanensis sp. nov. differs from the eight members with larger SVL values: M. baolongensis (42.0–45.0 mm in males), M. binlingensis (45.1–51.0 mm in males), M. caudoprocta (81.3 mm in male), M. jingdongensis (53.0–56.5 mm in males), M. omeimontis (56.0–59.5 mm in males), M. sangzhiensis (54.7 mm in single male), M. spinata (47.2–54.4 mm in males) and M. shuichengensis (102.0–118.3 mm in males).

Megophrys dongguanensis sp. nov. differs from 12 species occurring in eastern and southern China (M. acuta, M. brachykolos, M. boettgeri, M. cheni, M. huangshanensis, M. insularis, M. jinggangensis, M. kuatunensis, M. lini, M. lishuiensis, M. obesa and M. ombrophila) by the following combination of characters: presence of vomerine teeth (vs. absent in M. acuta, M. boettgeri, M. brachykolos, M. cheni, M. huangshanensis, M. kuatunensis, M. lini, M. lishuiensis, M. obesa and M. ombrophila), margin of tongue not notched posteriorly (vs. notched in M. boettgeri, M. cheni, M. huangshanensis, M. insularis and M. kuatunensis), absence of lateral fringes on toes (vs. presence of narrow lateral fringes on toes in M. acuta, M. jinggangensis and M. kuatunensis; presence of wide lateral fringes on toes in M. boettgeri, M. cheni and M. lini), toes with rudimentary webbing (vs. toes without webbing in M. lishuiensis, M. kuatunensis and M. ombrophila), hindlimbs short, with heels not meeting when the flexed hindlimbs are held at right angles to the body axis (vs. hindlimbs comparatively longer, with heels meeting or overlapping in M. cheni, M. boettgeri, M. kuatunensis, M. jinggangensis and M. lini), tibio-tarsal articulation reaching the region between tympanum and eye when hindlimb is stretched along the side of the body (vs. reaching forward to the shoulder in M. brachykolos and to the posterior edge of tympanum in M. insularis).

From the remaining 10 species occurring in China, Megophrys dongguanensis sp. nov. can be distinguished by the presence of vomerine teeth (vs. absent in M. binchuanensis, M. leishanensis, M. minor, M. tuberogranulatus, M. wuliangshanensis and M. wushanensis), by the unnotched tongue (vs. tongue notched in M. daweimontis, M. liboensis, M. minor and M. rubrimera), by the absence of lateral fringes on toes (vs. wide in M. binchuanensis, M. liboensis, M. palpebralespinosa and M. wushanensis (in males); narrow in M. rubrimera), by the rudimentary webbing on toes (vs. toes without webbing in M. rubrimera and M. wuliangshanensis; at least one-fourth webbed in M. palpebralespinosa), by the heels not meeting when the flexed hindlimbs are held at right angles to the body axis (vs. heels meeting in M. binchuanensis and M. tuberogranulatus; heels meeting or overlapping in M. minor and M. wushanensis; heels overlapping in M. leishanensis, M. liboensis, M. palpebralespinosa and M. wuliangshanensis).

Megophrys dongguanensis sp. nov. differs from the remaining species, M. fansipanensis, M. hoanglienensis and M. latidactyla, by the small horn-like tubercle at edge of upper eyelid (vs. slightly large in M. latidactyla), by the unnotched tongue (vs. tongue notched in M. fansipanensis, M. hoanglienensis and M. latidactyla), by the absence of lateral fringes on toes (vs. wide in M. latidactyla), by the presence of rudimentary webbing on toes (vs. webbing indistinct or absent in M. fansipanensis and M. hoanglienensis).

Megophrys dongguanensis sp. nov. further differs from M. feii, for which molecular data are lacking and cannot be allocated to any subgenus base on morphology only (Yang et al. 2018) by the larger body size, SVL 30.2–39.3 mm in males (VS. 24.3–25.1 mm in males), presence of nuptial pad with nuptial spines in males during breeding season (vs. absent), presence of vomerine teeth (vs. absent), unnotched tongue (vs. slightly notched), absence of lateral fringes on toes (vs. moderate or wide), heels not meeting when the flexed hindlimbs are held at right angles to the body axis (vs. heels overlapping).

Description of holotype

Adult male. Body moderate-sized, SVL 38.0 mm; head width slightly larger than head length, HWD/HDL 1.09; snout pointed in dorsal view, projecting, sloping backward to mouth in profile, protruding well beyond margin of lower jaw; top of head flat; eye large, ED/HDL 0.40, pupil vertical; nostril oblique ovoid; canthus rostralis well developed, forming the beginning of a fleshy, protruding ridge, that continues over the upper eyelid, and transitions into a supratympanic fold that terminates in the scapular region; loreal region slightly oblique; internasal distance slightly larger than interorbital distance; tympanum distinct, moderate-sized, TD/ED 0.54; large ovoid choanae at the base of the maxilla; presence of vomerine ridge bearing vomerine teeth; margin of tongue not notched posteriorly; internal vocal slits present near the rear of the lower mandible.

Radioulna length and hand length 0.24 of SVL; fingers without webbing and lateral fringes, relative finger length II < I < IV < III; tips of fingers slightly dilated, round; presence of subarticular tubercles on finger III, and one subarticular tubercle at the bases of each finger; outer metacarpal tubercles indistinct, inner metacarpal tubercles distinct and observably enlarged. Hindlimbs short, tibio-tarsal articulation reaching the region between tympanum and eye when hindlimb is stretched along the side of the body; heels not meeting when the flexed hindlimbs are held at right angles to the body axis; tibia length 0.41 of SVL and foot length 0.61 of SVL; relative toe length I < II < V < III < IV; tips of toes round and slightly dilated; presence of rudimentary webbing on toes but absence of lateral fringes and tarsal folds; one subarticular tubercle at the bases of each toe; presence of a long ovoid inner metatarsal tubercle and absence of outer metatarsal tubercle.

Dorsal skin texture rough with dense granules; granules forming discontinuous X-shaped ridge with two discontinuous dorsolateral ridges on both sides at the central trunk; several large tubercles present on dorsal surface of flanks, thighs, shanks and forearms; four small tubercles present on the edge of upper eyelid, one of which is more prominent; distinct narrow supratympanic fold curving posteroventrally from posterior corner of eye to a level above insertion of arm; ventral skin texture smooth, several granules present on surface of abandon, ventral and posterior surface of thighs; pectoral gland small, closer to axilla; single femoral gland on rear of thigh.

Measurements of holotype (in mm)

SVL 38.0, HDL 12.0, HDW 13.1, SNT 4.5, IND 3.9, IOD 3.6, ED 4.8, TD 2.6, TED 2.1, HND 9.1, RAD 9.2, FTL 23.2, TIB 15.6.

Coloration of holotype in life

(Fig. 3A–E) Yellowish brown dorsally, with a dark triangular marking between eyes. A wide oblique black band present on forearm. Dorsal surface of fingers and hindlimbs with dark grey transverse bands. Point of snout dark brown, presence of a vertical dark brown band below the eye. Tubercles on the edge of upper eyelid reddish. Supratympanic fold whitish tan. Ventral surface dark brown, with a black longitudinal band on surface of throat, several white spots present on ventral surface of limbs. Digits, inner and outer metacarpal tubercles greyish white, inner metatarsal tubercle greyish brown. Pectoral glands and femoral glands white. Iris yellowish brown.

Figure 3. 

Megophrys dongguanensis sp. nov. in life: A–E SYS a001973, the male holotype F SYS a001492, a male paratype with more distinct skin ridges, granules and tubercles on dorsal surface of body.

Coloration of holotype in preservative

Yellowish brown fades to greyish brown dorsally. Triangular marking between eyes, oblique bands on dorsal forearms, transverse bands on dorsal fingers and hindlimbs become indistinct. Color of ventral surface fades, all bands and spots become indistinct.

Variation

Measurements of type series are listed in Table 4. All paratypes are very similar to holotype in morphology and color pattern. However, one male (SYS a001492) has more distinct skin ridges, granules and tubercles on dorsal surface of body (Fig. 3, F).

Measurements (in mm; minimum-maximum, mean ± SD) of the type series of Megophrys dongguanensis sp. nov. and M. nankunensis. sp. nov., respectively.

Species Megophrys dongguanensis sp. nov. Megophrys nankunensis sp. nov.
Males (n = 9) Males (n = 11) Females (n = 2)
SVL 30.2–39.3 (36.3 ± 3.3) 29.9–34.9 (32.7 ± 1.5) 39.4–41.9
HDL 11.1–12.6 (12.0 ± 0.5) 9.0–11.3 (10.0 ± 0.7) 11.9–12.5
HDW 11.5–13.2 (12.8 ± 0.6) 10.1–12.6 (10.9 ± 0.7) 13.0–13.7
SNT 3.7–4.6 (4.2 ± 0.3) 2.8–3.7 (3.3 ± 0.3) 4.1–4.4
IND 3.7–3.9 (3.6 ± 0.2) 2.2–3.8 (3.1 ± 0.5) 3.1–3.9
IOD 3.2–3.6 (3.5 ± 0.2) 2.4–3.4 (2.8 ± 0.3) 3.1–3.2
ED 4.6–5.2 (4.9 ± 0.2) 3.1–4.4 (3.8 ± 0.4) 4.7–5.2
TD 2.1–2.8 (2.5 ± 0.3) 1.4–2.4 (1.9 ± 0.3) 2.6–2.7
TED 1.9–2.4 (2.1 ± 0.1) 0.8–1.7 (1.2 ± 0.2) 2.2–2.3
HND 8.3–9.8 (9.2 ± 0.5) 6.7–8.6 (7.5 ± 0.5) 9.7–10.2
RAD 8.2–9.7 (9.2 ± 0.5) 5.5–8.4 (6.6 ± 0.7) 8.0–8.4
FTL 13.4–16.5 (15.5 ± 1.08) 10.9–14.1 (12.2 ± 0.7) 14.6–15.3
TIB 19.9–23.4 (22.2 ± 1.11) 16.3–21.6 (18.2 ± 1.4) 22.6–25.5
HDL/SVL 0.32–0.37 (0.33 ± 0.02) 0.27–0.33 (0.31 ± 0.02) 0.30
HDW/SVL 0.33–0.40 (0.35 ± 0.02) 0.30–0.37 (0.33 ± 0.02) 0.33
HDW/HDL 1.04–1.09 (1.06 ± 0.02) 1.00–1.20 (1.10 ± 0.06) 1.09–1.10
SNT/HDL 0.33–0.37 (0.35 ± 0.01) 0.28–0.40 (0.33 ± 0.03) 0.35
SNT/SVL 0.11–0.12 (0.12 ± 0.01) 0.08–0.11 (0.10 ± 0.01) 0.10–0.11
IND/HDW 0.26–0.30 (0.28 ± 0.01) 0.21–0.34 (0.28 ± 0.04) 0.24–0.28
IOD/HDW 0.27–0.28 (0.27 ± 0.01) 0.23–0.30 (0.26 ± 0.02) 0.23–0.24
ED/HDL 0.37–0.44 (0.40 ± 0.02) 0.34–0.41 (0.38 ± 0.02) 0.39–0.42
ED/SVL 0.12–0.16 (0.14 ± 0.02) 0.09–0.13 (0.11 ± 0.01) 0.12
TD/ED 0.42–0.60 (0.51 ± 0.06) 0.43–0.61 (0.50 ± 0.06) 0.49–0.57
TED/TD 0.73–1.09 (0.88 ± 0.14) 0.58–0.74 (0.66 ± 0.05) 0.85
HND/SVL 0.24–0.28 (0.25 ± 0.01) 0.21–0.25 (0.23 ± 0.01) 0.24–0.25
RAD/SVL 0.24–0.28 (0.25 ± 0.01) 0.16–0.25 (0.20 ± 0.02) 0.20
TIB/SVL 0.41–0.46 (0.43 ± 0.02) 0.35–0.42 (0.37 ± 0.01) 0.37
FTL/SVL 0.58–0.70 (0.61 ± 0.04) 0.53–0.62 (0.56 ± 0.03) 0.57–0.61

Diagnostic characters separating the seven new species described in this study from Megophrys feii (incertae sedis) and 33 recognizing species of the Megophrys s.l. allocated to the subgenus Panophrys.

Species SVL Horn-like tubercle at edge of upper eyelid1 Vomerine teeth2 Tongue3 Lateral fringes on toes4 Toes5 TD/ED TIB/SVL
males (N) females (N)
M. dongguanensis 30.2–39.3 (9) / + + + 0.42–0.60 0.41–0.46
M. nankunensis 29.9–34.9 (11) 39.4–41.9 (2) + + + 0.43–0.61 0.35–0.42
M. jiulianensis 30.4–33.9 (9) 34.1–37.5 (2) + + + + 0.50–0.59 0.44–0.48
M. nanlingensis 30.5–37.3 (10) / + + + + + 0.43–0.57 0.45–0.51
M. wugongensis 31.0–34.1 (4) 38.5–42.8 (9) + + 0.45–0.53 0.37–0.44
M. mufumontana 30.1–30.8 (2) 36.3 (2) + + + 0.51–0.58 0.47–0.53
M. acuta 27.1–33.0 (10) 28.1–33.6 (4) ++ + + 0.57–0.71 0.38–0.45
M. baolongensis 42.0–45.0 (5) / + + 0.41 0.46
M. binchuanensis 32.0–36.0 (4) 40.2–42.5 (2) + or ‒ ++ + 0.33–0.50 0.46–0.48
M. binlingensis 45.1–51.0 (3) / + / + 0.47–0.52 0.52–0.53
M. boettgeri 34.5–37.8 (20) 39.7–46.8 (10) + + ++ + 0.40–0.67 0.45–0.49
M. brachykolos 33.7–39.3 (5) 33.9–45.9 (2) + + > 0.50 0.37–0.42
M. caudoprocta 81.3 (1) / ++ + / + 0.50 0.51
M. cheni 26.2–29.5 (15) 31.8–34.1 (3) + ++ ++ + 0.41–0.54 0.50–0.54
M. daweimontis 34.0–37.0 (18) 40.0–46.0 (3) + + / / 0.54
M. fansipanensis 30.9–44.3 (13) 41.7–42.5 (2) + + + 0.53–0.80 0.49–0.59
M. feii 24.3–25.1 (4) 28.2–28.9 (2) + + ++ + 0.51–0.58 0.48–0.55
M. hoanglienensis 37.4–47.6 (11) 59.6 (1) + + + 0.54–0.75 0.44–0.63
M. huangshanensis 36.0–41.6 (4) 44.2 (1) + + <0.50 0.42–0.45
M. insularis 36.8–41.2 (5) 47.1 (1) + + + + 0.46–0.57 0.40–0.43
M. jingdongensis 53.0–56.5 (3) 63.5 (1) + + + ++ +++ / 0.58–0.59
M. jinggangensis 35.1–36.7 (2) 38.4–41.6 (3) ++ + + + 0.73–0.88 0.47–0.50
M. kuatunensis 26.2–29.6 (13) 37.4 (1) + + + 0.44 0.38–0.48
M. latidactyla 38.9 (1) / ++ + ++ + 0.85 0.52
M. leishanensis 30.4–38.7 (10) 42.3 (2) + + / /
M. liboensis 34.7–67.7 (5) 60.8–70.6 (8) +++ + + ++ + 0.48–0.78 0.44–0.61
M. lini 34.1–39.7 (20) 37.0–39.9 (4) + ++ + 0.40–0.60 0.46–0.53
M. lishuiensis 30.7–34.7 (13) 36.9–40.4 (3) + / /
M. minor 34.5–41.2 (4) / + + 0.8–0.83 0.46–0.48
M. obesa 35.6 (1) 37.5–41.2 (6) + + 0.51–0.66 0.41–0.47
M. ombrophila 27.4–34.5 (5) 32.8–35.0 (4) + 0.52–0.69 0.32–0.41
M. omeimontis 56.0–59.5 (10) 68.0–72.5 (3) + + + + + / 0.52–0.56
M. palpebralespinosa 36.2–38.0 (2) / ++ + ++ +++ / 0.55
M. rubrimera 26.7–30.5 (8) / + + + + 0.58–0.76 0.48–0.56
M. sangzhiensis 54.7 (1) / + + + + + 0.62 0.59
M. shuichengensis 102.0–118.3 (7) 99.8–115.6 (6) ++ + ++ +++ 0.67 0.43–0.47
M. spinata 47.2–54.4 (18) 54.0–55.0 (2) + ++ +++ 0.43 0.56–0.58
M. tuberogranulatus 33.2–39.6 (9) 50.5 (1) + or ‒ + 0.50 0.45–0.51
M. wuliangshanensis 27.3–31.6 (10) 41.0–41.5 (2) + or ‒ 0.50 0.50–0.51
M. wushanensis 30.4–35.5 (10) 38.4 (1) ‒ (in female), ++ (in male) + 0.50 0.47–0.48

Etymology

The specific epithet “dongguanensis” is in reference to the type locality, Dongguan City of the new species. We propose the common English name “Dongguan Horned Toad” and Chinese name “Dong Guan Jiao Chan (东莞角蟾)”.

Distribution and natural history

Currently, Megophrys dongguanensis sp. nov. is only known from Mt. Yinping, Guangdong Province, China. It inhabits flowing montane streams and the nearby forest floor and leaf litter at elevations between 100–300 m. Advertisement calls of males were noticed from mid-December until April of the next year just before the rainy season. Males were found calling on rocks in the flowing streams. Tadpoles could be found in this period.

Megophrys (Panophrys) nankunensis J. Wang, Zeng & Y.Y. Wang, sp. nov.

Fig. 4, Table 4

Holotype

SYS a004498, adult male, collected by Jian Wang and Hai-Long He on 20 October 2015 from Mt. Nankun (23°38'19"N, 113°53'24"E; 400 m a.s.l.), Longmen County, Huizhou City, Guangdong Province, China.

Paratypes (10 males & two females)

Adult females, SYS a004506–4507, collected by Jian Wang and Hai-Long He on 20 October 2015; adult males, SYS a002023, 2032–2033, collected by Run-Lin Li on 20 March 2013, SYS a004499–4504, SYS a004505/CIB110007, collected by Jian Wang and Hai-Long He on 20 October 2015, all from Mt. Nankun at elevations between 300–650 m.

Diagnosis

(1) Body size small, SVL 29.9–34.9 mm in 11 adult males, 39.4–41.9 mm in two adult females; (2) head width slightly larger than head length, HDW/HDL ratio 1.00–1.20; (3) snout rounded in dorsal view, tip of snout slightly sharpened; (4) tympanum distinct, moderate-sized, TD/ED ratio 0.43–0.61; (5) strong vomerine ridge bearing vomerine teeth; (6) margin of tongue not notched behind; (7) shanks short, heels not meeting when the flexed hindlimbs are held at right angles to the body axis; tibia-tarsal articulation reaching forward to the region between tympanum and eye when hindlimb is stretched along the side of the body; (8) TIB/SVL ratio 0.35–0.42, FTL/SVL ratio 0.53–0.62; (9) absence of lateral fringes on fingers, presence of an indistinct subarticular tubercle on the bases of each finger, relative finger lengths II < I < IV < III; (10) toes with rudimentary webbing at their bases and without lateral fringes, subarticular tubercles only present on the bases of each toes; (11) dorsal surface with dense granules, surface of flanks and dorsal surface of limbs with large tubercles; (12) edge of eye lid with a small reddish horn-like tubercle; (13) supratympanic fold distinct, forming a depressed supraaxillary gland above insertion of arm; (14) dorsum beige to dark brown, with indistinct light brown patches, with an incomplete dark triangular marking between eyes; (15) males with a single subgular vocal sac, and dense dark villiform nuptial spines present on dorsal surface of first and second fingers during breeding season, respectively; (16) gravid females bear creamy yellow oocytes.

Comparisons

Comparative data of Megophrys nankunensis sp. nov. with M. dongduanensis sp. nov., M. feii and the 33 recognized members of Megophrys s.l. allocated to the subgenus Panophrys are listed in Table 5.

In the ML and BI phylogenetic trees (Fig. 2), Megophrys nankunensis sp. nov. is a sister taxon to M. dongguanensis sp. nov. (p=4.6–5.0%) with high node-supporting value (0.1 in BI, 100% in ML%), and differs from the later by the snout rounded in dorsal view, tip of snout slightly sharpened (vs. snout pointed in dorsal view, tip of snout not sharpened), supratympanic fold forming a depressed supraaxillary gland above insertion of arm (vs. supraaxillary gland absent).

With significantly smaller body size, SVL 29.9–34.9 mm in males and 39.4–41.9 mm in females, Megophrys nankunensis sp. nov. differs from the 12 members with larger SVL values: M. baolongensis (42.0–45.0 mm in males), M. binlingensis (45.1–51.0 mm in males), M. caudoprocta (81.3 mm in single male), M. hoanglienensis (37.4–47.6 mm in males), M. jingdongensis (53.0–56.5 mm in males, 63.5 mm in single female), M. latidactyla (38.9 mm in single male), M. omeimontis (56.0–59.5 mm in males, 68.0–72.5 mm in females), M. palpebralespinosa (36.2–38.0 mm in males), M. sangzhiensis (54.7 mm in single male), M. shuichengensis (102.0–118.3 mm in males, 99.8–115.6 mm in females), M. spinata (47.2–54.4 mm in males, 54.0–55.0 mm in females) and M. tuberogranulatus (50.5 in single female).

Megophrys nankunensis sp. nov. differs from 12 species occurring in eastern and southern China (M. acuta, M. brachykolos, M. boettgeri, M. cheni, M. huangshanensis, M. insularis, M. jinggangensis, M. kuatunensis, M. lini, M. lishuiensis, M. obesa and M. ombrophila) by the following combination of characters: presence of vomerine teeth (vs. absent in M. acuta, M. boettgeri, M. brachykolos, M. cheni, M. huangshanensis, M. kuatunensis, M. lini, M. lishuiensis, M. obesa and M. ombrophila), absence of lateral fringes on toes (vs. presence of narrow lateral fringes on toes in M. acuta, M. jinggangensis and M. kuatunensis; presence of wide lateral fringes on toes in M. boettgeri, M. cheni and M. lini), toes with rudimentary webbing (vs. toes without webbing in M. lishuiensis, M. kuatunensis and M. ombrophila), hindlimbs short, with heels not meeting when the flexed hindlimbs are held at right angles to the body axis (vs. hindlimbs comparatively longer, with heels meeting or overlapping in M. cheni, M. boettgeri, M. kuatunensis, M. jinggangensis and M. lini), tibio-tarsal articulation reaching forward to the region between tympanum and eye when hindlimb is stretched along the side of the body (vs. reaching forward to the shoulder in M. brachykolos and to the posterior edge of tympanum in M. insularis), relative finger lengths II < I < IV < III (vs. IV < I < II < III in M. brachykolos and I < II < IV < III in M. obesa and M. ombrophila); supratympanic fold forming a depressed supraaxillary gland above insertion of arm (vs. supraaxillary gland swollen in M. insularis; absent in other 11 species).

Megophrys nankunensis sp. nov. differs from the remaining nine members of the Megophrys s.l. allocated to the subgenus Panophrys which share a moderate or small body size, by the by the small horn-like tubercle at edge of upper eyelid (vs. horn-like tubercle indistinct or absent in M. binchuanensis, M. minor, M. wuliangshanensis and M. wushanensis; long point in M. liboensis), presence of vomerine teeth (vs. absent in M. binchuanensis, M. leishanensis, M. minor, M. wuliangshanensis and M. wushanensis), absence of lateral fringes on toes (vs. wide in M. binchuanensis, M. liboensis, M. wushanensis (wide in males); narrow in M. rubrimera), toes with rudimentary webbing (vs. toes without webbing in M. daweimontis, M. rubrimera, M. wuliangshanensis and M. wushanensis (in females); webbing indistinct or absent in M. fansipanensis), tibio-tarsal articulation reaching forward to the region between tympanum and eye when hindlimb is stretched along the side of the body (vs. reaching forward to the tip of snout in M. daweimontis), finger II shortest (vs. finger I shortest in M. liboensis), presence of an indistinct subarticular tubercle on the bases of each finger (vs. subarticular tubercle absent in M. fansipanensis), heels not meeting when the flexed hindlimbs are held at right angles to the body axis (heels meeting in M. binchuanensis; heels meeting or overlapping in M. minor and M. wushanensis; heels overlapping in M. leishanensis, M. liboensis and M. wuliangshanensis).

Megophrys nankunensis sp. nov. further differs from M. feii, for which molecular data are lacking and cannot be allocated to any subgenus base on morphology only (Yang et al. 2018) by the larger body size, SVL 29.9–34.9 mm in males and 39.4–41.9 mm in females (VS. 24.3–25.1 mm in males, 28.2–28.9 mm in females), presence of nuptial pad with nuptial spines in males during breeding season (vs. absent), presence of vomerine teeth (vs. absent), absence of lateral fringes on toes (vs. moderate or wide), heels not meeting when the flexed hindlimbs are held at right angles to the body axis (vs. heels overlapping).

Description of holotype

Adult male. Habitus small, SVL 31.3 mm; head width slightly larger than head length, HDW/HDL 1.12; snout rounded in dorsal view, tip of snout slightly sharpened, sloping backward to mouth in profile, protruding well beyond margin of lower jaw; top of head flat; eye large, ED/HDL 0.38; nostril oblique ovoid; pupil vertical; canthus rostralis well developed, forming the beginning of a fleshy, protruding ridge, that continues over the upper eyelid, and transitions into a supratympanic fold that terminates in the scapular region; loreal region vertical; internasal distance slightly larger than interorbital distance; tympanum distinct, moderate-sized, TD/ED 0.44; large ovoid choanae at the base of the maxilla; strong vomerine ridge bearing vomerine teeth; margin of tongue weakly notched posteriorly; internal vocal slits present near the rear of the lower mandible.

RAD/SVL 0.22, HND/SVL 0.22; absence of lateral fringes and webbing on fingers, relative finger lengths II < I < IV < III; tip of finger rounded, slightly swollen; presence of a distinct subarticular tubercle on the base of each finger; outer metacarpal tubercles indistinct, inner metacarpal tubercles distinct and observably enlarged. Hindlimbs short, tibio-tarsal articulation reaching forward the anterior margin of tympanum when hindlimb is stretched along the side of the body; heels not meeting when the flexed hindlimbs are held at right angles to the body axis; TIB/SVL 0.37 and FTL/SVL 0.55; relative toe lengths I < II < V < III < IV; tips of toes round and slightly dilated; presence of rudimentary webbing on toes but absence of lateral fringes and tarsal folds; presence of a subarticular tubercle only at the bases of each toes; presence of a long ovoid inner metatarsal tubercle and absence of outer metatarsal tubercle.

Dorsal skin texture smooth with dense granules, some of which forming a weak X-shaped skin ridge on center of trunk; surface of flanks with large tubercles; presence of a small horn-like tubercle at the edge of eyelid; distinct supratympanic fold curving posteroventrally from posterior corner of eye to a level above insertion of arm, forming a swollen supraaxillary gland above insertion of arm; ventral skin texture smooth with granules on the surface of abdomen; pectoral gland large, equal size to tip of fingers, closer to axilla; single large femoral gland on rear of thigh.

Measurements of holotype (in mm)

SVL 31.3, HDL 9.6, HDW 10.8, SNT 3.4, IND 3.4, IOD 2.4, ED 3.7, TD 1.6, TED 1.0, HND 6.9, RAD 7.0, FTL 17.3, TIB 11.6.

Coloration of holotype in life

(Fig. 4A–D) Dorsal surface beige with obscure darker patches, with a distinct and incomplete dark triangular marking between eyes, unconnected with an incomplete X-shaped marking on center of trunk. Forearm with dark bands dorsally; hindlimb with broad black transverse bands. Tip of snout dark brown. A dark brown vertical band below the eye. Supratympanic fold white. Horn-like tubercle at the edge of the upper eyelid orange. Surface of throat and chest dark brown, with scarlet spots. Posterior region of abdomen white, with dark brown and scarlet spots. Ventral surface of limbs white with brown patches. Ventral surface of hand and foot light brown, subarticular tubercle at the base of each fingers and toes, outer metacarpal tubercle, inner metatarsal tubercle and inner metacarpal tubercle pink. Pectoral and femoral glands white. Iris white.

Figure 4. 

Megophrys nankunensis sp. nov. in life: A–D SYS a004498, the male holotype E–F SYS a004507, the female paratype.

Coloration of holotype in preservative

On dorsal surface the beige fades to dark grey. Dark interorbital triangular marking becomes more indistinct. Ventral surface pale in color, grey-brownish grounding, markings and mottling more distinct, all scarlet spots absent.

Variation

Measurements and body proportions of type series are given in Table 4.

All paratype specimens were very similar in morphology and color pattern. However, the holotype has the dorsal surface beige (vs. reddish brown in paratypes SYS a002033, 4501, and dark brown in paratypes SYS a004502–4506, 4507 (Fig. 4E–F)), dorsal skin texture smooth, granules and tubercles weak (vs. dorsal skin texture relatively rough with more distinct granules and tubercles in paratypes SYS a004502, 4504–4507), and ventral surface of hand and foot light brown (vs. ventral surface of hand and foot grey white in paratypes SYS a004502–4504).

Etymology

The specific epithet “nankunensis” is in reference to the type locality of the new species: Mt. Nankun. We propose the common English name “Nankunshan Horned Toad” and Chinese name “Nan Kun Shan Jiao Chan (南昆山角蟾)”.

Distribution and habits

Currently, Megophrys nankunensis sp. nov. is known only from the type locality, Mt. Nankun in Longmen County, Guangdong Province, China. It inhabits forest floor, leaf litter and the nearby undergrowth rocky mountainous streams (2–3 m wide) surrounded by moist subtropical evergreen broadleaved forests at elevations between 300–600 m. Breeding season of M. nankunensis sp. nov. is from October to the following March, males were found calling under the leaf litter or rocks (Fig. 5A) on the ground in the flowing streams, besides, a pair were observed exposed on the floor in a flowing stream, about 2.5 m wide, prior to amplexus (Fig. 5B) at 20:09 P.M. on 20 October 2015. Tadpoles were not observed in this period.

Figure 5. 

Ecology and behavior of Megophrys nankunensis sp. nov. A an adult male observed under the rock in the flowing stream B pair of M. nankunensis sp. nov. observed exposed on leave litters in a flowing stream, about 2.5 m wide, prior to amplexus.

Megophrys (Panophrys) jiulianensis J. Wang, Zeng, Lyu & Y.Y. Wang, sp. nov.

Fig. 6, Table 6

Holotype

SYS a002112, adult male, collected by Yu-Long Li on 2 May 2013 from Daqiutian Protection Station (24°34'34.99"N, 114°26'28.53"E; 560 m a.s.l.) of Mt. Jiulian, Longnan County, Ganzhou City, Jiangxi Province, China.

Paratypes (nine males & two females)

SYS a002110, 2111, adult females, collected by Yu-Long Li on 3 May 2013 from Xiagongtang Protection Station (24°32'16.74"N, 114°27'56.82"E; 770 m a.s.l.) of Mt. Jiulian; SYS a001007, 1009, adult males, collected by Run-Lin Li on 23 July 2010 from Daqiutian Protection Station of Mt. Jiulian; SYS a002107–2109, 2113–2114, SYS a002115/CIB110008, adult males, collected by Yu-Long Li on 1–4 May 2013 from Xiagongtang Protection Station and Daqiutian Protection Station of Mt. Jiulian at elevations between 400–800 m a.s.l.; SYS a002031, adult male, collected by Run-Lin Li on 20 Marth 2013 from Mt. Nankun (23°38'21.94"N, 113°50'39.49"E; 610 m a.s.l.), Longmen County, Huizhou City, Guangdong Province, China.

Diagnosis

(1) Body slender and small-sized, SVL 30.4–33.9 mm in nine adult males, 34.1–37.5 mm in two adult females; (2) head width slightly larger than head length, HDW/HDL ratio 1.04–1.06; (3) snout rounded in dorsal view; (4) eye large, tympanum distinct, moderate-sized, TD/ED ratio 0.50–0.59; (5) weak vomerine ridge bearing vomerine teeth; (6) tongue weakly notched posteriorly; (7) hindlimbs slender, heels overlapping when the flexed hindlimbs are held at right angles to the body axis, tibia-tarsal articulation reaching forward to the middle of eye when hindlimb is stretched along the side of the body; (8) absence of lateral fringes on fingers, presence of an indistinct subarticular tubercle on the bases of each finger, relative finger lengths II < I < IV < III; (9) toes with rudimentary webbing at their bases and without lateral fringes, subarticular tubercles only present at the base of toe I and II; (10) dorsal skin rough, presence of black spines on granules of dorsal skin, and occasionally present on canthus rostralis and margin of tympanum, presence of large tubercles on flanks, dorsal body and limbs; (11) four prominent parallel dorsolateral ridges with granules bearing black spines on back of trunk, the middle two ridges forming a X-shaped ridge occasionally; (12) a reddish horn-like tubercle bearing a black spine at its tip at the edge of eye lid; (13) distinct supratympanic fold bearing black spines; (14) beige to brownish red above, with an hollow dark triangle between eyes and a rectangular dark marking on the center of the back of trunk; (15) males with a single subgular vocal sac, and presence of nuptial pads bearing darker nuptial spines on dorsal surface of the first and second fingers in adult males during breeding season, respectively; (16) gravid females bear creamy yellow oocytes.

Comparisons

Comparative data of Megophrys jiulianensis sp. nov. with M. dongduanensis sp. nov., M. nankunensis sp. nov., M. feii and the 33 recognized members of the Megophrys s.l. allocated to the subgenus Panophrys are listed in Table 5.

Megophrys jiulianensis sp. nov. is sympatric with M. nankunensis sp. nov. in Mt. Nankun, but it can be easily distinguished from the later by heels overlapping when the flexed hindlimbs are held at right angles to the body axis (vs. heels not meeting), TIB/SVL ratio 0.61–0.68 (vs. TIB/SVL ratio 0.35–0.42), supratympanic fold not forming a supraaxillary gland above insertion of arm (vs. supratympanic fold forming a depressed supraaxillary gland), presence of black spines on dorsal skin (vs. absent); besides, M. jiulianensis sp. nov. differs from M. dongguanensis sp. nov. by the notched tongue vs. (not notched), heels overlapping when the flexed hindlimbs are held at right angles to the body axis (vs. heels not meeting), TIB/SVL ratio 0.61–0.68 (vs. TIB/SVL ratio 0.41–0.46).

With significantly smaller body size, SVL 30.4–33.9 mm in males and 34.1–37.5 mm in females, M. jiulianensis sp. nov. differs from the 17 members with larger SVL values: M. baolongensis (42.0–45.0 mm in males), M. binchuanensis (40.2–42.5 mm in females), M. binlingensis (45.1–51.0 mm in males), M. caudoprocta (81.3 mm in single male), M. daweimontis (40.0–46.0 mm in females), M. fansipanensis (41.7–42.5 mm in females), M. hoanglienensis (37.4–47.6 mm in males, 59.6 mm in single female), M. jingdongensis (53.0–56.5 mm in males, 63.5 mm in single female), M. liboensis (34.7–67.7 mm in males, 60.8–70.6 mm in females), M. minor (34.5–41.2 mm in males), M. omeimontis (56.0–59.5 mm in males, 68.0–72.5 mm in females), M. palpebralespinosa (36.2–38.0 mm in males), M. sangzhiensis (54.7 mm in single male), M. shuichengensis (102.0–118.3 mm in males, 99.8–115.6 mm in females), M. spinata (47.2–54.4 mm in males, 54.0–55.0 mm in females), M. tuberogranulatus (50.5 mm in single female) and M. wuliangshanensis (41.3 mm in single female).

Megophrys jiulianensis sp. nov. differs from 12 species occurring in eastern and southern China (M. acuta, M. brachykolos, M. boettgeri, M. cheni, M. huangshanensis, M. insularis, M. jinggangensis, M. kuatunensis, M. lini, M. lishuiensis, M. obesa and M. ombrophila) by the following combination of characters: presence of vomerine teeth (vs. absent in M. acuta, M. boettgeri, M. brachykolos, M. cheni, M. huangshanensis, M. jinggangensis, M. kuatunensis, M. lini, M. lishuiensis, M. obesa and M. ombrophila), tongue notched posteriorly (vs. not notched in M. acuta, M. brachykolos, M. jinggangensis, M. lini, M. lishuiensis, M. obesa and M. ombrophila), absence of lateral fringes on toes (vs. narrow in M. acuta, M. jinggangensis and M. kuatunensis; wide in M. boettgeri, M. cheni and M. lini), heels overlapping when the flexed hindlimbs are held at right angles to the body axis (vs. heels not meeting in M. acuta, M. brachykolos, M. insularis, M. obesa and M. ombrophila).

Megophrys jiulianensis sp. nov. differs from the remaining four members of the Megophrys s.l. allocated to the subgenus Panophrys which share a moderate or small body size, by the presence of vomerine teeth (vs. absent in M. leishanensis and M. wushanensis), tongue notched posteriorly (vs. not notched in M. leishanensis, M. wushanensis and M. latidactyla), absence of lateral fringes on toes (vs. narrow in M. rubrimera; wide in M. latidactyla and M. wushanensis (wide in females)), toe webbing rudimentary (vs. absence of webbing on toes in M. rubrimera).

Megophrys jiulianensis sp. nov. further differs from M. feii, for which molecular data are lacking and cannot be allocated to any subgenus base on morphology only (Yang et al. 2018) by the larger body size, SVL 30.4–33.9 mm in males and 34.1–37.5 mm in females (VS. 24.3–25.1 mm in males, 28.2–28.9 mm in females), presence of nuptial pad with nuptial spines in males during breeding season (vs. absent), presence of vomerine teeth (vs. absent), absence of lateral fringes on toes (vs. moderate or wide).

Description of holotype

Adult male. Habitus slender and small, SVL 32.0 mm; head width slightly larger than head length, HDW/HWL 1.04; snout rounded in dorsal view, projecting, sloping backward to mouth in profile, protruding well beyond margin of lower jaw; top of head flat; eye large, ED/HDL 0.39; nostril oblique ovoid; pupil vertical; canthus rostralis well developed, forming the beginning of a fleshy, protruding ridge, that continues over the upper eyelid, and transitions into a supratympanic fold that terminates in the scapular region; loreal region vertical; internasal distance slightly larger than interorbital distance; tympanum distinct, moderate-sized, TD/ED 0.52; large ovoid choanae at the base of the maxilla; weak vomerine ridge bearing vomerine teeth; margin of tongue weakly notched posteriorly; internal vocal slits present near the rear of the lower mandible..

RAD/SVL 0.25; absence of lateral fringes and webbing on fingers, relative finger lengths II < I < IV < III; tip of finger rounded, slightly swollen; presence of an indistinct subarticular tubercle on the base of each finger; outer metacarpal tubercles indistinct, inner metacarpal tubercles distinct and observably enlarged. Hindlimbs long, tibio-tarsal articulation reaching forward to the middle of eye when hindlimb is stretched along the side of the body; heels overlapping when the flexed hindlimbs are held at right angles to the body axis; TIB/SVL 0.46 and FTL/SVL 0.62; relative toe lengths I < II < V < III < IV; tips of toes round and slightly dilated; presence of rudimentary webbing on toes but absence of lateral fringes and tarsal folds; presence of a subarticular tubercle only at the bases of the first and second toes; presence of a long ovoid inner metatarsal tubercle and absence of outer metatarsal tubercle.

Dorsum rough with dense granules bearing spines; canthus rostralis, margin of tympanum, supratympanic fold and upper lip with dense spines; presence of large tubercles bearing spines on dorsal surface of body, surface of flanks and dorsal and posterolateral surface of limbs; prominent parallel dorsolateral ridges with granules bearing spines on back of trunk; presence of a horn-like tubercle bearing a spine at its tip at the edge of eye lid; distinct supratympanic fold curving posteroventrally from posterior corner of eye to a level above insertion of arm; ventral skin texture smooth, the lower lip bears spines; sides of belly with large tubercles; ventral skin texture of thighs smooth with a few small tubercles, posterior surface and surface around anus with large tubercles bearing spines; surface of tibia-tarsal with a few tubercles bearing spines; presence of spines on lateral sides of fingers and toes; pectoral gland moderate-sized, closer to axilla; single femoral gland on rear of thigh, distinctly smaller than pectoral gland.

Measurements of holotype (in mm)

SVL 32.2, HDL 11.5, HDW 11.4, SNT 3.6, IND 3.5, IOD 3.3, ED 4.2, TD 2.3, TED 1.7, HND 8.0, RAD 8.1, FTL 20.5, TIB 14.7.

Coloration of holotype in life

(Fig. 6A–D) Dorsal surface yellowish brown, with an incomplete dark triangular marking between eyes. Spines on dorsal surface, granules and tubercles black. Forearm with a distinct, black oblique band. Transverse bands on hindlimb indistinct. Tip of snout grayish brown. A grayish-brown vertical band below the eye. Tubercle at the edge of the upper eyelid red. Ventral surface yellow, scattered with dense dark gray spots and black scarlet blotches; ventral surface of limbs flesh colored with pink and black spots. Palms and soles dark brown, inner metatarsal tubercle, outer metacarpal tubercle and inner metacarpal tubercle orange red, tip of digits orange-red. Pectoral glands and femoral glands white. Iris white.

Figure 6. 

General life aspect in life of Megophrys jiulianensis sp. nov.: A–D SYS a002112, the male holotype E–F SYS a002111, the female paratype.

Coloration of holotype in preservative

Dorsum yellowish brown fades to greyish brown, scattered with black spots. Greyish black triangular marking between the eyes become more distinct. Ventral surface paled in color, brown grounding, markings and mottling become more distinct.

Variation

Measurements and body proportions of type series are given in Table 6.

All paratype specimens were very similar in morphology and color pattern. However, dorsal skin texture is more rough with well-developed spines in the female specimen SYS a002111 (Fig. 6E–F), dorsal surface yellowish brown in the other female specimen SYS a002110, and the middle two ridges on dorsum forming an X-shape skin ridge in the male specimen SYS a002108.

Measurements (in mm; minimum-maximum, mean ± SD) of the type series of Megophrys jiulianensis sp. nov.

Species Megophrys jiulianensis sp. nov.
Males (n = 9) Females (n = 2)
SVL 30.4–33.9 (32.2 ± 1.2) 34.1–37.5
HDL 10.7–11.6 (11.2 ± 0.4) 12.0–12.4
HDW 10.9–11.8 (11.4 ± 0.4) 12.5–13.2
SNT 3.4–3.8 (3.6 ± 0.2) 3.9–4.1
IND 3.2–3.6 (3.5 ± 0.1) 3.5–3.8
IOD 3.2–3.5 (3.3 ± 0.1) 3.6
ED 3.9–4.4 (4.2 ± 0.2) 4.3–4.4
TD 2.1–2.5 (2.3 ± 0.1) 2.2–2.4
TED 1.6–2.0 (1.7 ± 0.1) 2.1–2.5
HND 7.4–10.6 (8.0 ± 0.4) 8.3–9.5
RAD 7.7–8.5 (8.1 ± 0.3) 8.3–9.8
FTL 14.1–15.2 (14.7 ± 0.4) 16.0–17.8
TIB 19.8–21.1 (20.5 ± 0.5) 21.6–25.5
HDL/SVL 0.34–0.37 (0.35 ± 0.01) 0.33–0.35
HDW/SVL 0.34–0.37 (0.35 ± 0.01) 0.35–0.37
HDW/HDL 1.00–1.04 (1.02 ± 0.02) 1.04–1.06
SNT/HDL 0.32–0.34 (0.32 ± 0.01) 0.33
SNT/SVL 0.11–0.12 (0.11 ± 0.00) 0.11
IND/HDW 0.29–0.33 (0.30 ± 0.01) 0.28–0.29
IOD/HDW 0.28–0.30 (0.29 ± 0.01) 0.27–0.29
ED/HDL 0.36–0.39 (0.38 ± 0.01) 0.35–0.36
ED/SVL 0.12–0.14 (0.13 ± 0.01) 0.12–0.13
TD/ED 0.50–0.59 (0.55 ± 0.03) 0.51–0.55
TED/TD 0.68–0.87 (0.75 ± 0.07) 0.95–1.04
HND/SVL 0.24–0.26 (0.25 ± 0.01) 0.24–0.25
RAD/SVL 0.24–0.27 (0.25 ± 0.01) 0.24–0.26
TIB/SVL 0.44–0.48 (0.46 ± 0.01) 0.47
FTL/SVL 0.61–0.67 (0.64 ± 0.02) 0.63–0.68

Etymology

The specific epithet “jiulianensis” is in reference to the known localities of the new species: Mt. Jiulian and Nankunshan Natuire Reserve located in the Jiulian Mountains range. We propose the common English name “Jiulianshan Horned Toad” and Chinese name “Jiu Lian Shan Jiao Chan (九连山角蟾)”.

Distribution and natural history

Currently, Megophrys jiulianensis sp. nov. is known from Mt. Nankun in Guangdong Province and the type locality, Jiulian Nature Reserve in Jiangxi Province, China. It inhabits forest floor, leaf litter and the nearby undergrowth mountainous streams surrounded by moist subtropical evergreen broadleaved forests at elevations between 500–800 m. Breeding season of M. jiulianensis sp. nov. is from March to July, males were usually found staying while calling on leaves (Fig. 7A), about 0.1–0.3 m above the ground. After the rain, numerous individuals can be easily found on the road, and a female individual from Mt. Nankun was observed feeding on an earthworm (Fig. 7B) on 20:45 p.m., 21 March 2016. Tadpoles could be found all year round.

Figure 7. 

Ecology and behavior of Megophrys jiulianensis sp. nov.: A The male paratype SYS a002031 observed calling on a leaf (showing subgular vocal sac) B a female individual observed feeding on an earthworm after rain, both from Mt. Nankun in Guangdong Province.

Megophrys jiulianensis sp. nov. is sympatric with M. nankunensis sp. nov. and M. mangshanensis at Mt. Nankun.

Megophrys (Panophrys) nanlingensis Lyu, J. Wang, Liu & Y.Y. Wang, sp. nov.

Fig. 8, Table 7

Holotype

SYS a001964, adult male, collected by Run-Lin Li on 21 December 2012 from Nanling Nature Reserve (24°54'48.80"N, 113°01'12.34"E; 1008m a.s.l.), Ruyuan County, Shaoguan City, Guangdong Province, China.

Paratypes (nine males)

SYS a001959–1962, SYS a001963/CIB110010, adult males, collected on 21 December 2012 by Run-Lin Li from the same stream as the holotype (1000–1300 m a.s.l.); SYS a002334, 2356–2358, collected on 1–3 October 2013 by Ying-Yong Wang and Zu-Yao Liu from Mt. Qiyun (25°52'22.84"N, 114°01'52.09"E; 691–1355m a.s.l.), Chongyi County, Ganzhou City, Jiangxi Province, China.

Diagnosis

(1) Body small-sized, SVL 30.5–37.3 mm in 10 adult males; (2) snout rounded in dorsal view; (3) tympanum distinct, moderate-sized, TD/ED ratio 0.43–0.57; (4) vomerine ridge and vomerine teeth present; (5) tongue notched posteriorly; (6) absence of lateral fringes and webbing on fingers, presence of narrow lateral fringes and rudimentary webbing on toes; (7) presence of a subarticular tubercle at the base of each finger and toe; (8) hindlimbs slender, heels overlapping, tibio-tarsal articulation reaching between the posterior corner to the center of eye; (9) TIB/SVL ratio 0.45–0.51 and FTL/SVL ratio 0.61–0.73; (10) dense conical granules present on surface of temporal region, upper lip, and from loreal region to the tip of snout; (11) granules and tubercles on dorsal surface forming a discontinuous X-shaped ridge and a pair of discontinuous dorsolateral ridges on back of trunk; (12) supratympanic fold distinct, whitish tan; (13) brown dorsally, with a dark triangular marking with light yellow edge between eyes, and an X-shaped or V-shaped marking with light yellow edge on the center of the back of trunk; (14) presence of a single subgular vocal sac in males; (15) nuptial pads and nuptial spines invisible in males during breeding season.

Comparisons

Comparative data of Megophrys nanlingensis sp. nov. with M. dongduanensis sp. nov., M. nankunensis sp. nov., M. jiulianensis sp. nov., M. feii and the 33 recognized members of Megophrys s.l. allocated to the subgenus Panophrys are listed in Table 5.

Megophrys nanlingensis sp. nov. differs from M. dongguanensis sp. nov., M. nankunensis sp. nov. and M. jiulianensis sp. nov. by the heels overlapping when hindlimb is stretched along the side of the body (vs. heels not meeting in M. dongguanensis sp. nov. and M. nankunensis sp. nov.), presence of lateral fringes on toes (vs. absent in M. dongguanensis sp. nov., M. nankunensis sp. nov. and M. jiulianensis sp. nov.), tongue notched posteriorly (vs. not notched in M. dongguanensis sp. nov. and M. nankunensis sp. nov.), skin relatively smooth and lacking black horny spines (vs. skin rough with black horny spines in M . jiulianensis sp. nov.).

With the smaller body size, SVL 30.5–37.3 mm in males, Megophrys nanlingensis sp. nov. differs from the nine members with larger SVL values: M. baolongensis (42.0–45.0 mm in males), M. binlingensis (45.1–51.0 mm in males), M. caudoprocta (81.3 mm in single male), M. jingdongensis (53.0–56.5 mm in males), M. latidactyla (38.9 mm in single male), M. omeimontis (56.0–59.5 mm in males), M. sangzhiensis (54.7 mm in single male), M. shuichengensis (102.0–118.3 mm in males) and M. spinata (47.2–54.4 mm in males).

Megophrys nanlingensis sp. nov. differs from 12 species occurring in eastern and southern China (M. acuta, M. brachykolos, M. boettgeri, M. cheni, M. huangshanensis, M. insularis, M. jinggangensis, M. kuatunensis, M. lini, M. lishuiensis, M. obesa and M. ombrophila) by the following combination of characters: presence of vomerine teeth (vs. absent in M. acuta, M. boettgeri, M. brachykolos, M. cheni, M. huangshanensis, M. kuatunensis, M. lini, M. lishuiensis, M. obesa and M. ombrophila), margin of tongue notched posteriorly (vs. not notched in M. acuta, M. brachykolos, M. jinggangensis, M. lini, M. lishuiensis, M. obesa and M. ombrophila), toes with narrow lateral fringes (vs. wide in M. boettgeri, M. cheni and M. lini; absent in M. brachykolos, M. huangshanensis, M. insularis, M. lishuiensis, M. obesa and M. ombrophila), toes with rudimentary webbing (vs. toes without webbing in M. lishuiensis, M. kuatunensis and M. ombrophila), hindlimbs comparatively longer, with heels overlapping when the flexed hindlimbs are held at right angles to the body axis (vs. hindlimbs short, with heels not meeting in M. acuta, M. brachykolos, M. huangshanensis, M. insularis, M. obesa and M. ombrophila).

Megophrys nanlingensis sp. nov. differs from the remaining 12 members of the Megophrys s.l. allocated to the subgenus Panophrys which share a moderate or small body size, by the small horn-like tubercle at edge of the upper eyelid (vs. horn-like tubercle indistinct or absent in M. binchuanensis, M. minor, M. wuliangshanensis and M. wushanensis; slightly large in M. palpebralespinosa; long point in M. liboensis), presence of vomerine teeth (vs. absent in M. binchuanensis, M. leishanensis, M. minor, M. wuliangshanensis and M. wushanensis), tongue notched posteriorly (vs. tongue not notched in M. palpebralespinosa, M. tuberogranulatus and M. wushanensis), toes with narrow lateral fringes (vs. wide in M. binchuanensis, M. liboensis, M. palpebralespinosa and M. wushanensis (in males)); absent in M. daweimontis, M. leishanensis, M. minor, M. tuberogranulatus, M. wuliangshanensis, M. wushanensis (in females); indistinct or absent in M. hoanglienensis), toes webbing rudimentary (vs. toes without webbing in M. daweimontis, M. fansipanensis, M. rubrimera and M. wuliangshanensis; indistinct or absent in M. fansipanensis and M. hoanglienensis; at least one-fourth webbed in M. palpebralespinosa), subarticular tubercles present (vs. absent in M. palpebralespinosa and M. rubrimera).

Megophrys nanlingensis sp. nov. further differs from M. feii, for which molecular data are lacking and cannot be allocated to any subgenus based on morphology only (Yang et al. 2018) by the larger body size, SVL 30.5–37.3 mm in males (VS. 24.3–25.1 mm in males), presence of nuptial pad with nuptial spines in males during breeding season (vs. absent), presence of vomerine teeth (vs. absent), presence of narrow lateral fringes on toes (vs. moderate or wide).

Description of holotype

Adult male. Body size small, SVL 32.5 mm; head length and head width almost isometric, HDW/HDL 0.99; snout rounded in dorsal view, projecting, sloping backward to mouth in profile, protruding well beyond margin of lower jaw; top of head flat; eye large, ED/HDL 0.37, pupil vertical; nostril oblique ovoid; canthus rostralis well developed; loreal region slightly oblique; internasal distance slightly larger than interorbital distance; tympanum distinct, moderate-sized, TD/ED 0.48; large ovoid choanae at the base of the maxilla; presence of vomerine ridge bearing vomerine teeth; margin of tongue notched posteriorly; internal vocal slits present near the rear of the lower mandible.

RAD/SVL 0.25, HND/SVL 0.24; fingers without webbing and lateral fringes, relative finger length II < I < IV < III; tips of fingers slightly dilated, round; one subarticular tubercle at the bases of each finger; outer and inner metacarpal tubercles distinct, and the inner one observably enlarged. Hindlimbs slender, tibio-tarsal articulation reaching forward to the center of the eye when hindlimb is stretched along the side of the body; heels overlapping when the flexed hindlimbs are held at right angles to the body axis; TIB/SVL 0.49 and FTL/SVL 0.69; relative toe length I < II < V < III < IV; tips of toes round and slightly dilated; toes with narrow lateral fringes, rudimentary webbing; one subarticular tubercle at the bases of each toes; presence of a long ovoid inner metatarsal tubercle and absence of outer metatarsal tubercle.

Dorsal skin texture rough; head surface rough, with small tapered granules densely covering from temporal region, upper lip, loreal region to tip of snout; granules forming discontinuous X-shaped ridge with two discontinuous dorsolateral ridges on both sides at the central trunk; large tubercles on flanks; a horn-like prominent tubercle on the edge of the upper eyelid; distinct supratympanic fold curving posteroventrally from posterior corner of eye to a level above insertion of arm; ventral skin texture smooth, with several large granules and tubercles on two sides; ventral skin texture of thighs smooth, with a few small tubercles; pectoral gland larger, closer to axilla; single femoral gland on rear of thigh.

Measurements of holotype (in mm)

SVL 32.5, HDL 11.5, HDW 11.4, SNT 3.7, IND 3.5, IOD 3.3, ED 4.2, TD 2.0, TED 1.7, HND 8.0, RAD 7.8, FTL 22.3, TIB 15.9.

Coloration of holotype in life

(Fig. 8A–D) Brown dorsally, with a dark triangular marking with light yellow edge between eyes, and an X-shaped marking with light yellow edge on the center of the back of trunk. Dark brown transverse bands dorsally on lower arms and hindlimbs. Surface of snout brown. Black brown vertical band below the eye on each side. Temporal region brown, supratympanic fold white. Ventral surface pale grey, an indistinct longitudinal stripe on surface of throat. Scarlet spots on surface of chest. Belly whitish grey with dark brown marbling. A pair of black longitudinal stripes scattered with several white tubercles on surface of lateroventral flanks. Ventral surface of limbs light red and scattered with white spots. Ventral surface of hands and feet dark brown, tips of digits pale-grey. Metacarpal tubercle and metatarsal tubercle light red. Pectoral glands and femoral glands white. Iris reddish brown.

Figure 8. 

Megophrys nanlingensis sp. nov. in life: A–D SYS a001964, male holotype E, F SYS a001963, female paratype.

Coloration of holotype in preservative

Coloration of dorsal and ventral surface turned pale; transverse bands on limbs, dark longitudinal stripe on surface of throat and black patches on surface of lateroventral flanks became more distinct; scarlet spots on surface of chest faded.

Variation

Measurement data of type series are listed in Table 7.

All paratypes are very similar to holotype in morphology and color pattern. However, the male specimen SYS a001963 (Fig. 8E, F) is obviously large in snout-vent length than other specimens, with lighter reddish-brown iris, yellowish brown background coloration and comparatively smooth skin. The heels are significantly overlapping in all specimens from Nanling Nature Reserve but slightly overlapping in specimens from Mt. Qiyun.

Measurements (in mm; minimum-maximum, mean ± SD) of the type series of Megophrys nanlingensis sp. nov.

Species Megopgrys nanlingensis sp. nov.
Males (n = 10)
SVL 30.5–37.3 (33.2 ± 1.9)
HDL 10.9–12.7 (11.6 ± 0.5)
HDW 10.7–13.8 (11.8 ± 0.9)
SNT 3.4–3.8 (3.6 ± 0.1)
IND 3.5–4.0 (3.7 ± 0.2)
IOD 3.2–4.0 (3.4 ± 0.3)
ED 4.1–4.9 (4.5 ± 0.3)
TD 1.9–2.5 (2.2 ± 0.2)
TED 1.6–2.2 (1.8 ± 0.2)
HND 7.1–9.6 (8.0 ± 0.6)
RAD 7.1–9.0 (8.1 ± 0.5)
FTL 18.6–27.1 (22.4 ± 2.3)
TIB 13.9–18.8 (16.0 ± 1.3)
HDL/SVL 0.33–0.36 (0.35 ± 0.01)
HDW/SVL 0.33–0.37 (0.35 ± 0.01)
HDW/HDL 0.97–1.09 (1.02 ± 0.04)
SNT/HDL 0.30–0.33 (0.31 ± 0.01)
SNT/SVL 0.10–0.12 (0.11 ± 0.01)
IND/HDW 0.29–0.35 (0.31 ± 0.02)
IOD/HDW 0.28–0.32 (0.29 ± 0.01)
ED/HDL 0.37–0.41 (0.39 ± 0.01)
ED/SVL 0.13–0.14 (0.14 ± 0.01)
TD/ED 0.43–0.57 (0.48 ± 0.04)
TED/TD 0.67–0.95 (0.83 ± 0.10)
HND/SVL 0.23–0.26 (0.24 ± 0.01)
RAD/SVL 0.23–0.26 (0.24 ± 0.01)
TIB/SVL 0.45–0.51 (0.48 ± 0.02)
FTL/SVL 0.61–0.73 (0.68 ± 0.04)

Etymology

The specific epithet “nanglingensis” is in reference to the type locality of the new species, Nanling Nature Reserve of the Nanling Mountains. We propose the common English name “Nanling Horned Toad” and Chinese name “Nan Ling Jiao Chan (南岭角蟾)”.

Distribution and natural history

Currently, Megophrys nanglingensis sp. nov. is known from Nanling Nature Reserve and the neighboring Mangshan Nature Reserve (between elevations of 1000–1300 m), together with Mt. Qiyun (between elevations of 690–1400 m). It inhabits streams in bamboo forests. Males are frequently heard calling during August and December. Tadpoles could be found in this period.

Megophrys nanlingensis sp. nov. is sympatric with M. mangshanensis and M. popei in Nanling Nature Reserve and the neighboring Mangshan Nature Reserve.

Megophrys (Panophrys) wugongensis J. Wang, Lyu & Y.Y. Wang, sp. nov.

Fig. 9, Table 8

Holotype

SYS a002625, adult male, collected by Guo-Ling Chen and Jian Zhao on 9 May 2014 from Yangshimu Scenic Area (27°34'47.93"N, 114°15'7.34"E; 550 m a.s.l.), Pingxiang City, Jiangxi Province, China.

Paratypes (three males & nine females)

Adult males, SYS a004777/CIB110011, SYS a004796, 4800, collected by Zhi-Tong Lyu and Ying-Yong Wang on 23 May 2016, and adult females, SYS a002610–2611, collected by Guo-Ling Chen and Jian Zhao on 8 May 2014, SYS a004797–4799, 4801–4804, collected by Zhi-Tong Lyu and Ying-Yong Wang on 23 May 2016, from Wugongshan Scenic Area (27°34'3.94"N, 114°10'28.38"E; 1050–1080 m a.s.l.), Anfu County, Ji’an City, Jiangxi Province, China.

Diagnosis

(1) Body size small, SVL 31.0–34.1 mm in four adult males and body size moderate, SVL 38.5–42.8 mm in nine adult females; (2) tympanum distinct, slightly convex, moderate-sized, TD/ED ratio 0.47–0.52; (3) vomerine teeth absent; (4) margin of tongue not notched posteriorly; (5) hindlimbs short, heels not meeting, tibia-tarsal articulation reaching forward to the region between posterior corner of eye and posterior margin of tympanum; (6) TIB/SVL ratio 0.39–0.44, FTL/SVL ratio 0.56–0.64; (7) fingers without lateral fringes, presence of a subarticular tubercle at the bases of each finger, relative finger lengths II < I = IV < III; (8) toes with rudimentary webbing at their bases and without lateral fringes, subarticular tubercles only present at the base of each toe; (9) numerous granules present on dorsal surface of body, several large tubercles present on surface of flanks and dorsal surface of limbs; (10) presence of a small horn-like tubercle at the edge of the upper eyelid; (11) supratympanic fold distinct, whitish; (12) yellowish brown or reddish brown dorsally, with an incomplete dark triangular marking between eyes and an X-shaped marking on back of trunk; (13) ventral surface greyish brown, ventral surface of abdomen with creamy white nebulous patches and black spots; (14) males with a single subgular vocal sac; (15) gravid females bear creamy yellow oocytes.

Comparisons

Comparative data of Megophrys wugongensis sp. nov. with M. dongduanensis sp. nov., M. nankunensis sp. nov., M. jiulianensis sp. nov., Megophrys nanlingensis sp. nov., M. feii and the 33 recognized members of Megophrys s.l. allocated to the subgenus Panophrys are listed in Table 5.

Megophrys wugongensis sp. nov. differs from M. dongguanensis sp. nov., M. nankunensis sp. nov., M. jiulianensis sp. nov. and M. nanlingensis sp. nov. by a combination of following characters: vomerine teeth absent (vs. vomerine teeth present), tongue not notched posteriorly (vs. tongue notched in M. jiulianensis sp. nov. and M. nanlingensis sp. nov.), absence of lateral fringes on toes (vs. presence of narrow lateral fringes on toes in M. nanlingensis sp. nov.), heels not meeting when the flexed hindlimbs are held at right angles to the body axis (vs. heels overlapping in M. jiulianensis sp. nov. and M. nanlingensis sp. nov.), absence of black spines on dorsal skin (vs. present in M. jiulianensis sp. nov.), relative finger lengths II < I = IV < III (vs. II < I < IV < III in M. nankunensis sp. nov., M. jiulianensis sp. nov. and M. nanlingensis sp. nov.), ventral surface with creamy white nebulous patches (vs. absence of such patched on ventral surface in M. dongguanensis sp. nov. and M. nankunensis sp. nov.).

With the smaller body size, SVL 31.0–34.1 mm in males and 38.5–42.8 mm in females, Megophrys wugongensis sp. nov. differs from the 13 members with larger SVL values: M. baolongensis (42.0–45.0 mm in males), M. binlingensis (45.1–51.0 mm in males), M. caudoprocta (81.3 mm in single male), M. hoanglienensis (37.4–47.6 mm in males, 59.6 mm in single female), M. jingdongensis (53.0–56.5 mm in males 63.5 in single female), M. latidactyla (38.9 mm in single male), M. liboensis (34.7–67.7 mm in males, 60.8–70.6 mm in females), M. omeimontis (56.0–59.5 mm in males, 68.0–72.5 mm in females), M. palpebralespinosa (36.2–38.0 mm in males), M. sangzhiensis (54.7 mm in single male), M. shuichengensis (102.0–118.3 mm in males, 99.8–115.6 mm in females), M. spinata (47.2–54.4 mm in males, 54.0–55.0 mm in females), and M. tuberogranulatus (33.2–39.6 mm in males, 50.5 mm in single female).

Megophrys wugongensis sp. nov. differs from 12 species occurring in eastern and southern China (M. acuta, M. brachykolos, M. boettgeri, M. cheni, M. huangshanensis, M. insularis, M. jinggangensis, M. kuatunensis, M. lini, M. lishuiensis, M. obesa and M. ombrophila) by the following combination of characters: vomerine teeth absent (vs. present in M. insularis and M. jinggangensis), tongue not notched posteriorly (vs. tongue notched in M. boettgeri, M. huangshanensis, M. kuatunensis and M. insularis), toes without lateral fringes (vs. laterals fringes on toes narrow in M. acuta, M. kuatunensis and M. jinggangensis; wide in M. boettgeri, M. cheni and M. lini), toes with rudimentary webbing (vs. toes without webbing in M. huangshanensis, M. lishuiensis and M. ombrophila), hindlimbs short, with heels not meeting when the flexed hindlimbs are held at right angles to the body axis (vs. hindlimbs comparatively longer, with heels overlapping in M. boettgeri, M. cheni, M. kuatunensis, M. jinggangensis and M .lini), relative finger lengths II < I = IV < III (vs. I < II ≤ IV < III in M. acuta and M. ombrophila; IV < II < I < III in M. brachykolos; I < II = IV < III in M. lishuiensis; I < II ≤ IV < III in M. obesa), males bearing nuptial pads with nuptial spines during breeding season (vs. nuptials absence in adult males of M. acuta), ventral surface with creamy white nebulous patches (vs. absence of such patched in M. brachykolos and M. obesa).

Megophrys nanlingensis sp. nov. differs from the remaining eight members of the Megophrys s.l. allocated to the subgenus Panophrys which share a moderate or small body size, by a combination of following characters: horn-like tubercle small at edge of the upper eyelid (vs. horn-like tubercle indistinct or absent in M. binchuanensis, M. minor, M. wuliangshanensis and M. wushanensis), absence of vomerine teeth (vs. present in M. daweimontis, M. fansipanensis and M. rubrimera), tongue not notched posteriorly (vs. tongue notched in M. minor, M. fansipanensis and M. rubrimera), toes without lateral fringes (vs. lateral fringes wide in M. binchuanensis, M. wushanensis (in males); narrow in M. rubrimera), toes with rudimentary webbing (vs. toes without webbing in M. daweimontis, M. fansipanensis, M. rubrimera and M. wuliangshanensis), heels not meeting when the flexed hindlimbs are held at right angles to the body axis (vs. heels overlapping in M. minor and M. wuliangshanensis), heels not meeting when the flexed hindlimbs are held at right angles to the body axis (heels meeting in M. binchuanensis; heels meeting or overlapping in M. minor and M. wushanensis; heels overlapping in M. leishanensis, and M. wuliangshanensis).

Megophrys wugongensis sp. nov. further differs from M. feii, for which molecular data are lacking and cannot be allocated to any subgenus base on morphology only (Yang et al. 2018) by the larger body size, SVL 31.0–34.1 mm in males and 38.5–42.8 mm in females (VS. 24.3–25.1 mm in males, 28.2–28.9 mm in females), presence of nuptial pad with nuptial spines in males during breeding season (vs. absent), absence of lateral fringes on toes (vs. moderate or wide).

Description of holotype

Adult male. Habitus small, SVL 31.0 mm; head width slightly larger than head length, HDW/HWL 1.03; snout rounded in dorsal view, tip of snout slightly sharpened, sloping backward to mouth in profile, protruding well beyond margin of lower jaw; top of head flat; eye large, ED/HDL 0.41; nostril oblique ovoid; pupil vertical; canthus rostralis well developed; loreal region vertical; internasal distance slightly larger than interorbital distance; tympanum distinct, moderate-sized, TD/ED 0.47; large ovoid choanae at the base of the maxilla; weak vomerine ridge present, vomerine teeth absent; margin of tongue not notched posteriorly; internal vocal slits present near the rear of the lower mandible.

RAD/SVL 0.24, HND/SVL 0.22; absence of lateral fringes and webbing on fingers, relative finger lengths II < I = IV < III; tip of finger rounded, slightly swollen; presence of a distinct subarticular tubercle on the base of each finger; outer metacarpal tubercles indistinct, inner metacarpal tubercles distinct and observably enlarged. Hindlimbs short, tibio-tarsal articulation reaching forward the posterior corner of eye when hindlimb is stretched along the side of the body; heels not meeting when the flexed hindlimbs are held at right angles to the body axis; TIB/SVL 0.43 and FTL/SVL 0.61; relative toe lengths I < II < V < III < IV; tips of toes round and slightly dilated; presence of rudimentary webbing on toes but absence of lateral fringes and tarsal folds; presence of a subarticular tubercle only at the bases of each toes; presence of a long ovoid inner metatarsal tubercle and absence of outer metatarsal tubercle.

Dorsal skin texture rough with dense granules, some of which forming an X-shaped skin ridge on center of trunk; surface of flanks with large tubercles; presence of a small horn-like tubercle at the edge of eye lid; distinct supratympanic fold curving posteroventrally from posterior corner of eye to a level above insertion of arm; superior margin of tympanum in connect with supratympanic fold; ventral skin texture smooth with granules on the surface of abdomen; pectoral gland large, closer to axilla; single large femoral gland on rear of thigh.

Measurements of holotype (in mm)

SVL 30.8, HDL 11.9, HDW 11.7, SNT 3.5, IND 3.0, IOD 2.8, ED 3.5, TD 1.8, TED 1.7, HND 8.5, RAD 7.2, FTL 21.8, TIB 15.1

Coloration of holotype in life

(Fig. 9A–C) Dorsal surface reddish brown, with a distinct and dark triangular marking with yellow edges between eyes. Hindlimb with broad black transverse bands. A dark brown vertical band below the eye. Canthus rostralis and supratympanic fold white. Horn-like tubercle at the edge of the upper eyelid yellow. Surface of throat and chest dark brown, with scarlet marbling, posterior region of abdomen white. Ventral surface of limbs brown with white spots and patches. Ventral surface of hand and foot brown, inner and outer metatarsal tubercles and inner metacarpal tubercle pink. Pectoral and femoral glands white. Iris reddish brown.

Figure 9. 

Megophrys wugongensis sp. nov. in life: A–C SYS a002625, male holotype D SYS a002610, female paratype E, F SYS a002611, female paratype.

Coloration of holotype in preservative

Dorsum dark brown, markings on dorsal surface became indistinct, transverse bands on limbs became dark grey and became more distinct. Surface of throat and chest light brown, posterior region of abdomen light yellow, ventral surface of limbs light brown, inner and outer metatarsal tubercles and inner metacarpal tubercle light yellow, all marbling, colored spots and patches absent.

Variation

Measurement data of type series are listed in Table 8.

All paratypes are very similar to holotype in morphology and color pattern. However, dorsal surface yellowish brown in female paratypes SYS a004798, 4801, 4804, markings on dorsal skin indistinct in male paratypes SYS a004777/CIB110011 and SYS a004796, and female paratypes SYS a002610 (Fig. 9D), 4797, 4799, presence of a rectangle marking on central back of trunk in the female paratype SYS a002611 (Fig. 9E–F).

Measurements (in mm; minimum-maximum, mean ± SD) of the type series of Megophrys wugongensis sp. nov.

Species Megophrys wugongensis sp. nov.
Males (n = 4) Females (n = 9)
SVL 31.0–34.1 (32.4 ± 1.3) 38.5–42.8 (40.8 ± 1.3)
HDL 10.2–11.2 (10.7 ± 0.4) 11.8–13.2 (12.6 ± 0.4)
HDW 10.4–11.9 (11.0 ± 0.6) 12.6–13.9 (13.4 ± 0.4)
SNT 3.4–3.9 (3.8 ± 0.2) 4.2–4.8 (4.6 ± 0.2)
IND 3.6–3.7 (3.7 ± 0.1) 3.6–4.2 (4.0 ± 0.2)
IOD 3.1–3.4 (3.2 ± 0.1) 3.6–3.8 (3.7 ± 0.1)
ED 4.1–4.4 (4.3 ± 0.1) 4.1–5.1 (4.4 ± 0.3)
TD 2.0–2.2 (2.1 ± 0.1) 2.1–2.3 (2.2 ± 0.1)
TED 1.7–2.2 (1.9 ± 0.2) 2.1–2.6 (2.4 ± 0.2)
HND 6.5–7.3 (7.0 ± 0.3) 8.2–9.7 (8.7 ± 0.5)
RAD 6.7–7.8 (7.4 ± 0.5) 8.1–9.8 (8.9 ± 0.6)
FTL 17.8–20.9 (19.2 ± 1.3) 21.8–25.0 (23.3 ± 1.1)
TIB 12.4–14.3 (13.3 ± 0.8) 15.0–17.9 (16.0 ± 0.9)
HDL/SVL 0.31–0.34 (0.33 ± 0.01) 0.30–0.33 (0.31 ± 0.01)
HDW/SVL 0.32–0.36 (0.34 ± 0.02) 0.32–0.35 (0.33 ± 0.01)
HDW/HDL 1.01–1.06 (1.03 ± 0.02) 1.03–1.08 (1.06 ± 0.02)
SNT/HDL 0.32–0.37 (0.35 ± 0.02) 0.33–0.40 (0.36 ± 0.02)
SNT/SVL 0.11–0.12 (0.12) 0.11–0.12 (0.11 ± 0.01)
IND/HDW 0.31–0.35 (0.33 ± 0.02) 0.27–0.32 (0.30 ± 0.02)
IOD/HDW 0.27–0.31 (0.29 ± 0.02) 0.27–0.30 (0.28 ± 0.01)
ED/HDL 0.37–0.41 (0.40 ± 0.02) 0.31–0.40 (0.35 ± 0.03)
ED/SVL 0.13–0.14 (0.13 ± 0.01) 0.10–0.13 (0.11 ± 0.01)
TD/ED 0.47–0.52 (0.49 ± 0.02) 0.45–0.53 (0.51 ± 0.03)
TED/TD 0.85–1.10 (0.92 ± 0.12) 0.91–1.14 (1.09 ± 0.07)
HND/SVL 0.20–0.22 (0.21 ± 0.01) 0.20–0.23 (0.21 ± 0.01)
RAD/SVL 0.21–0.24 (0.23 ± 0.02) 0.20–0.25 (0.22 ± 0.02)
TIB/SVL 0.39–0.44 (0.41 ± 0.02) 0.37–0.44 (0.39 ± 0.02)
FTL/SVL 0.56–0.64 (0.59 ± 0.04) 0.54–0.60 (0.57 ± 0.02)

Etymology

The specific epithet “wugongensis” is in reference to the type locality of the new species in the Wugong Mountains. We propose the common English name “Wugongshan Horned Toad” and Chinese name “Wu Gong Shan Jiao Chan (武功山角蟾)”.

Distribution and habits

Currently, Megophrys wugongensis sp. nov. is known from the type locality, Yangshimu Scenic Area, Pingxiang City, Jiangxi Province at approximate 550 m a.s.l., Wugongshan Scenic Area, Ji’an City, Jiangxi Province at approximate 1050–1080 m a.s.l., all located in the Luoxiao Mountains in eastern China. All specimens were collected on leaf litter near a stream in the bamboo forest, males were not heard calling. In consideration of the invisible nuptial pad and nuptial spines in all male specimens and the undeveloped fallopian tubes in all female specimens, the breeding season of M. wugongensis sp. nov. still remains unknown. Tadpoles were not observed. Megophrys wugongensis sp. nov. is sympatric with M. jinggangensis in all localities.

Megophrys (Panophrys) mufumontana J. Wang, Lyu & Y.Y. Wang, sp. nov.

Fig. 10, Table 9

Holotype

SYS a006391, adult male, collected by Zhi-Tong Lyu on 3 August 2017 from Mt. Mufu (28°58'18.45"N, 113°48'58.53"E; 1300 m a.s.l.), Pingjiang County, Yueyang City, Hunan Province, China.

Paratypes (one male & two females)

Adult females, SYS a006390/CIB110012, SYS a006419, and the other adult male, SYS a006392, all collected by Zhi-Ting Lyu on 3 August 2017 from the same locality as the holotype.

Diagnosis

(1) Body size small, SVL 30.1–30.8 mm in two adult males and SVL 36.3 mm in two adult females; (2) head length slightly larger than head width, HDW/HDL ratio 0.98–0.99; (3) tympanum distinct, moderate-sized, TD/ED ratio 0.51–0.58, upper 1/4 part of the tympanum concealed by supratympanic fold; (4) vomerine teeth absent; (5) margin of tongue not notched posteriorly; (6) heels overlapping, tibia-tarsal articulation reach forward to the tympanum in males and to the eye in females; (7) TIB/SVL ratio 0.47–0.53, FTL/SVL ratio 0.68–0.74; (8) fingers without lateral fringes, presence of a subarticular tubercle at the bases of each finger, relative finger lengths II = IV < I < III; (9) toes with rudimentary webbing at their bases and narrow lateral fringes, subarticular tubercles only present at the base of each toe; (10) numerous granules scattered with tubercles present on dorsal surface of body, limbs and surface of flanks, some of which forming a V-shaped, \ /-shaped or X-shaped skin ridge on central back of trunk; (11) presence of a small horn-like tubercle at the edge of the upper eyelid; (12) supratympanic fold distinct; (13) light brown to dark brown dorsally, with a dark triangular marking between eyes; (14) a pair of dark longitudinal and irregular marking with white edges on its upper side on ventrolateral surface of flanks; (15) surface of throat and chest greyish brown with dark brown patches and creamy white spots, surface of abdomen greyish white with creamy white and orange spots; (16) ventral surface of thighs with dense small whitish tubercles.

Comparisons

Comparative data of Megophrys mufumontana sp. nov. with M. dongduanensis sp. nov., M. nankunensis sp. nov., M. jiulianensis sp. nov., Megophrys nanlingensis sp. nov., Megophrys wugongensis sp. nov., M. feii and the 33 recognized members of Megophrys s.l. allocated to the subgenus Panophrys are listed in Table 5.

Megophrys mufumontana sp. nov. differs from M. dongguanensis sp. nov., M. nankunensis sp. nov., M. jiulianensis sp. nov. and M. wugongensis sp. nov. by upper 1/4 part of the tympanum concealed by supratympanic fold (vs. tympanum entirely visible), the heels overlapping when hindlimb is stretched along the side of the body (vs. heels not meeting in M. dongguanensis sp. nov., M. nankunensis sp. nov., M. jiulianensis sp. nov. and M. wugongensis sp. nov.), presence of narrow lateral fringes on toes (vs. absent in M. dongguanensis sp. nov., M. nankunensis sp. nov. and M. jiulianensis sp. nov.), absence of vomerine teeth (vs. present in M. dongguanensis sp. nov., M. nankunensis sp. nov., M. jiulianensis sp. nov. and M. nanlingensis sp. nov.), tongue not notched posteriorly (vs. tongue notched in M. jiulianensis sp. nov. and M. nanlingensis sp. nov.), skin relatively smooth and lacking black horny spines (vs. skin rough with black horny spines in M . jiulianensis sp. nov.).

With the smaller body size, SVL 30.1–30.8 mm in males and 36.3 mm in females, Megophrys mufumontana sp. nov. differs from the 19 members with larger SVL values: M. baolongensis (42.0–45.0 mm in males), M. binchuanensis (32.0–36.0 mm in males, 40.2–42.5 mm in females), M. binlingensis (45.1–51.0 mm in males), M. caudoprocta (81.3 mm in single male), M. daweimontis (34.0–37.0 mm in males, 40.0–46.0 mm in females), M. fansipanensis (41.7–42.5 mm in females), M. hoanglienensis (37.4–47.6 mm in males, 59.6 mm in single female), M. jingdongensis (53.0–56.5 mm in males 63.5 in single female), M. latidactyla (38.9 mm in single male), M. liboensis (34.7–67.7 mm in males, 60.8–70.6 mm in females), M. minor (34.5–41.2 mm in males), M. omeimontis (56.0–59.5 mm in males, 68.0–72.5 mm in females), M. palpebralespinosa (36.2–38.0 mm in males), M. sangzhiensis (54.7 mm in single male), M. shuichengensis (102.0–118.3 mm in males, 99.8–115.6 mm in females), M. spinata (47.2–54.4 mm in males, 54.0–55.0 mm in females), M. tuberogranulatus (33.2–39.6 mm in males, 50.5 mm in single female), M. wushanensis (38.4 mm in single female) and M. wuliangshanensis (41.3 mm in single female).

Megophrys mufumontana sp. nov. differs from 12 species occurring in eastern and southern China (M. acuta, M. brachykolos, M. boettgeri, M. cheni, M. huangshanensis, M. insularis, M. jinggangensis, M. kuatunensis, M. lini, M. lishuiensis, M. obesa and M. ombrophila) by the following combination of characters: upper 1/4 part of the tympanum concealed by supratympanic fold (vs. tympanum entirely visible in the 12 species above), absence of vomerine teeth (vs. present in M. insularis and M. jinggangensis), tongue not notched posteriorly (vs. tongue notched in M. boettgeri, M. cheni, M. huangshanensis, M. insularis and M. kuatunensis), presence of narrow lateral fringes on toes (vs. absent in M. brachykolos, M. huangshanensis, M. insularis, M. lishuiensis, M. obesa and M. ombrophila; wide in M. boettgeri and M. cheni), toes with rudimentary webbing (vs. toes without webbing in M. huangshanensis, M. kuatunensis, M. lishuiensis and M. ombrophila), the heels overlapping when hindlimb is stretched along the side of the body (vs. heels not meeting in M. acuta, M. brachykolos, M. insularis, M. obesa and M. ombrophila).

Megophrys mufumontana sp. nov. differs from the remaining M. leishanensis and M. rubrimera allocated to the subgenus Panophrys by the absence of vomerine teeth (vs. present in M. rubrimera), tongue not notched posteriorly (vs. tongue notched in M. rubrimera), upper 1/4 part of the tympanum concealed by supratympanic fold (vs. tympanum entirely visible in M. leishanensis and M. rubrimera), toes with narrow lateral fringes (vs. absent in M. leishanensis; indistinct or absent in M. rubrimera).

Megophrys mufumontana sp. nov. further differs from M. feii, for which molecular data are lacking and cannot be allocated to any subgenus base on morphology only (Yang et al. 2018) by the larger body size, SVL 30.1–30.8 mm in males and 36.3 mm in females (VS. 24.3–25.1 mm in males, 28.2–28.9 mm in females), tongue not notched posteriorly (vs. tongue notched), toes with narrow lateral fringes (vs. moderate or wide).

Description of holotype

Adult male. Habitus small, SVL 30.8 mm; head length slightly larger than head width, HDW/HWL 0.98; snout rounded in dorsal view, sloping backward to mouth in profile, protruding well beyond margin of lower jaw; top of head flat; eye large, ED/HDL 0.30; nostril oblique ovoid; pupil vertical; canthus rostralis well developed; loreal region vertical; internasal distance slightly larger than interorbital distance; tympanum distinct, moderate-sized, TD/ED 0.56; large ovoid choanae at the base of the maxilla; weak vomerine ridge present, vomerine teeth absent; margin of tongue not notched posteriorly.

RAD/SVL 0.25, HND/SVL 0.30; absence of lateral fringes and webbing on fingers, relative finger lengths II = IV < I < III; tip of finger rounded, slightly swollen; presence of a distinct subarticular tubercle on the base of each finger; outer metacarpal tubercles indistinct, inner metacarpal tubercles distinct and observably enlarged. Hindlimbs long, tibio-tarsal articulation reaching forward to the tympanum when hindlimb is stretched along the side of the body; heels overlapping when the flexed hindlimbs are held at right angles to the body axis; TIB/SVL 0.53 and FTL/SVL 0.74; relative toe lengths I < II < V < III < IV; tips of toes round and slightly dilated; presence of rudimentary webbing and narrow lateral fringes on toes but absence of tarsal folds; presence of a subarticular tubercle only at the bases of each toes; presence of a long ovoid inner metatarsal tubercle and absence of outer metatarsal tubercle.

Dorsal skin texture rough with dense granules and scattered with small tubercles, some of which forming a \ /-shaped skin ridge on central back of trunk; presence of a small horn-like tubercle at the edge of upper eye lid; distinct supratympanic fold curving posteroventrally from posterior corner of eye to a level above insertion of arm; upper 1/4 part of the tympanum covered by supratympanic fold; ventral skin texture smooth with granules; pectoral gland large, closer to axilla; single large femoral gland on rear of thigh.

Measurements of holotype (in mm)

SVL 30.1, HDL 11.6, HDW 11.4, SNT 3.5, IND 3.0, IOD 2.8, ED 3.5, TD 18, TED 1.7, HND 8.5, RAD 7.2, FTL 21.8, TIB 15.1.

Coloration of holotype in life

(Fig. 10A–D) Dorsal surface brown, with a distinct and incomplete dark triangular marking between eyes. Hindlimb with black transverse bands. A dark brown vertical band below the eye. Horn-like tubercle at the edge of the upper eyelid red. Surface of throat and chest greyish brown with dark brown patches. Surface of abdomen greyish white with creamy white and orange spots. Ventral surface of limbs pink with white spots and light-yellow patches. Ventral surface of hand and foot brown, inner and outer metatarsal tubercles and inner metacarpal tubercle pink. Pectoral and femoral glands white. Iris white.

Figure 10. 

Megophrys mufumontana sp. nov. in life: A–D SYS a006391, male paratype E–F SYS a006392, female paratype.

Coloration of holotype in preservative. Coloration of dorsum dark brown, markings on dorsal surface and transverse bands on limbs became indistinct. Ventral surface of throat, chest and abdomen dark grey. All patches on ventral surface indistinct, all colored spots absent. Ventral surface of limbs light yellow.

Variation

Measurement data of type series are listed in Table 9.

All paratypes are very similar to holotype SYS a006391 in morphology and color pattern. However, tibia-tarsal articulation reaching forward to the eye when hindlimb is stretched along the side of the body in all females, and granules and tubercles forming a \ /-shaped skin ridge on central back of trunk in the holotype (vs. X-shaped in SYS a006390, 6419; V-shaped in SYS a006392 (Fig. 10E–F)).

Measurements (in mm; minimum-maximum, mean ± SD) of the type series of Megophrys mufumontana sp. nov.

Species Megophrys mufumontana sp. nov.
Males (n = 2) Females (n = 2)
SVL 30.1–30.8 36.3
HDL 11.6–11.9 11.8–12.4
HDW 11.4–11.7 11.7–12.3
SNT 3.5–3.7 3.7–4.2
IND 3.0–3.1 3.5–3.6
IOD 2.8–2.9 3.2–3.3
ED 3.5–3.6 3.7–3.8
TD 1.7–1.8 2.1–2.2
TED 1.7–1.8 1.8–1.9
HND 8.5–9.2 9.4–9.9
RAD 7.2–7.7 8.0–8.2
FTL 21.8–22.9 24.8–25.1
TIB 15.1–16.3 16.9–17.5
HDL/SVL 0.39 0.33–0.34
HDW/SVL 0.38 0.32–0.34
HDW/HDL 0.98 0.99
SNT/HDL 0.30–0.31 0.31–0.34
SNT/SVL 0.12 0.10–0.12
IND/HDW 0.26 0.29–0.30
IOD/HDW 0.25 0.27
ED/HDL 0.30 0.31
ED/SVL 0.12 0.10
TD/ED 0.51–0.56 0.57–0.58
TED/TD 0.90–0.94 0.86
HND/SVL 0.28–0.30 0.26–0.27
RAD/SVL 0.24–0.25 0.22–0.23
TIB/SVL 0.50–0.53 0.47–0.48
FTL/SVL 0.72–0.74 0.68–0.69

Etymology

The specific epithet “mufumontana” is in reference to the type locality of the new species, Mt. Mufu. We propose the common English name “Mufushan Horned Toad” and Chinese name “Mu Fu Shan Jiao Chan (幕阜山角蟾)”.

Distribution and habits

Currently, Megophrys mufumontana sp. nov. is known only from Mt. Mufu, Pingjiang County, Yueyang City, Hunan Province, China at approximate 1300 m a.s.l.. All specimens were collected on leaf litter near a stream (about 5 m wide) surrounded by moist subtropical evergreen broadleaved forests, males were not heard calling. Tadpoles were not observed. Because none of the males have nuptial pads developed and none of the females have fallopian tubes and eggs developed, the breeding season of M. mufumontana sp. nov. remains unknown.

Discussion

Megophrys dongguanensis sp. nov. is easily confused with M. brachykolos because of the relatively short shanks. In addition, the type locality of the new species is at a straight-line distance of approximately 72 km from the type locality (Hongkong Island), and at a straight-line distance of approximately 32 km from the closest locality (Sanzhoutian of Shenzhen City) of M. brachykolos. Currently, eight Megophrys species in the subgenus Panophrys were found to have comparatively short shanks with heels not meeting when thighs are adpressed at right angles with respect to the body axis: M. dongguanensis sp. nov., M. nankunensis sp. nov., M. wugongensis sp. nov., M. acuta, M. brachykolos, M. insularis, M. megacephala and M. obesa.

In our previous study (Liu et al. 2018), 41 cryptic species within the subgenus Panophrys were revealed, and one of them was recently described as Megophrys leishanensis by Li et al. (2018). Moreover, except for M. mufumontana sp. nov. (not mentioned in Liu et al. (2018)), five of them are described in this study. Currently, the total number of recognized species of the subgenus Panophrys rises to 39, which makes it the most species-rich subgenus of Megophrys (≈46.4%). It’s worth noting that there remain still 33 undescribed species according to Liu et al. (2018), and 27 of them are found in southeastern China, which further reveals the unusually high level of species diversity in this region.

As the diversity of the subgenus Panophrys was confirmed to be extremely underestimated (Chen et al. 2017; Mahony et al. 2017; Liu et al. 2018), a number of new Panophrys species have been described since 2017 (i.e. Megophrys lishuiensis, M. insularis, M. rubrimera, M. liboensis, and six new species in this study). However, all of these species have narrow distributions. For example, M. insularis is currently known only from an offshore island in Guangdong (Wang et al. 2017b), and M. liboensis is currently known only from a cave in Libo, Guizhou (Zhang et al. 2017). For the six new species in this study, M. dongguanensis sp. nov., M. nankunensis sp. nov., M. wugongensis sp. nov. and M. mufumontana sp. nov. are currently only found in their type localities. This situation of “micro-endemism” (Liu et al. 2018) has brought great challenges for the protection of these unique toads.

Among the six new species described in this paper, M. jiulianensis sp. nov. is sympatric with M. nankunensis sp. nov. in Mt. Nankun while also being sympatric with M. hongshanensis sp. nov. in Mt. Jiulian. Further, M. mufumontana sp. nov. is sympatric with a known congener M. jinggangensis in Mt. Mufu and M. wugongensis sp. nov. is sympatric with M. jinggangensis in Mt. Wugong. By combining the localities of these species in our phylogenetic trees (Fig. 2), our results also support the conclusion of “sympatric but distant phylogenetically” (Liu et al. 2018), that is, sympatric distribution is very common in horned toads within the subgenus Panophrys while they are distantly related in the phylogeny (Fei et al. 2012; Li et al. 2014; Wang et al. 2014; Liu et al. 2018). These geographical patterns of “sympatric but distant phylogenetically” and “micro-endemism” indicate that the Asian horned toads would be good candidates for studies on speciation and biogeography.

Acknowledgments

We thank Yao Li from the Biology Museum of Sun Yet-sen University, for her help in the lab work, and thank Runlin Li, Hailong He, Chaoyu Lin, Siyu Zhang and Guoling Chen from the Biology Museum of Sun Yet-sen University, for their help in the field work.

The work was supported by Project of Comprehensive Scientific Survey of Luoxiao Mountains Range (No. 2013FY111500) of Ministry of Science and Technology of China to Ying-Yong Wang.

References

  • Boulenger GA (1899) Descriptions of three new reptiles and a new batrachian from Mount Kina Balu, North Borneo. Annals and Magazine of Natural History Series 7(4): 453. https://doi.org/10.1080/00222939908678228
  • Chen JM, Zhou WW, Poyarkov NA, Stuart BL, Brown RM, Lathrop A, Wang YY, Yuan ZY, Jiang K, Hou M, Chen HM, Suwannapoom C, Nguyen SN, Duong TV, Papenfuss TJ, Murphy RW, Zhang YP, Che J (2017) A novel multilocus phylogenetic estimation reveals unrecognized diversity in Asian horned toads, genus Megophrys sensu lato (Anura: Megophryidae). Molecular Phylogenetics and Evolution 106: 28–43. https://doi.org/10.1016/j.ympev.2016.09.004
  • Deuti K, Grosjean S, Nicolas V, Vasudevan K, Ohler A (2017) Nomenclatural puzzle in early Xenophrys nomina (Anura, Megophryidae) solved with description of two new species from India (Darjeeling hills and Sikkim). Alytes (34): 20–48.
  • Dubois A (1987) Miscellanea taxinomica batrachologica (I). Alytes 1987 [1986]: 7–95.
  • Dubois A, Ohler A (1998) A new species of Leptobrachium (Vibrissaphora) from northern Vietnam, with a review of the taxonomy of the genus Leptobrachium (Pelobatidae, Megophyinae). Dumerilia 4(14): 1–32.
  • Fei L, Hu SQ, Ye CY, Huang YZ (2009) Fauna Sinica. Amphibia Vol. 2 Anura. Science Press, Beijing, 957 pp. [In Chinese]
  • Fei L, Ye CY (2016) Amphibians of China. Vol. 1. Science Press, Beijing, 1040 pp.
  • Fei L, Ye CY, Jiang JP (2012) Colored atlas of Chinese amphibians and their distributions. Sichuan Publishing House of Science & Technology, Chengdu, 619 pp. [In Chinese]
  • Frost DR, Grant T, Faivovich J, Bain RH, Haas A, Haddad CFB, De Sa, Rafael O, Channing A, Wilkinson M, Donnellan SC, Raxworthy CJ, Campbell JA, Blotto BL, Moler P, Drewes RC, Nussbaum RA, Lynch JD, Green DM, Wheeler WC (2006) The amphibian tree of life. Bulletin of the American Museum of natural History 297: 1–370. https://doi.org/10.1206/0003-0090(2006)297[0001:TATOL]2.0.CO;2
  • Huang YZ, Fei L (1981) Two new species of amphibians from Xizang. Acta Zootaxonomica Sinica/Dong Wu Fen Lei Xue Bao 6: 211–215.
  • Inger RF (1989) Four new species of frogs from Borneo. Malayan Nature Journal, Kuala Lumpur 42: 229–243.
  • Jiang JP, Ye CY, Fei L (2008) A new horn toad Megophrys sangzhiensis from Hunan, China (Amphibia, Anura). Zoological Research 29(2): 219–222. [in Chinese with English abstract] https://doi.org/10.3724/SP.J.1141.2008.00219
  • Jiang JP, Yuan FR, Xie F, Zheng ZH (2003) Phylogenetic relationships of some species and genera in megophryids inferred from partial sequences of mitochondrial 12S and 16S rRNA genes. Zoological Research 24: 241–248.
  • Li C, Wang YZ (2008) Taxonomic review of Megophrys and Xenophrys, and a proposal for Chinese species (Megophryidae, Anura). Acta Zootaxonomica Sinica 33(1): 104−106.
  • Li YL, Jin MJ, Zhao J, Liu ZY, Wang YY, Pang H (2014) Description of two new species of the genus Xenophrys (Amphibia: Anura: Megophryidae) from Heishiding Natural Resreve, Fengkai, Guangdong, China, based on molecular and morphological data. Zootaxa 3795: 449–471. https://doi.org/10.11646/zootaxa.3795.4.5
  • Li SZ, Xu N, Liu J, Jiang JP, Wei G, Wang B (2018) A new species of the asian toad genus Megophrys sensu lato (Amphibia: Anura: Megophryidae) from Guizhou Province, China. Asian Herpetological Research 9(4): 224–239.
  • Liu ZY, Chen GL, Zhu TQ, Zeng ZC, Lyu ZT, Wang J, Messenger K, Greenberg AJ, Guo ZX, Yang ZH, Shi SH, Wang YY (2018) Prevalence of cryptic species in morphologically uniform taxa – fast speciation and evolutionary radiation in Asian toads. Molecular Phylogenetics and Evolution. https://doi.org/10.1016/j.ympev.2018.06.020
  • Mahony S, Foley NM, Biju SD, Teeling EC (2017) Evolutionary history of the Asian Horned Frogs (Megophryinae): integrative approaches to timetree dating in the absence of a fossil record. Molecular Biology and Evolution 34(3): 744–771. https://doi.org/10.1093/molbev/msw267
  • Mahony S, Kamei RG, Teelin Ec, Biju SD (2018) Cryptic diversity within the Megophrys major species group (Amphibia: Megophryidae) of the Asian Horned Frogs: Phylogenetic perspectives and a taxonomic revision of South Asian taxa, with descriptions of four new species. Zootaxa 4523: 1–96. https://doi.org/10.11646/zootaxa.4523.1.1
  • Messenger KR, Dahn HA, Liang Y, Xie P, Wang Y, Lu C (2019) A new species of the genus Megophrys Gunther, 1864 (Amphibia: Anura: Megophryidae) from Mount Wuyi, China. Zootaxa 4554: 561–583. https://doi.org/10.11646/zootaxa.4554.2.9
  • Malkmus R, Manthey U, Vogel G, Hoffmann P, Kosuch J (2002) Amphibians & Reptiles of Mount Kinabalu (North Borneo). Ruggell: A.R.G. Gantner Verlag, 424pp.
  • Mo XY, Shen YH, Li HH, Wu XS (2010) A new species of Megophrys (Amphibia: Anura: Megophryidae) from the northwestern Hunan Province, China. Current Zoology 56(4): 432−436.
  • Orlov NL, Poyarkov NA, Nguyen TT (2015) Taxonomic notes on Xenophrys frogs (Megophryidae: Anura) of Vietnam, with description of a new species. Russian Journal of Herpetology 22: 206–218.
  • Rao DQ, Yang DT (1997) The karyotypes of Megophryinae (Pelobatinae) with a discussion on their classification and phylogenetic relationships. Asiatic Herpetological Research 7: 93–102. https://doi.org/10.5962/bhl.part.18858
  • Ronquist F, Teslenko M, Van Der Mark P, Ayres DL, Darling A, Höhna S, Larget B, Liu L, Suchard MA, Huelsenbeck JP (2012) MrBayes 3.2: efficient Bayesian phylogenetic inference and model choice across a large model space. Systematic Biology 61: 539–542. https://doi.org/10.1093/sysbio/sys029
  • Stuart BL, Sok K, Neang T (2006) A collection of amphibians and reptiles from hilly eastern Cambodia. Raffles Bulletin of Zoology, Singapore 54: 129–155.
  • Simon C, Frati F, Beckenbach A, Crespi B, Liu H, Flook P (1994) Evolution, weighting, and phylogenetic utility of mitochondrial gene sequences and a compilation of conserved polymerasechain reaction primers. Annals of the Entomological Society of America 87: 651–701. https://doi.org/10.1093/aesa/87.6.651
  • Tapley B, Cutajar TP, Mahony S, Chung NT, Dau VQ, Nguyen TT, Luong HV, Rowley JJL (2017) The Vietnamese population of Megophrys kuatunensis (Amphibia: Megophryidae) represents a new species of Asian horned frog from Vietnam and southern China. Zootaxa 4344: 465–492. https://doi.org/10.11646/zootaxa.4344.3.3
  • Tapley B, Cutajar TP, Mahony S, Nguyen CT, Dau VQ, Luong AM, Le DT, Nguyen TT, Nguyen TQ, Portway C, Luong HV, Rowley JJL (2018) Two new and potentially highly threatened Megophrys Horned frogs (Amphibia: Megophryidae) from Indochina’s highest mountains. Zootaxa 4508: 301–333. https://doi.org/10.11646/zootaxa.4508.3.1
  • Tamura K, Stecher G, Peterson D, Filipski A, Kumar S (2013) MEGA6: molecular evolutionary genetics analysis, version 6.0. Molecular Biology and Evolution 30: 2725–2729. https://doi.org/10.1093/molbev/mst197
  • Thompson JD, Gibson TJ, Plewniak F, Jeanmougin F, Higgins DG (1997) The CLUSTAL_X windows interface: flexible strategies for multiple sequence alignment aided by quality analysis tools. Nucleic Acids Research 25: 4876–4882. https://doi.org/10.1093/nar/25.24.4876
  • Tian YZ, Gu XM, Sun AQ (2000) A new species of Xenophrys in China (Amphibia: Pelobatidae). Acta Zootaxonomica Sinica 25: 462−466.
  • Wang YF, Liu BQ, Jiang K, Jin W, Xu JN, Wu CH (2017a) A new species of the horn toad of the genus Xenophrys from Zhejiang, China (Amphibia: Megophryidae). Chinese Journal of Zoology 52(1): 19–29. [in Chinese with English abstract]
  • Wang J, Liu ZY, Lyu ZT, Zeng ZC, Wang YY (2017b) A new species of the genus Xenophrys (Amphibia: Anura: Megophryidae) from an offshore island in Guangdong Province, southeastern China. Zootaza 4324: 541–556. https://doi.org/10.11646/zootaxa.4324.3.8
  • Wang YY, Zhao J, Yang JH, Zhou ZX, Chen GL, Liu Y (2014) Morphology, molecular genetics, and bioacoustics support two new sympatric Xenophrys (Amphibia: Anura: Megophryidae) species in Southeast China. PLoS ONE 9: e93075. https://doi.org/10.1371/journal.pone.0093075
  • Wang YY, Zhang TD, Zhao J, Sung YH, Yang JH, Pang H, Zhang Z (2012) Description of a new species of the genus Xenophrys Günther, 1864 (Amphibia: Anura: Megophryidae) from Mount Jinggang, China, based on molecular and morphological data. Zootaxa 3546: 53–67. https://doi.org/10.11646/zootaxa.3546.1.4
  • Ye CY, Fei L (1995) Taxonomic studies on the small type Megophrys in China including descriptions of the new species (subspecies) (Pelobatidae: genus Megophrys). Acta Herpetologica Sinica/Liangqi Paxing Dongwu Yanjiu 4–5: 72–81.
  • Ye CY, Fei L, Xie F (2007) A new species of MegophryidaeMegophrys baolongensis from China (Amphibia, Anura). Herpetologica Sinica 11: 38–41.
  • Zhang YG, Li N, Xiao JL, Pan T, Wang H, Zhang B, Zhou J (2017) A new species of the genus Xenophrys (Amphibia: Anura: Megophryidae) from Libo County, Guizhou, China. Asian Herpetological Research 8: 75–85.
  • Zheng YC, Zeng XM, Yuan YZ, Liu ZJ (2004) Phylogenetic positions of Ophryophryne and four Leptobrachium group genera in Megophryidae (Anura). Sichuan Journal of Zoology 23: 290–295.

Appendix 1

Specimens of comparative species examined

Megophrys boettgeri (n = 13): China: Jiangxi Provence: Guixi City: Yangjifeng Nature Reserve (the middle area of Wuyi Mountains, 600–883 m a.s.l.): SYS a000312, 000315, 000328–000330, 000376, 000378; Guangfeng County: Tongboshan Nature Reserve (the eastern area of Wuyi Mountains, 450–821 m a.s.l.): SYS a001671–001673, 001683, 001700.

Megophrys brachykolos (n = 21): China: Hong Kong: SYS a001502–001503; Guangdong Province: Shenzhen City: Yangtaishan Forest Park (60–150 m a.s.l.): SYS a2051–002056, 002069–002074, 002413; Qiniangshan Geological Park (30–50 m a.s.l.): SYS a002405–002410.

Megophrys caudoprocta (n = 3): China: Hunan Province: Zhangjiajie City: Sangzhi County: Mt. Badagong (1100–1200 m a.s.l.): SYS a004281, 004308–4309.

Megophrys cheni (n = 19): China: Jiangxi Province: Jinggangshan City: Mt. Jinggang (1200–1260 m a.s.l.): SYS a001427–001429, SYS a001871–001873; Hunan Province: Yanling County: Taoyuandong Nature Reserve: Lishuzhou Village (1480–1530 m a.s.l.): SYS a002123–002127, Dayuan Farm (1480 m a.s.l.): SYS a002140–002145.

Megophrys huangshanensis (n = 10): China: Jiangxi Province: Wuyuan County: Mount Dazhang (600–900 m a.s.l.): SYS a001314–001323.

Megophrys insularis (n = 6): China: Guangdong Province: Shantou City: Nan’ao Island (50–500 m a.s.l.): SYS a002167–002171, SYS a003666/CIB 106881.

Megophrys jingdongensis (n = 2): Yunnan Province: Jingdong County: Mt. Wuliang (1800 m a.s.l.): SYS a003928–3929.

Megophrys jinggangensis (n = 10): China: Jiangxi Province: Jinggangshan City: Mt. Jinggang (700–900m a.s.l.): SYS a001413–001416, 001430; Hunan Province: Yanling County: Taoyuandong Nature Reserve (800–1000 m a.s.l.): SYS a001859–001863.

Megophrys kuatunensis (n = 3): China: Fujian Province: Wuyishan City (=Ch’ungan Hsien): Guadun Village (= Kuatun Village, 1060–1220 m a.s.l.): SYS a001579 and 001590; Jiangxi Province: Guixi City: Yangjifeng Nature Reserve (the middle area of Wuyi Mountains, 950 m a.s.l.): SYS a000241.

Megophrys lini (n = 27): China: Jiangxi Province: Jinggangshan City: Mt. Jinggang (1100–1610 m a.s.l.): SYS a001417–001424, SYS a002375–002386; Suichuan County: Nanfengmian Nature Reserve (1150–1250 m a.s.l.): SYS a002369–002374; Hunan Province: Yanling County: Taoyuandong Nature Reserve: Niushiping Village (1360 m a.s.l.): SYS a002128.

Megophrys minor (n = 4): China: Sichuan Province: Dujiangyan City: Mt. Qingcheng: SYS a003209, 003211–3213.

Megophrys omeimontis (n = 6): China: Sichuan Province: Mt. Emei: SYS a001798–001801, 001940–001941.

Megophrys sangzhiensis (n = 6): China: Hunan Province: Zhangjiajie City: Sangzhi County: Mt. Badagong (1100–1200 m a.s.l.): SYS a004306–004307, 004313–004316.

Megophrys spinata (n = 2): China: Guizhou Province: Leishan County: Mt. Leigong: SYS a002226–002227.

Megophrys tuberogranulatus (n = 1): China: Hunan Province: Zhangjiajie City: Sangzhi County: Mt. Badagong (1100–1200 m a.s.l.): SYS a004310.Megophrys wuliangshanensis (n = 2): Yunnan Province: Jingdong County: Mt. Wuliang (1800 m a.s.l.): SYS a003924–3925.