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Research Article
A new species of Nidirana (Anura, Ranidae) from northern Guangxi, China
expand article infoWei-Cai Chen‡, Jian-Ping Ye§, Wan-Xiao Peng‡, Peng Li‡, Tong-Ping Su‡, Gui-Dong Yu|, Zhi-Ying Cheng|
‡ Nanning Normal University, Nanning, China
§ Maoershan National Nature Reserve, Guilin, China
| Guangxi Forest Inventory and Planning Institute, Nanning, China

Corresponding author: Wei-Cai Chen ( chenweicai2003@126.com )

Academic editor: Annemarie Ohler
© 2022 Wei-Cai Chen, Jian-Ping Ye, Wan-Xiao Peng, Peng Li, Tong-Ping Su, Gui-Dong Yu, Zhi-Ying Cheng.
This is an open access article distributed under the terms of the Creative Commons Attribution License (CC BY 4.0), which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited.
Citation: Chen W-C, Ye J-P, Peng W-X, Li P, Su T-P, Yu G-D, Cheng Z-Y (2022) A new species of Nidirana (Anura, Ranidae) from northern Guangxi, China. ZooKeys 1135: 119-137. https://doi.org/10.3897/zookeys.1135.94371
ZooBank: urn:lsid:zoobank.org:pub:3C8C2AAD-B83C-4FB6-9A72-34ADE05DA610
Open Access

Abstract

A new species of music frog, Nidirana guibeiensis sp. nov., is described from northern Guangxi, China. Based on two mtDNA fragments analyzed, phylogenetic trees reveal that N. guibeiensis sp. nov. is most closely related to N. leishanensis. However, the new species can be identified by conspicuous diagnostic morphological characteristics as well as bioacoustics. In contrast to the known Nidirana species, the advertisement calls of the new species can be divided into three types, calls with one, two, and three notes. In addition, the new species has nest construction behavior, which is inconsistent with N. leishanensis. Nidirana guibeiensis sp. nov. occurs in paddy fields or still pools at 300–1300 m a.s.l.

Keywords

Bioacoustics, morphology, nest construction, phylogeny

Introduction

The genus Nidirana Dubois, 1992 is widespread in eastern and southeastern Asia (Frost 2021; AmphibiaChina 2022). Recently, the known diversity of Nidirana has increased dramatically, due to combined morphological, molecular, and bioacoustical analyses (Li et al. 2019; Lyu et al. 2020a, 2020b, 2021). To date, there are 17 recognized Nidirana species. Most have been reported in the past five years (AmphibiaChina 2022). Five Nidirana species have been confirmed to occur in Guangxi: N. guangxiensis Mo, Lyu, Huang, Liao & Wang, 2021, N. leishanensis Li, Wei, Xu, Cui, Fei, Jiang, Liu & Wang, 2019, N. shiwandashanensis Chen, Peng, Li & Liu, 2022, N. xiangica Lyu & Wang, 2020a, and N. yaoica Lyu, Mo, Wan, Li, Pang & Wang, 2019. Nidirana adenopleura (Boulenger, 1909) has always been considered to occur throughout Guangxi (Fei et al. 2009; Mo et al. 2014). However, recent research has indicated the misidentifications of specimens allocated to N. adenopleura, and no evidence supports the occurrence of N. adenopleura in Guangxi (Lyu et al. 2021; Chen et al. 2022).

In 2022, we conducted surveys in northern Guangxi and collected 15 Nidirana specimens. These specimens differ from the known Nidirana species in morphology, phylogeny, and bioacoustics. Herein, we describe these specimens as a new species of Nidirana.

Materials and methods

Sampling and morphological examination

Fourteen adults and one subadult were collected at the Maoershan National Nature Reserve (n = 3), Zhongfeng Town, Ziyuan County (n = 11), and Lingtan Town, Xing’an County (n = 1) in northern Guangxi, China (Fig. 1). After euthanasia with isoflurane, all specimens were fixed in 10% formalin, then transferred to 75% ethanol, and finally deposited at Nanning Normal University (NNU). Before being fixed, muscle samples were taken and stored in 100% ethanol for molecular analysis. The definition of morphological characteristics and measurements followed Chen et al. (2022). The following measurements were taken with digital calipers to the nearest 0.1 mm:

Figure 1. 

Localities of the new species and its sister taxa. The source of the map came from WorldClim (http://www.worldclim.com/version2).

SVL snout-vent length (from the tip of snout to posterior margin of vent);

HDL head length (from the tip of snout to the articulation of jaw);

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

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

IND internasal distance (distance between nares);

IOD interorbital distance (minimum distance between upper eyelids);

ED eye diameter (from the anterior corner of eye to posterior corner of eye);

TD tympanum diameter (horizontal diameter of tympanum);

HND hand length (from the proximal border of the outer palmar tubercle to the tip of Finger III);

FTL foot length (from the distal end of the shank to the tip of Toe IV);

TIB tibial length (from the outer surface of the flexed knee to the heel).

Sex was identified by examining the nuptial pad and suprabrachial gland. The webbing formula followed Savage (1975).

Geographically and phylogenetically, the new species, N. leishanensis, and N. xiangica are close to each other. Fourteen adults were measured for comparison, whereas the morphological data of N. leishanensis and N. xiangica came from the references (Li et al. 2019; Lyu et al. 2020a). A principal component analysis (PCA) and the Mann-Whitney U tests were performed on SPSS, based on the adult male specimens. To reduce the impact of allometry, the correct value from the ratio of each character to SVL was calculated, then log-transformed for analyzing. The significance level was set at 0.05. Morphological comparison data came from the collected specimens (Appendix 1) and the references in Table 1.

Table 1.
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Diagnostic characters separating Nidirana guibeiensis sp. nov. from all congeners. Labial tooth row formula from Dubois (1995).

ID Species SVL of males (mm) SVL of females (mm) Fingers tips Lateroventral groove on fingers Relative length of fingers Toes tips Lateroventral groove on toes Tibio-tarsal articulation Subgular vocal sacs Nuptial pad Spinules on dorsal skin Nest construction Tadpole labial tooth row formula Calling References
1 N. guibeiensis sp. nov. 50.2–63.6 54.6 Dilated Present except finger I II < IV < I < III Dilated Present Eye-snout Present One on finger I Absent Present ? 1–3 notes This study
2 N. adenopleura 43.1–57.6 47.6–60.7 Dilated Present except finger I II < I < IV < III Dilated Present Snout tip or eye-snout Present One on finger I Entire or posterior Absent I:1+1/1+1:II or I:0+0/1+1:I 2–5 regular notes Lyu et al. (2017, 2020b)
3 N. chapaensis 35.5–42.5 41.0–51.8 Dilated Present except finger I II < I = IV < III Dilated Present Nostril Present Two on finger I Absent or few above vent Present I:1+2/1+1:II 3 notes Chuaynkern et al. (2010)
4 N. daunchina 40.6–51.0 44.0–53.0 Dilated Absent or rarely present II < I < IV < III Dilated Present Nostril Present One on finger I Absent Present I:1+1/1+1:II or I:1+1/2+2:I 2–5 notes containing a specific first note Liu (1950); Lyu et al. (2017)
5 N. guangdongensis 50.0–58.4 55.3–59.3 Dilated Present except finger I II < I < IV < III Dilated Present Nostril Present One on finger I Entire Absent ? 2–4 regular notes Lyu et al. (2020a)
6 N. guangxiensis 40.2–47.6 49.9–51.0 Dilated Present on fingers III and IV II < I < IV < III Dilated Present Nostril Present One on finger I Absent Present I: 1+1/1+1:II 6–11 rapidly repeated regular notes Lyu et al. (2021)
7 N. hainanensis 32.8–44.4 ? Dilated Present II < I < IV < III Dilated Present Nostril Present Absent Absent Present ? 2–4 fast-repeated double-notes Fei et al. (2007, 2009)
8 N. leishanensis 49.5–56.4 43.7–55.3 Dilated Present II < IV < I < III Dilated Present Eye-snout Present Two on fingers I and II Absent Absent I:1+2/ 1+1:II 1 single note Li et al. (2019)
9 N. lini 44.1–63.1 57.7–68.6 Dilated Present except finger I II < I < IV < III Dilated Present Beyond snout Present One on finger I Posterior Absent I:1+1/1+1:II 5–7 notes containing a specific first note Chou (1999); Lyu et al. (2017)
10 N. mangveni 53.6–59.7 59.7–65.1 Dilated Present on fingers III and IV I < II < IV < III Dilated Present Anterior corner of eye Present One on finger I Entire or posterior Absent ? 2–7 regular notes Lyu et al. (2020a)
11 N. nankunensis 33.3–37.1 37.8–39.5 Dilated Present except finger I II < I < IV < III Dilated Present Nostril Present One on finger I Absent or few above vent Present I:1+1/1+1:II 13–15 notes containing a specific first note Lyu et al. (2017)
12 N. occidentalis 44.5–53.0 55.6–61.3 Not dilated Absent II < I < IV < III Not dilated Absent Eye Present One on finger I Posterior Absent ? 3–5 regular notes Lyu et al. (2020b)
13 N. okinavana 35.5–42.8 44.6–48.8 Dilated Present except finger I II < I < IV < III Dilated Present Eye center-near nostril Absent Poorly one on finger I Absent Present I:1+1/1+1:II 10–25 fast-repeated notes Chuaynkern et al. (2010); Lyu et al. (2017)
14 N. pleuraden 46.2–52.3 46.9–61.7 Not dilated Absent II < I < IV < III Not dilated Absent Nostril Present One on finger I Posterior Absent I:1+1/1+1: II 1–4 regular notes Lyu et al. (2017, 2020b)
15 N. shiwandashanensis 46.2–50.8 48.3 Dilated Present II < IV < I < III Dilated Present Eye Present One on finger I Absent ? I: 1+1/1+1:II 6–8 double-notes Chen et al. (2022)
16 N. xiangica 56.3–62.3 53.5–62.6 Dilated Present II < I < IV < III Dilated Present Eye-snout Present One on finger I Entire Absent ? 2–3 notes containing a specific first note Lyu et al. (2020a)
17 N. yaoica 42.1–45.6 ? Dilated Present II < I < IV < III Dilated Present Nostril Present One on finger I Absent ? (Probably present) ? 1–3 fast-repeated regular notes Lyu et al. (2019)
18 N. yeae 41.2–43.5 44.7 Dilated Absent II < IV < I < III Dilated Present Eye Present One on finger I Absent ? (Probably absent) I: 1+1/1+1:II 2–6 notes containing a specific first note Wei et al. (2020)

Phylogenetic analyses

Bayesian inference (BI) and maximum likelihood (ML) methods were used to analyze phylogenetical relationships based on partial 16S ribosomal RNA gene (16S, ~1050 bp) and partial cytochrome oxidase subunit I gene (COI, ~640 bp) sequences analyses. The two mtDNA fragments were amplified and sequenced following Lyu et al. (2019). PCR reaction conditions included 94 °C for 5 min; 35 cycles of denaturing at 94 °C for 35 s, annealing at 55 °C (for 16S)/52 °C (for COI) for 45 s, and extending at 72 °C for 60 s. Sequences were sequenced using an ABI3730 automated DNA sequencer in Sangon Biotech (Shanghai) Co., Ltd (Guangzhou, China). In addition, homologous sequences of Nidirana species downloaded from GenBank were included in our phylogenetic analysis (Table 2). These sequences contain all holotypes or paratypes of Nidirana species known in China. The BI analysis was implemented using MRBAYES v. 3.1.2 (Ronquist and Huelsenbeck 2003). The best-fit model (GTR+I+G) was chosen using JMODELTEST v. 2.1.2 (Posada 2008) based on Akaike and Bayesian information criteria. Two independent runs with four Markov Chain Monte Carlo simulations were performed for 30 million iterations and sampled every 1000th iteration. The first 25% of samples were discarded as burn-in. ML was analyzed on the CIPRES science gateway with 100 rapid bootstrap replicates (Miller et al. 2010) (https://www.phylo.org/portal2). Outgroups follow Chen et al. (2022).

Table 2.
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Information for samples used in phylogenetic analyses in this study. Type locality indicated by an asterisk (*). NNU represents Nanning Normal University; SYS, Sun Yat-sen University; MNHN, Muséum National d’Histoire Naturelle, Paris; NHMG, Natural History Museum of Guangxi; CIB, Chengdu Institute of Biology, Chinese Academy of Sciences.

ID Species Locality Voucher no. 16S COI References
1 N. guibeiensis sp. nov. China: Guangxi: Xing’an: Maoershan (paratype) NNU 00917 ON985180 ON968962 This study
2 N. guibeiensis sp. nov. China: Guangxi: Xing’an: Maoershan (paratype) NNU 00918 ON985181 ON968963 This study
3 N. guibeiensis sp. nov. China: Guangxi: Xing’an: Maoershan (paratype) NNU 00919 ON985182 ON968964 This study
4 N. guibeiensis sp. nov. China: Guangxi: Xing’an: Yanguang (paratype) NNU 00810 ON985179 ON968961 This study
5 N. guibeiensis sp. nov. China: Guangxi: Xing’an: Zhongfeng (paratype) NNU 00694 ON985176 ON968958 This study
6 N. guibeiensis sp. nov. China: Guangxi: Xing’an: Zhongfeng (paratype) NNU 00769 ON985177 ON968959 This study
7 N. guibeiensis sp. nov. China: Guangxi: Xing’an: Zhongfeng (paratype) NNU 00770 ON985178 ON968960 This study
8 N. guibeiensis sp. nov. China: Guangxi: Xing’an: Zhongfeng (paratype) NNU 00867 ON985183 ON968965 This study
9 N. adenopleura China: Taiwan: Taichung City SYS a007358 MN946445 MN945201 Lyu et al. (2020a)
10 N. adenopleura China: Taiwan: Taichung City SYS a007359 MN946446 MN945202 Lyu et al. (2020a)
11 N. chapaensis Vietnam: Lao Cai: Sapa* MNHN 2000.4850 KR827711 KR087625 Grosjean et al. (2015)
12 N. daunchina China: Sichuan: Mt Emei* SYS a004594 MF807822 MF807861 Lyu et al. (2020a)
13 N. daunchina China: Sichuan: Mt Emei* SYS a004595 MF807823 MF807862 Lyu et al. (2020a)
14 N. guangdongensis China: Guangdong: Yingde City (holotype) SYS a005767 MN946406 MN945162 Lyu et al. (2020a)
15 N. guangdongensis China: Guangdong: Yingde City (paratype) SYS a005768 MN946407 MN945163 Lyu et al. (2020a)
16 N. guangxiensis China: Guangxi: Mt Daming (paratype) NHMG 202007001 MZ677222 MZ678729 Lyu et al. (2021)
17 N. guangxiensis China: Guangxi: Mt Daming (paratype) NHMG 202007002 MZ677223 MZ678730 Lyu et al. (2021)
18 N. hainanensis China: Hainan: Mt Diaoluo* SYS a007669 MN946451 MN945207 Lyu et al. (2020a)
19 N. hainanensis China: Hainan: Mt Diaoluo* SYS a007670 MN946452 MN945208 Lyu et al. (2020a)
20 N. leishanensis China: Guizhou: Mt Leigong* SYS a007908 MN946453 MN945209 Lyu et al. (2021)
21 N. leishanensis China: Guizhou: Mt Fanjing SYS a007195 MN946454 MN945210 Lyu et al. (2021)
22 N. leishanensis China: Guizhou: Mt Fanjing SYS a007196 MN946455 MN945211 Lyu et al. (2021)
23 N. leishanensis China: Guangxi: Mt Jiuwan NHMG 202007021 MZ677227 MZ678734 Lyu et al. (2021)
24 N. leishanensis China: Guangxi: Mt Jiuwan NHMG 202007022 MZ677228 MZ678735 Lyu et al. (2021)
25 N. leishanensis China: Guangxi: Mt Jiuwan NHMG 202007023 MZ677229 MZ678736 Lyu et al. (2021)
26 N. leishanensis China: Guangxi: Mt Jiuwan NHMG 202007025 MZ677230 MZ678737 Lyu et al. (2021)
27 N. lini China: Yunnan: Jiangcheng County* SYS a003967 MF807818 MF807857 Lyu et al. (2017)
28 N. lini China: Yunnan: Jiangcheng County* SYS a003968 MF807819 MF807858 Lyu et al. (2017)
29 N. mangveni China: Zhejiang: Mt Dapan (paratype) SYS a006310 MN946424 MN945180 Lyu et al. (2020a)
30 N. mangveni China: Zhejiang: Mt Dapan (paratype) SYS a006311 MN946425 MN945181 Lyu et al. (2020a)
31 N. nankunensis China: Guangdong: Mt Nankun (paratype) SYS a005718 MF807839 MF807878 Lyu et al. (2017)
32 N. nankunensis China: Guangdong: Mt Nankun (holotype) SYS a005719 MF807840 MF807879 Lyu et al. (2017)
33 N. occidentalis China: Yunnan: Mt Gaoligong (paratype) SYS a003775 MF807816 MF807855 Lyu et al. (2020a)
34 N. occidentalis China: Yunnan: Mt Gaoligong (holotype) SYS a003776 MF807817 MF807856 Lyu et al. (2020a)
35 N. okinavana Japan: Okinawa: Iriomote Island* Unknown NC022872 NC022872 Kakehashi et al. (2013)
36 N. pleuraden China: Yunnan: Kunming City* SYS a007858 MT935683 MT932858 Lyu et al. (2020b)
37 N. pleuraden China: Yunnan: Wenshan City SYS a007717 MT935671 MT932850 Lyu et al. (2020b)
38 N. shiwandashanensis China: Guangxi: Shangsi County (holotype) NNU00238 MZ787977 MZ782098 Chen et al. (2022)
39 N. shiwandashanensis China: Guangxi: Shangsi County (paratype) NNU00239 MZ787978 MZ782099 Chen et al. (2022)
40 N. xiangica China: Hunan: Mt Dawei (paratype) SYS a006491 MN946433 MN945189 Lyu et al. (2020a)
41 N. xiangica China: Hunan: Mt Dawei (holotype) SYS a006492 MN946434 MN945190 Lyu et al. (2020a)
42 N. xiangica China: Hunan: Mt Dawei (paratype) SYS a006493 MN946435 MN945191 Lyu et al. (2020a)
43 N. xiangica China: Hunan: Mt Yangming (paratype) SYS a007269 MN946436 MN945192 Lyu et al. (2020a)
44 N. xiangica China: Hunan: Mt Yangming (paratype) SYS a007270 MN946437 MN945193 Lyu et al. (2020a)
45 N. xiangica China: Hunan: Mt Yangming (paratype) SYS a007271 MN946438 MN945194 Lyu et al. (2020a)
46 N. xiangica China: Hunan: Mt Yangming (paratype) SYS a007272 MN946439 MN945195 Lyu et al. (2020a)
47 N. xiangica China: Hunan: Mt Yangming (paratype) SYS a007273 MN946440 MN945196 Lyu et al. (2020a)
48 N. xiangica China: Jiangxi: Mt Wugong (paratype) SYS a002590 MN946441 MN945197 Lyu et al. (2020a)
49 N. xiangica China: Guangxi: Mt Dupangling SYS a006568 MN946442 MN945198 Lyu et al. (2020a)
50 N. xiangica China: Guangxi: Mt Dupangling SYS a006569 MN946443 MN945199 Lyu et al. (2020a)
51 N. xiangica China: Guangxi: Mt Dupangling SYS a006570 MN946444 MN945200 Lyu et al. (2020a)
52 N. yaoica China: Guangxi: Mt Dayao (paratype) SYS a007020 MK882276 MK895041 Lyu et al. (2019)
53 N. yaoica China: Guangxi: Mt Dayao (paratype) SYS a007021 MK882277 MK895042 Lyu et al. (2019)
54 N. yeae China: Guizhou: Tongzi County (paratype) CIB TZ20190608005 MN295228 MN295234 Wei et al. (2020)
55 N. yeae China: Guizhou: Tongzi County (paratype) CIB TZ20160714016 MN295231 MN295237 Wei et al. (2020)
56 Babina holsti Japan: Okinawa* Unknown NC022870 NC022870 Kakehashi et al. (2013)
57 Babina subaspera Japan: Kagoshima: Amami Island* Unknown NC022871 NC022871 Kakehashi et al. (2013)

Bioacoustics analysis

Advertisement calls of five individuals were recorded in the fields using a SONY ICX–0471 recorder on 7 May, and 3 and 28 June 2022. The ambient temperature was measured with a digital hygrothermograph. Calls were recorded at 21 °C, 23 °C, and 18 °C. Advertisement calls were analyzed using Raven Pro v. 1.6 (Cornell Laboratory of Ornithology, USA) as per Köhler et al. (2017). The acoustic properties were set to a window size of 512 points, fast Fourier transform, and Hanning window with no overlap. We performed the following measurements: call duration (measured from the beginning to the end of the call), note duration (measured from the beginning to the end of the note), inter-note interval (measured from the end of one note to the beginning of the consecutive note), and dominant frequency (the peak frequency of the call). The published bioacoustics data were obtained from the literature (Table 1).

Results

PCA results were shown in Fig. 2. The extracted components PC1 eigenvectors accounted for 29.6% of the variance, PC2 for 16.8%, and PC3 for 13.3%. The new specimens can be significantly distinguished from N. leishanensis and N. xiangica. The results of Mann-Whitney U tests implied that the new specimens were significantly different from N. leishanensis and N. xiangica on many morphometric characters, including HDW, SNT, IND, ED, and FTL for N. leishanensis, and SNT, IND, ED, HND, FTL, and TIB for N. xiangica (Table 3). Additionally, the new specimens can be easily identified by a series of diagnostic characters, such as relatively larger body size, smooth dorsum with tubercles on the posterior of the back, lateroventral grooves present on all fingers and toes but not on Finger I, and tibiotarsal articulation reaching the level between the eye and nostril (Table 1). ML and BI analyses led to identical topologies based on the two mtDNA fragments (Fig. 3). Phylogenetical trees indicated that our newly collected specimens were strongly clustered into a monophyletic group and sister to N. leishanensis with robust support (PP = 0.97, BS = 94).

Figure 2. 

Scatter plot of PC1 and PC2 of PCA based on the morphometric measurements, distinguishing Nidirana guibeiensis sp. nov., N. leishanensis, and N. xiangica.

Figure 3. 

Maximum-likelihood tree based on 16S + COI fragments with bootstrap supports/Bayesian posterior probabilities on branches.

Table 3.
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Measurements of Nidirana guibeiensis sp. nov. (in mm) and morphometric comparisons with N. leishanensis and N. xiangica. Abbreviations defined in Material and methods. * p-values < 0.05, ** p-values < 0.01.

Male (n = 13) Female p-values from Mann-Whitney U tests
Ranging mean ± SD NNU 00694 New species vs N. leishanensis New species vs N. xiangica
SVL 50.2–63.6 55.2 ± 3.7 54.6 0.054 0.215
HDL 18.6–22.5 19.9 ± 1.1 19.8 0.380 0.137
HDW 18.3–23.1 20.1 ± 1.4 17.3 0.011* 0.321
SNT 7.0–9.5 7.5 ± 0.7 7.0 0.002** 0.003**
IND 5.3–6.6 6.0 ± 0.5 5.3 0.028* 0.000**
IOD 4.1–5.6 4.9 ± 0.4 5.0 0.661 0.121
ED 4.4–6.9 5.6 ± 0.7 5.4 0.028* 0.000**
TD 4.6–6.3 5.4 ± 0.5 4.8 0.161 0.094
HND 12.6–15.6 14.0 ± 1.0 14.3 0.726 0.041*
FTL 26.2–33.4 29.3 ± 2.0 30.5 0.001** 0.000**
TIB 25.7–29.9 28.4 ± 1.5 30.8 0.861 0.001**

The call spectrograms of the new specimens are shown in Fig. 4. Three types of calls were recorded: calls with one note (Type A), two notes (Type B), and three notes (Type C). Note number of a call, call duration, note duration, and inter-note duration are listed in Table 4. The newly collected specimens had different call characteristics distinguishing them from the call types of their congeners, a call with notes, call duration, and note duration (Table 4). However, the calls of N. leishanensis consist of one strophe with one syllable, and call durations last 330–430 ms. The calls of N. xiangica consist of two or three notes, and call durations last 331.9–624.8 ms. The two species above are distinct from the new specimens based on acoustic data (Table 4).

Figure 4. 

Advertisement call spectrograms of Nidirana guibeiensis sp. nov.

Table 4.
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Vocalization parameters of Nidirana guibeiensis sp. nov.

Type A (n = 21) Type B (n = 37) Type C (n = 13)
Notes number of one call 1 2 3
Call duration 155–232, (mean 179.6 ± 24.4) ms 349–471, (mean 383.4 ± 28.6) ms 561–777, (mean 658.6 ± 71.7) ms
First note duration 153–210, (mean 167.4 ± 13.1) ms 160–210, (mean 187.8 ± 18.4) ms
Second note duration 71–90, (mean 79.3 ± 4.2) ms 71–86, (mean 77.2 ± 5.5) ms
Third note duration 75–93, (mean 82 ± 5.3) ms
First inter-note duration 125–171, (mean 136.6 ± 11.3) ms 113–167, (mean 139.5 ± 18.3) ms
Second inter-note duration 142–221, (mean 171.8 ± 24.1) ms

Phylogeny, bioacoustics, and morphology support the recognition of the newly collected specimens from northern Guangxi as a previously undescribed Nidirana species, which is described below.

Taxonomic account

Nidirana guibeiensis Chen, Ye, Peng & Li, sp. nov.

https://zoobank.org/22E61BF0-A1D2-4E44-83A5-F4EA99A50B78
Fig. 5

Holotype

NNU 00771; adult ♂; China, Guangxi, Ziyuan County, Zhongfeng Town; 110.6882°E, 25.9750°N; Wei-Cai Chen leg., 8 May 2022.

Paratypes

NNU 00769–770, 772–773; 4 adult ♂♂; same locality and date as holotype • NNU 00694; 1 adult ♀; same locality as holotype; Gui-Dong Yu leg., 29 April 2022 • NNU 00810; 1 adult ♂; China, Guangxi, Xing’an County, Lingtan Town; 110.5622°E, 25.5907°N; Wei-Cai Chen leg., 9 May 2022 • NNU 00864–867; 4 adult ♂♂; same locality as holotype; Wei-Cai Chen leg., 2 June 2022 • NNU 00917–919; 3 adult ♂♂; Maoershan National Nature Reserve; 110.4937°E, 25.8823°N; Wei-Cai Chen, Tong-Ping Su & Gui-Dong Yu leg., 28 June 2022.

Etymology

The species name refers to its distribution in northern Guangxi. ‘Guibei’ means northern Guangxi.

We suggest the English name Guibei Music Frog and the Chinese name Gui Bei Qin Wa (桂北琴蛙).

Diagnosis

Nidirana guibeiensis sp. nov. differs from its congeners in the combination of the following characteristics: larger body size (SVL 50.2–63.6 mm in males; 54.6 mm in the only sampled female); dorsum smooth with tubercles on the posterior of the back; surfaces of throat, chest, and upper part of the belly with grey clouding, lower part of the belly near immaculate creamy white; dorsal midline with a discrete creamy-white line; lateroventral grooves present on all fingers and toes but not on Finger I; tibiotarsal articulation reaching the level between eye and nostril; a pair of subgular vocal sacs present; three types of calls: one note, two notes, or three notes.

Description of holotype

Adult male, SVL 56.3 mm; head length slightly larger than width (HDL/HDW = 1.05); snout oval, significantly protruding beyond lower jaw; canthus rostralis distinct; loreal region concave; nostril oval and closer to snout than eye, laterally opening; a creamy white stripe on upper lip, beginning at the tip of snout along with upper lip and ending above insertion of arm; supratympanic fold visible; IOD/IND = 0.72; eye diameter almost equal to tympanum diameter (ED/TD = 1.02); vomerine teeth oval, closer to each other than to choana; tongue pyriform with a deep notch on posterior; a pair of subgular vocal sacs present. Relative finger lengths: II < IV < I < III; all tips of fingers but Finger I slightly dilated with lateroventral grooves; finger webbing and dermal fringes absent; subarticular tubercles prominent and conical; two palmar tubercles distinct; nuptial pad present on lateral Finger I with velvety spinules, extending from hand base to level of subarticular tubercle. Relative toe lengths: I < II < V < III < IV; all tips of toes slightly dilated, forming elongated and pointed discs with lateroventral grooves; toe webbing formula: I 2 – 2 II 1½ – 3– III 2 – 3+ IV 3+ – 2– V; toes with lateral fringes; subarticular tubercles prominent and oval; inner metatarsal tubercles elongated, but outer metatarsal tubercles conical; heels not meeting when thighs are held at right angles to body; tibiotarsal articulation reaching the level between eye and nostril. Dorsum smooth with tubercles on the posterior of the back; hindlimbs smooth with several tubercles; dorsolateral fold beginning at the posterior of eye and ending above groin; pineal gland distinct; flanks with suprabrachial glands at each side; peripheral vent with some small tubercles (Fig. 5A–F).

Figure 5. 

The holotype of Nidirana guibeiensis sp. nov. (NNU 00771) A dorsal view B dorsolateral view C ventral view D ventral view of hand E ventral view of foot F tubercles on the rear of the back.

Color of holotype

Alive, dorsum moss grey without spots; pineal gland light yellow; dorsal midline with a discrete creamy-white line, beginning at pineal gland and ending at vent; tympanum light brown; presence of a creamy-white linear gland on upper jaw; maxillary gland creamy-white; flank with several black spots and tubercles, and a large grey suprabrachial gland; thigh and tibia with three distinct black bars; surfaces of throat, chest, and upper part of belly with grey clouding, lower part of belly near immaculate creamy white; ventral limbs incarnadine; anterior of base of forelimb with a dark stripe; the anterior and posterior of iris reddish-brown, whereas the upper and lower part of iris brown (Fig. 5A–F). In preservation, dorsal surface faded to deep grey; black spots turned darkish black.

Variations

Measurements of type series are listed in Table 3 and Table S1. Paratypes were similar to the holotype in morphology and color pattern. Some had a discrete, rusty dorsal midline, and a rusty line along the dorsolateral folds, and rusty blotches on the flanks (Fig. 6A). Various tubercles on the rear of the back, some denser but some sparse (Fig. 6B). Some had five or six black bars on thigh and tibia (Fig. 6C).

Figure 6. 

Nidirana guibeiensis sp. nov. A dorsal view of NNU 00769 B rough tubercles on the rear of the back (NNU 00865) C five bars on thigh and tibia (NNU 00867) D female with creamy yellow eggs with pigmented poles (NNU 00694).

Ecology and distribution

Nidirana guibeiensis sp. nov. was found in paddy fields or still pools at 300–1 300 m a.s.l. We heard the advertisement calls in the field during the surveys, from April to July. We observed that the new species has nest construction behavior (Fig. 7). The nest was made of rice stems and 20–30 cm in diameter without a covering. Eggs were observed in the nest (Fig. 7). Females were gravid with creamy-yellow eggs with black poles (Fig. 6D). The new species is widespread in northern Guangxi.

Figure 7. 

Nests of Nidirana guibeiensis sp. nov. with eggs.

Comparison

A summary of morphological characteristics is listed in Table 1. Nidirana guibeiensis sp. nov. differs from its congeners in the following characteristics: (1) SVL 50.2–63.6 mm in males; (2) dorsum smooth with tubercles on the posterior of the back; (3) surfaces of throat, chest, and upper part of belly with grey clouding, lower part of belly near immaculate creamy white; (4) dorsal midline with a discrete creamy-white line; (5) tibiotarsal articulation reaching the level between eye and nostril; (6) lateroventral grooves present on all fingers and toes but not on Finger I; (7) nuptial pad present on Finger I; (8) a pair of subgular vocal sacs present; (9) a call comprised of one, two, or three notes.

Phylogenetically, N. guibeiensis sp. nov. is closest to N. leishanensis (Fig. 3). However, N. guibeiensis sp. nov. differs from N. leishanensis in the absence of dermal fringes on fingers (vs broad lateral fringes on inner sides of Fingers II, III, and IV but absent on Finger I); the presence of lateroventral grooves on all fingers except Finger I (vs lateroventral grooves present on Fingers III and IV); toe webbing formula: I 2 – 2 II 1½ – 3– III 2 – 3+ IV 3+ – 2– V (vs I 1⅓ –2 II 1⅓ –2⅓ III 1⅔ –3 IV 3⅓ –1⅓ V); dorsum smooth with tubercles on the posterior of the back (vs dorsal skin rough with dense granules but not concentrated on the posterior of the back); heels not meeting when thighs are held at right angles to body (vs heels overlapping); outer metatarsal tubercle present (vs absent); surfaces of throat, chest, and upper part of belly with grey clouding, lower part of belly near immaculate creamy white (vs surface and throat smooth and incarnadine); supratympanic fold present (vs absent); two palmar tubercles distinct (vs three palmar tubercles elliptic, distinct); nuptial pad present on Finger I (vs nuptial pad on the inner side of base of Fingers I and II); a call comprised of one, two, or three notes (vs a call with one strophe with one syllable). The new species has nest construction behavior (vs no nest construction behavior).

Nidirana guibeiensis sp. nov. differs from N. xiangica in having a dorsal midline with a discrete creamy-white line (vs absent dorsal midline); heels not meeting when thighs are held at right angles to body (vs heels meeting); a smooth dorsum with tubercles on the posterior of the back (vs extremely rough dorsal surface with dense tubercles and white horny spinules on the entire dorsum); supratympanic fold present (vs absent); the anterior and posterior of iris reddish-brown, whereas the upper and lower parts of iris brown (vs upper ⅓ iris brownish-white and lower ⅔ iris reddish-brown); advertisement calls contained 1–3 notes (vs 2 or 3 notes with a specific first note). Additionally, the new species has nest construction behavior (vs no nest construction behavior).

Due to the larger body size in males (SVL 50.2–63.6 mm), N. guibeiensis sp. nov. differs from males of N. nankunensis (SVL 33.3–37.1 mm), N. chapaensis (SVL 35.5–42.5 mm), N. guangxiensis (SVL 40.2–47.6 mm), N. hainanensis (SVL 32.8–44.4 mm), N. okinavana (SVL 35.5–42.8 mm), N. yaoica (SVL 42.1–45.6 mm), and N. yeae (SVL 41.2–43.5 mm). Other differences are (Table 1): lateroventral grooves present on all fingers but not on Finger I (vs absent in N. guangxiensis, N. occidentalis, N. pleuraden, and N. yeae); lateroventral grooves present on all toes (vs absent on all toes in N. pleuraden and N. occidentalis) ; tibiotarsal articulation reaching the level between the eye and nostril (vs beyond the tip of snout in N. lini); dorsum smooth with tubercles on the posterior of the back (vs no tubercles on the posterior of the back in N. chapaensis, N. daunchina, N. guangdongensis, N. guangxiensis, N. hainanensis, N. leishanensis, N. nankunensis, N. okinavana, N. shiwandashanensis, N. yaoica, and N. yeae); single nuptial pad present on Finger I (vs absent in N. hainanensis; two parts on Finger I in N. chapaensis; present on Fingers I and II in N. leishanensis).

Discussion

To date, six recognized Nidirana species have been reported from Guangxi, indicating an impressive species diversity. Lyu et al. (2021) and Chen et al. (2022) pointed out that Nidirana species have relatively narrow ranges. Rivers and mountains contribute to the speciation of Nidirana. Phylogenetically, the new species, N. leishanensis, and N. xiangica are clustered together. The three species are geographically close but occur in different mountain ranges (Fig. 1). Nidirana guibeiensis sp. nov. occurs in the Yuechengling Mountains, N. leishanensis in the Miaoling Mountains, and N. xiangica in the Dupanling, Dawei, and Yangming mountains. The other three species of Nidirana occurring in Guangxi (N. guangxiensis, N. shiwandashanensis and N. yaoica) resemble the abovementioned species but occur in separated mountain ranges (Lyu et al. 2019, 2021; Chen et al. 2022).

The discovery of the new species indicates that the diversity of the genus Nidirana is still underestimated. The taxonomic validity of reports of N. adenopleura in Guangxi must be reconsidered. In recent years, we have carried out a series of field surveys, but no evidence supports the occurrence of N. adenopleura in Guangxi. Nidirana guibeiensis sp. nov., which was reported by Mo et al. (2014) as N. adenopleura, is a good example of a misidentification of a new Nidirana species.

Acknowledgements

The authors are grateful to the Maoershan National Nature Reserve staff for assistance during fieldwork. The authors also thank to Annemarie Ohler and the anonymous reviewers for their valuable comments on the manuscript. This work was supported by the National Natural Science Foundation of China (32060116) and Guangxi Natural Science Foundation, China (2020GXNSFDA238022).

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Appendix 1

Table A1.
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Specimens examined. Type locality indicated by an asterisk (*).

Species Locality Voucher no.
Nidirana guangxiensis Damingshan National Nature Reserve, Guangxi, China* NNU 00213
Nidirana guangxiensis Damingshan National Nature Reserve, Guangxi, China* NNU 00214
Nidirana guangxiensis Damingshan National Nature Reserve, Guangxi, China* NNU 00215
Nidirana guangxiensis Damingshan National Nature Reserve, Guangxi, China* NNU 00216
Nidirana guangxiensis Damingshan National Nature Reserve, Guangxi, China* NNU 00217
Nidirana guangxiensis Damingshan National Nature Reserve, Guangxi, China* NNU 00218
Nidirana leishanensis Yuanbaoshan National Nature Reserve, Guangxi, China NNU 201908032
Nidirana leishanensis Yuanbaoshan National Nature Reserve, Guangxi, China NNU 201908033
Nidirana shiwandashanensis Shiwandashan National Nature Reserve, Guangxi, China* NNU 00238
Nidirana shiwandashanensis Shiwandashan National Nature Reserve, Guangxi, China* NNU 00239
Nidirana shiwandashanensis Shiwandashan National Nature Reserve, Guangxi, China* NNU 00605
Nidirana shiwandashanensis Shiwandashan National Nature Reserve, Guangxi, China* NNU 00606
Nidirana yaoica Dayaoshan National Nature Reserve, Guangxi, China* NNU 201907008
Nidirana yaoica Dayaoshan National Nature Reserve, Guangxi, China* NNU 201907009

Supplementary material

Supplementary material 1 

Supplementary data

Wei-Cai Chen, Jian-Ping Ye, Wan-Xiao Peng, Peng Li, Tong-Ping Su, Gui-Dong Yu, Zhi-Ying Cheng

Data type: Morphological.

Explanation note: Measurements of Nidirana guibeiensis sp. nov. (in mm). Abbreviations defined in Material and methods. *= holotype.

This dataset is made available under the Open Database License (http://opendatacommons.org/licenses/odbl/1.0/). The Open Database License (ODbL) is a license agreement intended to allow users to freely share, modify, and use this Dataset while maintaining this same freedom for others, provided that the original source and author(s) are credited.
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