A new species of the genus Gracixalus (Amphibia: Anura: Rhacophoridae) from Southern Guangxi, China Author Mo, Yunming Author Zhang, Wei Author Luo, Yu Author Zhou, Shichu Author Chen, Weicai text Zootaxa 2013 3616 1 61 72 journal article 10.11646/zootaxa.3616.1.5 3cc81418-e23b-4ae3-999c-7f334682fdf6 1175-5326 283369 659E6372-9311-4E50-9B04-9D2729E5C111 Gracixalus nonggangensis sp. nov. ( Fig. 2 ) Holotype . NHMG200809044 ( Fig. 2 ), adult male, from Nonggang National Nature Reserve, Longzhou country, Guangxi, China , near the Sino-Vietnam border ( 22.5233º N , 106.9523º E , alt. 216 m a.s.l) ( Fig. 1 A), collected by Yunming Mo, Wei Zhang and Shichu Zhou on 7 September 2008 . Paratypes . NHMG200809043, adult male collected near locality of the holotype ( 22.4567º N , 106.9742º E , alt. 432 m a.s.l) on 7 September 2008 ; NHMG 20091001 , NHMG 20091003 , NHMG 20091009 –10, adult males, and NHMG 20091002 , adult female, from the same site as the holotype , collected by Yunming Mo, Wei Zhang and Shichu Zhou on 14 October 2009 . NHMG201005008 and NHMG 111024 , adult females, from the same site as the holotype , collected by Yunming Mo, Weicai Chen and Wei Zhang on 7 May 2010 , and 14 October 2011 . NHMG1005046, adult male, from Longrui station of Nonggang National Nature Reserve, Ningming county ( 22.2463° N , 107.0729° E , alt. 252 m a.s.l), collected by Yunming Mo, Wei Liao and Zhuqiu Song on 10 May 2010 . NHMG 111025 , adult male, from the same site as the holotype , collected by Yunming Mo, Weicai Chen and Wei Zhang on 14 October 2011 . Diagnosis. The new species is assigned to the genus Gracixalus by the tips of digits expanded into large disks, vomerine teeth absent, horizontal pupil, two outer palmar tubercles, tibiotarsal articulation reaching the tip of snout. Gracixalus nonggangensis sp. nov. can be distinguished from all other species of Gracixalus by the following combination of characters: medium size (SVL ranging from 29.9–35.3 mm for males, 33.6–38.0 mm for female); vomerine teeth absent; distinct tympanum as wide as the disc of finger III; lower part of tympanum with many small tubercles; tibiotarsal articulation reaching the tip of snout; dermal fringes along outer side of forearm, tibia and tarsus absent; smooth, yellowish-olive dorsum in life, with a wide dark-green irregular mark; throat, chest and belly white with light grey-blue tint and brown marbling in life; dark green, broad transverse stripes on limbs: 2 on the forearm, 3 on the thigh and tibia; finger webbing absent, toes one-third webbed; male with internal subgular vocal sac. Description of holotype . Adult male (SVL 30.0 mm), body moderately elongate; head slightly longer than wide (HL 12.0 mm; HW 11.2 mm ; MN 10.0 mm; MFE 7.8 mm ; MBE 3.4 mm ); snout rounded in dorsal view and slightly protruding, SL ( 4.7 mm ) longer than horizontal diameter of eye (EL 3.8 mm ); canthal edges rounded, loreal region oblique, slightly concave; nostrils oval, markedly protuberant, directed laterally, closer to tip of snout (NS 1.6 mm ) than to eye (EN3.0 mm); internasal distance (IN 2.9 mm ) as wide as upper eyelid (UEW 2.8 mm ), less than interorbital space (IUE 3.7 mm ); internarial space concave; distance between front of eyes (IFE 5.9 mm ) about 60% distance between back of eyes (IBE 9.9 mm ); pupil horizontally oval; tympanum ( TYD 1.6 mm ) distinct, rounded, 42% of eye diameter, and equal to the disc of finger III ( 1.6 mm ); tympanum-eye distance (TYE 0.9 mm ), 56% of tympanum diameter; supratympanic fold indistinct, extending from behind of eye to above the shoulder; parotoid glands absent; cephalic ridges absent; co-ossified skin on the head absent; vomerine teeth absent; tongue pear-shaped with deep notch posteriorly. Forelimbs short and gracile ( Fig. 2 ; Fig. 3 A); forearm (FLL 5.9 mm ) shorter than hand (HAL 8.9 mm ), hand 67% as long as foot (FOL 13.2 mm ); forearm and hand length ( 14.8 mm ) slightly smaller than half of SVL; dermal fringe along outer side of forearm absent; fingers I short and thin; fingers II, III (TFL 5.4 mm ) and IV long and thin; relative length of fingers: I <II<IV<III ( Fig. 3 A); tips of all fingers with moderate discs, with circummarginal grooves, almost two times the width of finger (fd1/fw1 1.4/ 0.7 mm ; fd2/fw2 1.9/1.0 mm; fd3/fw3 2.3/ 1.1 mm ; fd4/ fw4 2.3/ 1.1 mm ); finger webbing absent; subarticular tubercles distinct, bluntly rounded: one on fingers I and II, and two on fingers III and IV; prepollex distinct, oval; two small palmar tubercles, the outer indistinct; supernumerary tubercles distinct on finger II, III, IV, absent on fingers I. Hindlimbs moderately long ( Fig. 2 ; Fig. 3 B); heels slightly overlapping when legs are folded at right angle to body; when adpressed to body, the tibiotarsal articulation reaches to the tip of snout; tibia 4.7 times longer (TL 16.5 mm ) than wide (TW 3.9 mm ), longer than thigh (FL 15.8 mm ) and foot length (FOL 13.2mm ), approximately 44% of SVL; length of foot and tarsus (TFOL 21.7 mm ) about 72% of SVL; toes moderately long and thin, toe IV (FTL 7.1 mm ) 33% the distance from base of tarsus to tip of toe IV; relative length of toes: I<II<III<V<IV; tips of all toes with small discs, with distinct circummarginal grooves, rather wide compared to toe width, discs almost one and a half times width of toes (td1/tw1 1.2/ 0.9 mm ; td2/tw2 1.5/ 0.9 mm ; td3/tw3 1.5/ 0.9 mm ; td4/tw4 2.0/1.0 mm; td5/tw5 1.8/ 0.9 mm ); webbing formula I 2–2 1/ 2 II 1 1/2 –3+ III 2 –3 1/ 2 IV 3+– 2- V (MTTF: 8.0 mm; MTFF 5.9 mm ; TFTF 8.4 mm ; FFTF 5.4 mm ); subarticular tubercles prominent, rounded, simple and all present: 1, 1, 2, 3, 2; tarsal fold absent; supernumerary tubercles absent; inner metatarsal tubercle distinct (IMT 1.3 mm ), 1.5 times in length of toe I (ITL2.0 mm); outer metatarsal tubercle absent. FIGURE 2. Color of holotype in life (NHMG200809044). ( A) dorsal view, (B) dorsolateral view, (C) ventral view; Color of holotype in preservative. (D) dorsal view, (E) ventral view, (F) head from lateral view. Photos: Yunming Mo. FIGURE 3. Ventral surface of right hand (A), and right foot (B) of the preserved holotype (NHMG 200809044) of Gracixalus nonggangensis sp. nov. Photos: Yunming Mo. Dorsum and dorsal parts of head, forelimbs, and hindlimbs overall smooth but with some small tubercles; lower part of tympanum with many small tubercles; ventral part of forearm, tibia and tarsus smooth; dorsolateral folds absent; throat, chest, belly and ventral part of thighs granular; macroglands absent; dermal appendage at vent absent. Coloration in life. Dorsal parts of head and dorsum, flank, loreal region, tympanic region and tympanum yellowish-olive, a dark-green irregular patch running from between eyes to shoulder, divaricating to the posterior part of the dorsum; upper and lower lip yellowish-olive with some creamy white spots; dorsal part of forelimb, thigh, tibia and foot yellowish-olive with transverse olive bands, forelimbs with 3 and hindlimbs with 6; lower part of flanks and tympanum with some dark brown spots; throat and margin of throat, chest, and belly marbled with white, ventral part of forelimbs and hindlimbs greyish-white; iris olive with brown reticulations; pupil periphery lined with yellow ( Fig. 2 A, B, C). Coloration in formalin. Dorsal parts of head and dorsum, and upper part of flanks brownish grey; the dorsum with irregularly dark brown mark, a triangular spot between eyes bifurcating into two bands continuing posteriorly; lower flank beige with brown small spots; snout, canthus and loreal region, tympanum region, tympanum, and upper lip brown or brownish with white spots; vent region brown or brownish with white spots; forelimb, dorsal surface of thigh, tibia and foot light brown or grey with some dark brown bands; anterior and posterior part of thigh, and underneath the vent grey with small white spots; throat, margin of throat, chest and belly creamy white with brown marbling; ventral part of forelimbs and thighs creamy white; webbing greyish ( Fig. 2 D, E, F). Male secondary sexual characters. Nuptial pad absent; internal subgular vocal sac present; lineae masculinae absent. Measurements of holotype . SVL, 30.0; HL, 12.0; HW, 11.2; MN, 10.0; MFE, 7.8; MBE, 3.4; IFE, 5.9; IBE, 9.9; IN, 2.9; EN, 3.0; EL, 3.8; NS, 1.6; SL, 4.7; TYD , 1.6; TYE, 0.9; IUE, 3.7; UEW, 2.8; HAL, 8.9; FLL, 5.9; TEL, 5.4; fd3, 1.6; FL, 15.8; TL, 16.5; FOL, 13.2; FTL, 7.1; TFOL, 21.7; IMT, 1.3; ITL, 2.0; MTTF, 8.0; TFTF, 5.9; MTFF, 8.4; FFTF, 5.4. TABLE 2. Measurements (mm) of Gracixalus nonggangensis sp. nov. (Abbreviations see Material and Methods).

Character	Male	Female
Body and head	Range; mean ± S. D. (N=8)	Range; mean ± S. D. (N=3)
SVL	29.9–35.3; 32.4±2.2	33.6–38.0; 35.2±2.4
HL	11.1–13.2; 12.3±0.7	12.9–13.6; 13.1±0.4
HW	10.5–13.0; 11.7±0.8	12.2–12.7; 12.5±0.3
MN	10.0–12.1; 10.8±0.7	11.3–12.1; 11.6±0.4
MFE	7.1–9.0; 8.1±0.6	8.2–8.9; 8.5±0.4
MBE	3.2–4.7; 3.9±0.4	3.7–4.8; 4.3±0.6
IFE	5.9–7.5; 6.7±0.5	6.7–7.3; 6.9±0.3
IBE	9.9–11.6; 10.8±0.6	10.7±11.5; 11.2±0.4
IN	2.9–3.8; 3.4±0.3	3.6–3.9; 3.7±0.2
EN	3.0–3.6; 3.2±0.2	3.3–3.9; 3.5±0.3
EL	3.8–5.3; 4.7±0.5	4.8–5.3; 5.0±0.3
NS	1.6–2.3; 2.0±0.3	2.1–2.6; 2.3±0.3
SL	4.7–5.7;5.4±0.4	5.5–5.9; 5.7±0.2
TYD	1.5–2.1; 1.9±0.2	2.0–2.5; 2.2±0.3
TYE	0.9–1.3; 1.1±0.2	1.2–1.5; 1.3±0.2
IUE	3.3–4.3; 3.8±0.3	3.8–4.6; 4.1±0.5
UEW	2.8–3.6; 3.1±0.3	2.9–3.7; 3.3±0.4
Forelimbs
HAL	8.5–10.1; 9.5±0.6	9.6–11.2; 10.2±0.9
FLL	5.7–6.9; 6.4±0.4	5.9–7.6; 6.8±0.9
TEL	4.9–6.2; 5.7±0.4	5.7–6.4; 6.1±0.4
fd3	1.5–2.1; 1.8±0.2	1.7–2.0; 1.8±0.2
Hindlimbs
FL	15.5–18.7; 17.3±1.1	17.4–20.3; 18.7±1.5
TL	16.5–19.1; 18.0±0.9	19.0–20.9; 19.7±1.0
FOL	13.2–16.2; 14.4±1.0	14.5–16.8; 15.5±1.2
FTL	7.1–8.5; 7.7±0.5	7.5–8.8; 8.1±0.7
TFOL	21.2–24.6; 23.1±1.2	24.4–26.3; 25.1±1.0
IMT	1.2–1.9; 1.5±0.3	1.3–1.6; 1.5±0.2
ITL	1.9–2.9; 2.2±0.4	2.2–2.8; 2.4±0.3
Webbing
MTTF	7.8–8.8; 8.2±0.3	9.1–9.7; 9.4±0.3
TFTF	5.8–6.7; 6.3±0.3	5.4–7.3; 6.1±1.0
MTFF	8.1–9.3; 8.7±0.4	9.9–10.7; 10.2±0.5
FFTF	5.4–6.5; 5.9±0.3	5.4–7.2; 6.1±1.0

Variation. The tympanum almost equal to the width of the disc of finger III in males and obviously bigger than the disc of finger III in two female specimens (NHMG 20091002 and NHMG201005008); morphometrics and coloration in the five paratypes are similar to those of the holotype . Measurements of type series are shown in Table 2 . Habitat and distribution. The new species was found at elevations between 200– 500 m . Between May and October vocalizing males were encountered at night (18:00–23:00 h, air temperature <27°C) on bushes and grassy vegetation at a height of 0.5–1.0 m above the ground in secondary karst evergreen forest without nearby waterbodies ( Fig. 1 B). This species is only known from Nonggang National Nature Reserve, including Longzhou and Ningming country (about 22.2333°– 22.4667° N , 106.9500°– 107.0667° E , alt. 216–432 m a.s.l), in southern Guangxi, China . Etymology . This species is named after the locality in which it was collected. The suggested English name is the Nonggang small treefrog. Sequence divergence. Uncorrected sequence divergences between G. nonggangensis sp. nov. sequences and all homologous sequences available on the genus Gracixalus were greater than 6%. The genetic distance is smallest between G. nonggangensis sp. nov. and G. jinxiuensis ( EF564525 ), at 6.1%, whereas the intraspecific variation among G. nonggangensis sp. nov. range from 0 to 2.6%. Among them, three specimens (NHMG 20091009 –10, NHMG 111024 ) came from Nonggang station of Nonggang National Nature Reserve, which shared the same haplotype, whereas the genetically most divergent specimen (NHMG1005046) came from Longrui station of Nonggang National Nature Reserve, at a distance of about 20 km from Nonggang station. Preliminary hypothesis of phylogenetic relationship. Based upon our preliminary molecular data, the genus Gracixalus formed a monophyletic group with well-supported values (BP=98; BBP=1.00), and was divided two clades (Clade I and Clade II) with a strong support values ( Fig. 4 ), consistent with Rowley et al. (2011). Clade I consists of G. gracilipes , G. supercornutus , G. quangi , and G. quyeti (= Clade I in Rowley et al. (2011)); Clade II consists of G. nonggangensis sp. nov. , G. jinxiuensis , G. cf. jinxiuensis , Kurixalus cf. ananjevae , K. carinensis , K. odontotarsus and Gracixalus sp. (= Clade II in Rowley et al. (2011)). Among them, G. j i n x i u e n s i s did not form monophyletic groups, and indicated G. jinxiuensis group was likely a species complex. Based on the 16S ribosomal RNA mitochondrial gene, we suggested that G. nonggangensis sp. nov. is sister to this group ( G. cf. jinxiuensis , K. cf. ananjevae , K. carinensis , K. odontotarsus and Gracixalus sp.) with moderate supported values (BP=90; BBP=0.89). FIGURE 4. Bayesian posterior probability tree was reconstructed from 16S ribosomal RNA mitochondrial gene sequences with Philautus aurifasciatus , Kurixalus eiffingeri and K. odontotarsus as outgroups. Maximum-likelihood tree produced nearidentical topology. Two reliability indices are given on nodes: the Bayesian posterior probabilities/the maximum likelihood bootstrap percentages. Comparisons. We compared G. nonggangensis sp. nov. with all rhacophorid treefrogs with a SVL under 45 mm from China and adjoining countries. Firstly, G. nonggangensis sp. nov . can be distinguished from all other species of the genus Gracixalus . Morphologically, the new species is most similar to G. quyeti (Nguyen et al. 2008) . However, the species differs from G. quyeti by having a smooth, yellowish-olive dorsum, lacking tubercles (versus dorsal surface of head, back and upper portion of flanks covered with small sharp tubercles in G. quyeti ) (Nguyen et al. 2008). Molecularly, phylogenetic trees indicated that G. nonggangensis sp. nov. did not cluster with G. quyeti in the same clade ( Fig. 4 ), and the genetic distance (uncorrected p distance) between them reaches 12.7%. G. nonggangensis sp. nov. differs from G. j i n x i u e n s i s , from Dayaoshan National Nature Reserve, Guangxi, by having a larger size in adults (SVL 29.9–35.3 mm male versus 23.5 mm male in G. jinxiuensis ), and by having a yellowish-olive dorsum (versus brown dorsum in G. j i n x i u e n s i s ) (Hu et al. 1978; Fei et al . 2009). G. nonggangensis sp. nov. differs from G. gracilipes by having a larger size in adults (SVL 29.9–35.3 mm male versus SVL< 25 mm male in G. gracilipes ) and lacking white patch under the eye to the tympanum (present in G. gracilipes ) (Delorme et al. 2005; Fei et al. 2009). G. nonggangensis sp. nov. can be distinguished from G. medogensis by having tibiotarsal articulation reaching to the tip of snout (versus to the front of eye in G. medogensis ) (Fei et al. 2009). Furthermore, G. medogensis has a brown inverse “V” mark on the dorsum, and an orange mark on the flank of thigh (absent in G. nonggangensis sp. nov. ) (Fei et al. 2009). G. nonggangensis sp. nov. differs from G. quangi by having a rounded snout (versus a triangularly pointed snout in G. quangi ) and lacking black spots on the flanks and ventral surface of the thighs and a tibiotarsal projection (present in G. quangi ), and having a larger size in adults (SVL 29.9–35.3 mm male versus SVL< 25 mm male in G. quangi ) (Rowley et al. 2011). G. nonggangensis sp. nov. differs from G. supercornutus by having a larger size in adults (SVL 29.9–35.3 mm male versus SVL< 25 mm male in G. supercornutus ) and lacking prominent tubercles or spines on the whole dorsum and the upper eyelids (versus present in G. supercornutus ) (Orlov et al. 2004). The yellow-olive dorsum, finger webbing absent, toes one-third webbed, and lacking vomerine teeth and macroglands separate G. nonggangensis sp. nov. from all species of Buergeria , Kurixalus , Polypedates and Rhacophorus . Buergeria japonica has a brown dorsum (Fei et al. 2009). Buergeria oxycephala has fingers lightly webbed and toes fully webbed (Fei et al. 2009). The species of Chiromantis have an opposable finger (versus noopposable fingers in G. nonggangensis sp. nov. ). Feihyla palpebralis has a distinct, broad silvery white bands below the eyes, and a nuptial pad on finger I and II. Feihyla fuhua has a long and wide white streak from below eye to the base of shoulder (Fei et al. 2009; Fei et al. 2010). Kurixalus carinensis has distinct supratympanic fold and macrogland. Kurixalus ordontotarsus has dermal fringes and tubercles on the limbs, and sharp tubercles on the dorsum (Ye et al. 1993; Nguyen et al. 2008). Kurixalus eiffingeri has some white glands in the forearms and hindlimbs, and well developed vomerine teeth (Fei et al. 2009). Kurixalus bisacculus is dark brown with only a tubercle at the heel (Taylor 1962). Kurixalus verrucosus have a brown dorsum with only a tubercle at the heel (Boulenger 1893). Liuixalus hainanus has brown oval mark on the dorsum (Liu & Wu 2004). Liuixalus ocellatus has a brown dorsum with some glands (Liu & Hu 1973). Liuixalus romeri has a small size (SVL 15–18 mm in male versus 29.9–35.3 mm in male for G. nonggangensis sp. nov. ) (Smith 1953). Philautus abditus , Philautus cardamonus and Philautus kempii have a brown dorsum (Inger et al. 1999, Ohler et al. 2002, Annandale 1912). Philautus maosonensis has a brown dorsum, and truncated snout (Bourret 1937). Philautus truongsonensis has a brownish dorsum, an obtusely pointed snout (Orlov & Ho 2005). Philautus tytthus is dark greyish (Smith 1940). The species of Polypedates have vomerine teeth, and finger webbing. Rhacophorus arvalis has a large size (SVL ranging from 42–44 mm in male versus 29.9–35.3 mm in male for G. nonggangensis sp. nov. ), and finger webbing. Rhacophorus chenfui has white flank lines that clearly divide dorsal and ventral portions and purple flanks in life (Fei et al. 2009). Rhacophorus dugritei and Rhacophorus hungfuensis have vomerine teeth and finger webbing. Rhacophorus laoshan lack a green dorsum and has vomerine teeth. Rhacophorus leucofasciatus and Rhacophorus minimus have vomerine teeth, and a white line running from snout to the lateral of body and ending before the hindlimbs. Rhacophorus moltrechti has finger webbing. Rhacophorus rhodopus has vomerine teeth, and finger and toe fully webbed (Rao et al. 2006; Fei et al. 2009). Rhacophorus taipeianus has green dorsum with small glands, and finger wedding. Rhacophorus translineatus has triangularly pointed snout, and finger webbing. Rhacophorus verrucopus has a tibiotarsal projection, and finger webbing. Rhacophorus yaoshanensis has weak tuberculous dermal ridge along outer sides of forearm and tarsus, and finger webbing (Fei et al. 2009). Rhacophorus yinggelingensis has a lager size (SVL> 40 mm in male versus 29.9–35.3 mm in male for G. nonggangensis sp. nov. ), and the front of thigh is yellow and red-tinged, and the rear of thigh and inner side of tibia are red (Chou et al. 2007). Raorchestes kakachi has indistinct tympanum and blotches of dark brown on flanks (Seshadri et al. 2012). Raorchestes longchuanensis and Raorchestes menglaensis have a brown dorsum (Yang & Li 1978; Kou 1990). Raorchestes parvulus has a brownish grey dorsum (Boulenger 1893). Raorchestes resplendens has a bright orange to reddish coloration, multiple macroglands on the body and extremely short limbs (Biju et al. 2010). Theloderma andersoni , Theloderma asperum , Theloderma baibengensis , Theloderma kwangsiensis , Theloderma moloch , Theloderma lateriticum , Theloderma rhododiscus , and Theloderma stellatum lack a green dorsum or have some glands on the dorsum (Ahl 1927; Boulenger1886; Jiang et al. 2009; Bain et al. 2009; Liu & Hu 1962; Taylor 1962; Fei et al. 2009).