The fourth species of Leptobrachella (Anura, Megophryidae) found at Shiwandashan National Nature Reserve, Guangxi, China Author Chen, Wei-Cai https://orcid.org/0000-0002-2398-4079 Key Laboratory of Environment Change and Resources Use in Beibu Gulf Ministry of Education, Nanning Normal University, Nanning 530001, China chenweicai2003@126.com Author Li, Peng https://orcid.org/0000-0001-8311-0544 Key Laboratory of Environment Change and Resources Use in Beibu Gulf Ministry of Education, Nanning Normal University, Nanning 530001, China Author Peng, Wan-Xiao https://orcid.org/0000-0001-5635-9061 Key Laboratory of Environment Change and Resources Use in Beibu Gulf Ministry of Education, Nanning Normal University, Nanning 530001, China Author Liu, You-Jun https://orcid.org/0000-0001-7285-4943 Guangxi Key Laboratory of Earth Surface Processes and Intelligent Simulation, Nanning Normal University, Nanning 530001, China Author Huang, Yong https://orcid.org/0000-0002-3493-9468 Shiwandashan National Nature Reserve, Fangcheng 538000, China text ZooKeys 2024 2024-02-22 1192 257 279 http://dx.doi.org/10.3897/zookeys.1192.98352 journal article http://dx.doi.org/10.3897/zookeys.1192.98352 1313-2970-1192-257 96474EC442B14BEEA68D4F736A8A4EB9 8D4DA76882CC5FCDAE6B8906C96F03FD Leptobrachella guinanensis Chen, Li, Peng & Liu sp. nov. Figs 5 , 6 Material examined. Holotype . NNU 00876, adult male, collected at the Shiwandashan National Nature Reserve , Shangsi County , Guangxi , China ( 21°55'1.2"N , 107°54'10.8"E ; elevation 512 m ), collected by Wei-Cai Chen on 18 June 2022 . Paratypes . NNU 00560-561, two adult males, NNU 00557-559, three adult females, collected at the same locality as the holotype on 10 June 2021 ; NNU 00569-571, three adult females, collected at the same locality as the holotype on 1 July 2021 ; NNU 00875, one adult male, NNU 00877-880, four adult females, collected at the same locality and time as the holotype . All specimens were collected by Wei-Cai Chen. Etymology. The species name Leptobrachella guinanensis is derived from the geographic distribution of this species, specifically the southern Guangxi region. The suggested English name for this species is Gui Nan Leaf Litter Toad, while the Chinese name is Gui Nan Zhang Tu Chan (桂南掌突蟾). Diagnosis. Leptobrachella guinanensis sp. nov. can be distinguished from its congeners by a combination of the following characters: (1) SVL 30.5-32.5 mm in males; 38.7-41.8 mm in females; (2) 1/3 toe webbing, wide lateral fringes; (3) dorsal surface shagreened with small, raised tubercles and longitudinal ridges; (4) ventral surface creamy white without dark brown spots; (5) throat immaculate creamy white and its margin concentrated brown spots; (6) iris bicoloured, upper half light copper, transitioning to silver in lower half; (7) crossbars of hindlimbs with tubercles; (8) distinct dermal ridges under the toes; (9) a pair of glands under the vent; (10) tibia-tarsal articulation reaching to centre of eye; (11) relatively higher dominant frequency of advertisement calls (7.3-8.3 kHz). Description of holotype. Adult male, SVL = 30.5 mm, head width less than length (HW/HL = 0.93); snout protruding, projecting over the lower jaw; nostril oval, closer to the tip of snout than eye; canthus rostralis distinct; loreal region sloping and slightly concave; interorbital region flat; pupil vertical; eye diameter near equal to snout length (ED/SNT = 0.99); tympanum distinct and rounded, and its diameter conspicuously less than eye diameter (TD/ED = 0.41); supratympanic fold distinct, raised from corner of eye to supra-axillary gland; vomerine teeth absent; vocal sac openings located laterally on the floor of mouth; tongue with a shallow notch at the posterior tip. Tips of fingers rounded and slightly swollen; relative finger lengths I <II <IV <III; nuptial pad absent; subarticular tubercles absent; prominent inner palmar tubercle, separated from the small outer palmar tubercle; finger webbing and dermal fringes absent. Tips of toes rounded, slightly swollen; relative toe lengths I <II <V <III <IV; subarticular tubercles absent, replaced by distinct dermal ridges; pronounced large, oval inner metatarsal tubercle; outer metatarsal tubercle absent; 1/3 toe webbing; lateral fringes wide. TIB/SVL = 0.51; tibia-tarsal articulation reaching to the centre of eye; heels not meeting when thighs are appressed at right angles to body (Fig. 5 ). Figure 5. The holotype of L. guinanensis sp. nov. A dorsal view B ventral view C dorsolateral view D rear of the back and dorsal view of thighs E ventral view of hand F ventral view of foot. 1, tubercles on the crossbars; 2, femoral gland; 3, a pair of glands under the vent; 4, toe webbing; 5, wide lateral fringes on toe. Dorsal surface shagreened with small, raised tubercles and longitudinal ridges; belly and chest smooth without tubercles; anterior throat with several tubercles; ventral surface of limbs with creamy white tubercles; crossbars of hindlimbs with tubercles; flanks with several tubercles; pectoral glands oval, ~ 1.2 mm in diameter; femoral glands oval, ~ 1.3 mm in diameter, located on the posteroventral surface of thighs, closer to the knee than to the vent; supra-axillary glands distinct and rounded, ~ 0.9 mm in diameter; a pair of glands under the vent; and continued ventrolateral glandular line distinct (Fig. 5 ). Colour of holotype in life. Dorsal surface brown, an inverted triangle marking between eyes, irregular markings on shoulder and the rear of back; flanks with light orange tubercles; tympanum pale brown; supratympanic line black from posterior corner of eye to supra-axillary glands; posterior corner of eye silver; wide brown bars on upper lip; flanks with irregular black spots; brown transverse bars distinct on dorsal surface of forelimbs and hindlimbs; upper arm surfaces light orange; ventral surface creamy white without dark brown spots; throat immaculate creamy white and its margin concentrated brown spots; ventral surfaces of limbs purplish grey; pectoral and femoral glands, and a pair of creamy white glands under the vent, supra-axillary glands light orange; pupil black; iris bicoloured, upper half light copper, transitioning to silver in lower half (Fig. 5 ). Colour of holotype in preservative. Dorsum and limbs surfaces faded to a uniform grey; brown, inverted triangle marking distinctly visible between eyes; irregular black spots distinct on flanks; throat, chest, and belly creamy white; pectoral, femoral, supra-axillary, and ventrolateral glands creamy white; dark crossbars on limbs, fingers and toes remained distinct; upper arm and tibiotarsus faded to grey. Variation. Measurements of the type series are provided in Table 3 and Suppl. material 1: table S5. The black spots and tubercles on the flanks exhibit variation between individuals. Certain individuals possess more tubercles and longitudinal ridges on their dorsum and hindlimb surfaces (Fig. 6A ), while others display a light brown colouration on their dorsum (Fig. 6B ). Figure 6. A more tubercles and longitudinal ridges on dorsum and hindlimbs surfaces (NNU00875) B light brown on dorsum (NNU00569) C ventral view of the gravid female (NNU00880) D eggs creamy white without black poles. Ecology and distribution. Leptobrachella guinanensis sp. nov. was discovered in the evergreen forest at SWDS, at an elevation of 400-600 m. The individuals were observed near rocky streams between 20:00-24:00 h. Males were found calling while sitting on rocks near the stream ~ 0.5-1.0 m. Females were found to be gravid with creamy white eggs (Fig. 6C ) and laid their eggs in a bag while being raised indoors (Fig. 6D ). Currently, L. guinanensis sp. nov. is only known from SWDS. So far within this reserve, we have identified four species of Leptobrachella , namely L. guinanensis sp. nov., L. shangsiensis , L. shiwandashanensis , and L. sungi . Comparison. Table 2 presents a concise overview of the diagnostic morphological characters of species found north of the Isthmus of Kra. Leptobrachella guinanensis sp. nov. can clearly be distinguished from its phylogenetically close congeners, L. ventripunctata . Leptobrachella guinanensis sp. nov. differs from L. ventripunctata by a larger body size (SVL 30.5-32.5 mm in males; 38.7-41.8 mm in females vs 25.5-28.0 mm in males, 31.5-35.0 mm in females), ventral surface creamy white without brown spots (vs chest and belly creamy white with many scattered brown spots), ventral surfaces of limbs purplish grey (vs ventral surface of limbs grey-brown with dark brown and white speckling or dots), 1/3 toe webbing and toe lateral fringes wide (vs no toe webbing and lateral fringes), dermal ridges distinct under toes (vs absent) (Fig. 7 ), tibia-tarsal articulation reaching the centre of eye (vs the level between tympanum and posterior of eye), heels not meeting when thighs are appressed at right angles to body (vs heels overlapping). In addition, L. guinanensis sp. nov. differs from L. ventripunctata by relatively high dominant frequencies (7.3-8.3 kHz vs 6.1-6.4 kHz), call durations (mean 25.5 ms, ranging 23-31 ms vs mean 145 ms, ranging 65-430 ms) and call intervals (mean 91.2 ms, ranging 55-133 ms vs mean 134 ms, ranging 31-416 ms) (Table 4 ). Secondly, L. guinanensis sp. nov. can be easily distinguished from its sympatric species, L. shangsiensis , L. shiwandashanensis , and L. sungi . Leptobrachella guinanensis sp. nov. differs from L. shangsiensis by a larger body size (SVL 30.5-32.5 mm in males, 38.7-41.8 mm in females vs 24.9-29.4 mm in males, 30.8-35.9 mm in females), crossbars of hindlimbs with tubercles (vs lack of tubercles on crossbars of hindlimbs), 1/3 toe webbing (vs toe webbing rudimentary), head width less than length (HW/HL = 0.93 vs HW/HL = 1.15), eye diameter near equal to snout length (ED/SNT = 0.99 vs ED/SNT = 0.78), a pair of glands under the vent (vs absent glands under the vent), dominant frequencies (7.3-8.3 kHz vs 5.5-6.5 kHz), call duration (mean 25.5 ms, ranging 23-31 ms vs mean 66.0 ms, ranging 64-69 ms; Table 4 ). Leptobrachella guinanensis sp. nov. differs from L. shiwandashanensis by relatively larger body size (SVL 30.5-32.5 mm in males; 38.7-41.8 mm in females vs 26.8-29.7 mm in males, 33.7-35.9 mm in females), 1/3 toe webbing and wide lateral fringes on toe (vs no webbing and no lateral fringes on toe), tibia-tarsal articulation reaching to the centre of eye (vs posterior of eye), a pair of glands under the vent (vs absent glands under the vent), dominant frequencies (7.3-8.3 kHz vs 5.3-5.7 kHz), call duration (mean 25.5 ms vs mean 226.6 ms; Table 4 ). Leptobrachella guinanensis sp. nov. differs from L. sungi by conspicuously smaller body size (SVL 30.5-32.5 mm in males; 38.7-41.8 mm in females vs SVL 48.3-52.7 mm in males, 56.7-58.9 mm in females); iris bicoloured, upper half light copper, transitioning to silver in lower half (vs uniform gold green iris), finger II longer than finger I (vs finger I and II equal in length), tympanum distinct and rounded (vs indistinct), dorsal surface brown, an inverted triangle marking between eyes, irregular markings on shoulder and the rear of back (vs dorsum uniformly light brown or with light spots), dominant frequencies (7.3-8.3 kHz vs 2.0-2.7 kHz), call duration (mean 25.5 ms vs mean 59.4 ms), call intervals (mean 91.2 ms vs mean 478.4 ms) (Table 4 ). Figure 7. Morphological characters compared between L. guinanensis sp. nov. and L. ventripunctata A1-6 L. guinanensis sp. nov. B1-6 L. ventripunctata A1, B1 dorsal view A2, B2 dorsolateral view A3, B3 ventral view A4, B4 ventral view of hand A5, B5 ventral view of foot A6, B6 ventral view of the gravid female. Finally, L. guinanensis sp. nov. can be differentiated from other species in the genus Leptobrachella based on distinctive bioacoustics and morphological diagnostic characters (for details see Table 2 , Suppl. material 1: table S4), as well as genetic divergences (Suppl. material 1: table S3).