Leptobrachium tengchongense, Yang, Jian-Huan, Wang, Ying-Yong & Chan, Bosco Pui-Lok, 2016

Yang, Jian-Huan, Wang, Ying-Yong & Chan, Bosco Pui-Lok, 2016, A new species of the genus Leptobrachium (Anura: Megophryidae) from the Gaoligongshan Mountain Range, China, Zootaxa 4150 (2), pp. 133-148 : 136-145

Leptobrachium tengchongense sp. nov.

Holotype. SYS a004603, adult male, calling on the floor in shrub about 3 m from a rocky stream. The stream was 3–5 m wide, flowing pass mature montane evergreen broadleaf forest at Datang substation of the Tengchong Section of Gaoligongshan National Nature Reserve, Tengchong County, Yunnan Province, China (25°44'26.23'' N, 98°41'45.74'' E, 2060 m above sea level), collected at 2100 h on 13 March 2015 by J.H. Yang.

Paratypes. SYS a004623–4627, five adult males, from the same locality as holotype, collected at 2000–0130 h on 17–18 March 2016 by J.H. Yang . SYS a004628–4632, five adult males, from Cizhuhe substation of the Tengchong Section of Gaoligongshan National Nature Reserve (25°46'16.85'' N, 98°37'54.27'' E, 2240 m above sea level; 25°47'22.33'' N, 98°37'33.60'' E, 2510 m above sea level), collected at 2000–0130 h on 20–21 March 2016 by J.H. Yang.

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6) Leptobrachium sp.

!9) L. ailaonicum

) L. banae

) L. boringii

) L. buchardi

) L. chapaense

) L. guangxiense

) L. hainanense

) L. hasseltii

!21) L. huashen

) L. ingeri !

) L. leishanense 1 4.7!

) L. leucops 1 2.2 1 1.6!

) L. liui 1 5.3 2.3 1 2.2!

) L. masatakasatoi 1 4.5 4.3 1 1.0 5.2!) L. mouhoti 1 3.3 1 1.0 4.8 1 1.6 1 1.0!) L. ngoclinhense 1 3.0 1 0.8 4.6 1 1.8 1 0.4 2.3!) L. promustache 15.5 4.6 12.2 5.6 7.0 11.2 11.6!) L. pullum 14.7 11.6 5.2 12.2 11.4 2.1 3.3 11.0!) L. rakhinensis 13.3 14.1 14.1 14.7 15.1 14.3 14.1 15.9 14.5!) L. xanthops 13.3 11.0 5.0 11.2 10.6 3.3 2.9 11.2 3.9 14.3!) L. xanthospilum 13.9 12.8 7.9 13.7 12.4 6.6 5.6 12.6 6.8 15.1 6.4!) Oreolalax rhodostigmatus 13.3 11.6 11.6 12.6 11.4 11.8 11.0 11.0 11.4 14.9 11.8 13.9!) Scutiger chintingensis 14.5 12.6 12.2 13.7 12.8 12.0 11.6 13.3 12.0 13.9 11.8 14.3 9.9!

Referred specimens. SYS a004604, seven tadpoles (a004604a–g), collection data identical to holotype, not assigned as types.

Etymology. The specific epithet, “ tengchongense ”, is in reference to the type locality, Tengchong County of Yunnan Province, China.

Diagnosis. Assigned to the genus Leptobrachium on the basis of having head width larger than tibia length; skin above with a network of ridges; oval and large axillary glands present; extremities of digits rounded; breeding males lacking spines on fingers and breast; and iris bicolored ( Dubois & Ohler 1998). The new species, Leptobrachium tengchongense sp. nov., can be distinguished from other congeners by a combination of following characters: (1) relatively small size (adult males SVL 41.7–51.5 mm); (2) head width slightly larger than head length; (3) tympanum indistinct; (4) two palmar tubercles oval and distinct, inner one larger than outer one; (5) sexually active males without spines on the upper lip; (6) dorsal skin smooth with distinct network of ridges; (7) dorsum pinkish grey and scattered with irregular black markings; (8) venter dark purplish-gray with numerous small white spots on tubercles, solid white chest; (9) iris bicolored, upper one-third light blue, lower two-third dark brown.

Description of holotype. Adult male, SVL 45.3 mm. Habitus moderately stocky; body tapering to groin. Head broad and depressed; head length (HDL 18.4 mm) slightly shorter than head width (HDW 19.9 mm). Snout rounded in dorsal view, sharply sloping in profile, barely projecting beyond lower jaw in profile; nostril slightly closer to eye than to tip of snout, below canthus; internarial distance (IND 4.8 mm) shorter than interorbital distance (IOD 6.0 mm); canthus rostralis distinct; lores oblique, moderately concave; eye large, slightly projecting from side of head, diameter (EYE 6.7 mm) shorter than snout length (SNT 7.8 mm), interorbital distance slightly greater than upper eyelid width (UEW 5.0 mm); no pineal ocellus; tympanum round, nearly invisible, tympanum diameter (TYM 2.5 mm) about 37.3% eye diameter and slightly greater than distance between tympanum and eye (TED 2.0 mm); tongue heart-shaped, notched posteriorly; large, slit-like vocal sac openings on floor of mouth near lateral margin of tongue; vomerine teeth absent.

Forelimb slender. Fingers moderately slender, without webbing. Tip of fingers blunt, that of fingers I slightly swollen; relative finger lengths II = IV <I <III; two oval palmar tubercles not in contact, inner larger than outer; inner palmar tubercle larger than tip of fingers I; low callous tissue on ventral surface of fingers; nuptial pad absent. Hindlimb slender and relatively short; heels not meeting when limbs are held at right angles to body; tibiotarsal articulation of adpressed limb reaching to the posterior margin of jaws. Toes moderately slender; webbing poorly developed, formula: I 2–2 ¼ II 1 ½–3 III 2–3 ½ IV 4–2 V. Tips of toes similar to those of fingers; relative toe lengths I<II<V<III<IV; toes with lateral fringes; subarticular tubercles obscure, but elongate, replaced by low callous tissue; distinct, oval, inner metatarsal tubercle, length (IML 3.0 mm) about 76.9 % distance between tip of toe I and tubercle; no outer metatarsal tubercle.

Skin above smooth with distinct network of ridges, lower flank scattered with small tubercles; no spines on upper lip; low supratympanic ridge from posterior edge of eye to axilla; temporal region granular; tubercles on sacrum absent; sparsely granulated around vent; chin, chest and belly densely covered with white small tubercles; limbs with distinct ridges dorsally and smooth ventrally; a large, round axillary gland (2.5 mm in diameter) present on ventrolateral surface slightly posterior to insertion of forelimb with body, slightly visible ( Fig. 3 F); a flat, oblong femoral gland (4.5 mm in length) present on distal half of posteroventral surface of thigh, not in white color and imperceptible ( Fig. 3 D).

Color of the holotype in life. Dorsum pinkish grey, head slightly brighter, with distinct, black irregular markings; upper flank like dorsum, lower flank like belly, with small white spots on small tubercles; dorsal limbs with distinct wide black bandings; lower two-third of iris dark brown, upper one-third of iris light blue; black streak under canthus and supratympanic fold; a small black spot present on lower anterior corner of eye; ventral surface of body and limbs dark purplish-gray except the chest remarkably white; numerous minute white spots on tubercles on chin, chest, and belly; axillary gland and femoral gland not in white color and imperceptible ( Fig. 3 D, F).

Color of the holotype in preservative. Color in preservative dark gray-brown, dark markings on dorsum and bandings on limbs still visible. Upper part of iris and scleral arc faded to white.

Variation. All ten paratypes match the overall characters of the holotype (for measurements of the type series see Table 3). The femoral gland imperceptible in the holotype and other nine paratypes, but is dull white and slightly visible in a single paratype SYS a004627; the axillary gland is distinctly visible in five paratypes (SYS a004623, a004624, a004627, a004630 and a004632), but barely visible or imperceptible in the other five paratypes and holotype.

Advertisement call. A single, pulsed note of approximately 0.13 s duration similar to other Leptobrachium for which the advertisement call is known. A full description of the advertisement call will be presented elsewhere (Yang et al. in prep).

Tadpoles. The following description based on specimen SYS a004604a preserved in 96 % ethanol: stage 40, SVL 31.6 mm, total length 91.2 mm, tail height 18.6 mm; body elliptical in dorsal view, snout rounded; eye moderate, positioned dorsolaterally, not visible in ventral view; spiracle tube short with a free distal end, positioned laterally; tail musculature strong, relatively straight anteriorly and gradually tapering posteriorly, almost reaching the tip; the fin relatively high, tip of tail pointed; oral disk ventral, labial tooth row formula: 1:(5+5)/(4+4). Body and tail light brown, with a wide light V-shaped marking at posterior body, the base of the V slightly extending to the upper fin; dorsal and lateral body scattered with moderate dark brown spots; tail with large dark brown blotches in the proximal half and small dark spots in the distal half.

Distribution and natural history. Leptobrachium tengchongense sp. nov. is so far known from Datang substation and Cizhuhe substation of the Tengchong Section of Gaoligongshan National Nature Reserve, Tengchong County, Yunnan Province of China. The new species was found mainly on the riparian forest floor, near shallow, clear-water rocky streams in well-preserved montane evergreen broadleaf forest at elevations between 2000–2500 m. Most adult males were found calling with their bodies buried in the moist topsoil and heads hidden under leaf litter, making them difficult to detect ( Fig. 5 A). A few males were calling amongst rock crevices ( Fig. 5 B). No females were detected during the surveys.

We conducted field surveys in the type localities of the new species, namely Datang and Cizhuhe substations of GLGS, between April 2014 and March 2016. A few males were heard calling during the survey on 13–14 March 2015 in Datang, and only one calling adult male (holotype) was collected; but during the surveys on 17–18/ 20–21 March 2016 in Datang and Cizhuhe respectively, calling males of the new species were ubiquitous along streams and riparian forests, and ten calling adult males were collected from both localities. In March 2016, montane forests between 2200–2900 m a. s.l. were surveyed in Cizhuhe, but mating calls and individuals of the new species were only detected from the lower elevation band (i.e. 2200–2510 m a.s.l.). No calls and specimens of the new species were detected during the survey on 26–27 April 2014 in Datang and 18–19 May 2015 in Cizhuhe, except the tadpoles that were found in the stream in both localities in all surveys. Sympatric herpetofauna recorded in the locality during the survey included the Spiny stream toad Bufo tuberospinius (Yang, Liu & Rao) , Yunnan spiny frog Nanorana yunnanensis (Anderson) , Glandular horned toad Megophrys glandulosa Fei, Ye & Huang , Anderson's frog Odorrana andersonii (Boulenger) , Chaochiao brown frog Rana chaochiaoensis Liu , Large-eyed bamboo snake Pseudoxenodon macrops (Blyth) , and Leonard's Keelback Rhabdophis leonardi (Wall) .

Comparison. We compared Leptobrachium tengchongense sp. nov. to all other recognized species of Leptobrachium from China and Indochina, all of which formed a monophyletic clade in molecular studies ( Brown et al. 2009; Matsui et al. 2011; this study): i.e., L. ailaonicum , L. banae , L. boringii , L. buchardi , L. chapaense , L. guangxiense , L. hainanense , L. huashen , L. leishanense , L. leucops , L. liui , L. masatakasatoi , L. mouhoti , L. ngoclinhense , L. promustache , L. pullum , L. xanthops and L. xanthospilum . We also compared L. tengchongense sp. nov. with the newly described L. bompu from India of which comparative 16S mtDNA data was unavailable from the GenBank. Unrelated Sunderland/ Thailand clade taxa (occur in southern Myanmar, Thailand, Peninsular Malaysia, Sumatra, Java, Bali, Borneo, and the Philippines) are omitted from comparisons for simplicity.

By the absence of spines on the upper lip in sexually active males, Leptobrachium tengchongense sp. nov. distinctly differs from six other congeners, i.e., L. ailaonicum , L. boringii , L. leishanense , L. liui , L. ngoclinhense and L. promustache , all of which process spines on the upper lip in sexually active males and were formerly referred to the genus Vibrissaphora ( Rao et al. 2006; Fei et al. 2005, 2009). By having a bicolored iris with upper one-third light blue, Leptobrachium tengchongense sp. nov. can be distinguished from L. bompu (iris uniformly grey-blue), L. chapaense (iris uniformly dark brown), L. huashen (iris uniformly dark brown, [see discussion below]), L. leishanense (upper part of iris light green), L. leucops (upper part of iris white), L. liui (upper part of iris light green), L. mouhoti (iris black with an orange-yellow crescent on outer margin), L. ngoclinhense (iris uniformly dark brown), L. pullum (upper part of iris red), L. xanthops (upper part of iris pale yellow), and L. xanthospilum (upper half of iris white). By having a relatively smaller body size (SVL 41.7–51.5 mm in adult males), Leptobrachium tengchongense sp. nov. can be distinguished from the larger L. ailaonicum (68.5–87.6 mm in males), L. banae (57.2–70.0 mm in males), L. boringii (69.8–89.0 mm in males), L. guangxiense (53.5–58.1 mm in males), L. leishanense (69.0– 95.7 mm in males), L. liui (67.8–90.5 mm in males), L. masatakasatoi (55.2 mm in male), L. ngoclinhense (77.1 mm in male) and L. xanthospilum (62.8–73.4 mm in males). By having inner palmar tubercle larger than outer one, Leptobrachium tengchongense sp. nov. can be distinguished from L. buchardi , L. ngoclinhense , L. pullum and L. hainanense (versus two palmar tubercles subequal in size in former three species, and outer one slightly larger or equal to the inner one in L. hainanense ). Leptobrachium tengchongense sp. nov. further differs from L. guangxiense and L. hainanense by having an indistinct tympanum (versus distinct in both species).

Besides those differences mentioned above, Leptobrachium tengchongense sp. nov. further differs from the phylogenetically closely related congeners without spines on the upper lip in adult males, namely L. chapaense , L. huashen and L. masatakasatoi ( Fig. 2 ), by having a pinkish grey dorsal surface of body in life (versus brown in life in L. chapaense and L. huashen , Fig. 4 .), presence of a distinct network of ridges on the dorsum (versus absent in L. masatakasatoi ); inner metatarsal tubercle larger than first toe (versus smaller in L. masatakasatoi ); presence of a dark black spot on lower anterior corner of eye (versus absent in L. chapaense and L. huashen ), relatively wide black bands on the dorsal surface of the limbs (versus relatively narrower bands in these species); and large and distinct cream spots on lower flanks absent (versus present in these species).

TABLE ³. Measurements (mm) of type specimens of Leptobrachium tengchongense sp. nov.. Abbreviations đefineđ in text.

SYS SYS SYS SYS SYS SYS SYS SYS SYS SYS SYS Mean ± SD

a004603 a004623 a004624 a004625 a004626 a004627 a004628 a004629 a004630 a004631 a004632 (N=11)

Holotype Paratype Paratype Paratype Paratype Paratype Paratype Paratype Paratype Paratype Paratype

Male Male Male Male Male Male Male Male Male Male Male

45.3 51.5 48.0 45.9 47.9 50.5 48.2 48.9 41.7 48.6 50.3 47.89±2.76 HDL 18.4 20.7 19.4 18.7 19.4 20.2 19.7 20.8 17.5 18.9 20.2 19.45±1.02 HDW 19.9 21.7 20.4 20.0 20.2 21.2 20.9 22.3 18.2 20.5 21.9 20.65±1.14

7.8 8.4 8.1 7.9 7.7 8.1 8.1 7.9 7.0 8.2 8.5 7.97±0.40 EYE 6.7 7.4 6.8 6.2 6.5 6.9 6.7 6.9 5.4 6.5 6.4 6.58±0.50 UEW 5.0 5.6 5.1 5.0 5.0 5.3 5.2 5.8 4.8 4.8 4.6 5.11±0.35

6.0 6.0 6.7 6.1 6.3 6.9 6.6 6.3 6.5 6.6 6.6 6.42±0.30

4.8 5.1 5.1 4.7 4.6 4.9 5.1 5.2 4.7 5.2 5.1 4.95±0.22 LAL 14.1 14.4 14.8 13.7 13.6 14.4 13.7 14.1 12.6 14.3 14.0 13.97±0.58 HND 11.9 13.5 12.9 12.6 12.4 13.7 12.0 13.3 11.5 13.2 13.2 12.75±0.72

16.9 19.0 17.5 17.2 17.6 18.8 17.6 18.4 15.9 17.9 18.0 17.71±0.88 TGH 20.6 22.2 20.9 20.1 20.6 22.0 20.4 21.6 16.9 20.3 21.6 20.65±1.44

9.8 9.8 9.2 9.7 9.4 9.5 9.9 9.8 9.3 10.0 10.7 9.74±0.41

18.3 20.4 19.6 18.4 18.3 20.9 18.5 20.0 16.6 20.0 20.3 19.21±1.28

3.0 3.4 3.4 2.7 2.9 3.7 3.0 3.3 2.6 3.6 3.6 3.20±0.38 IMW 1.5 1.8 1.5 1.3 1.5 1.6 1.5 1.8 1.3 1.6 1.7 1.55±0.17

Figures

FIGURE 2. Bayesian inference (BI) tree derived from partial sequences of the mitochondrial 16 S rRNA gene. Numbers above branches are bootstrap support values for maximum likelihood analyses (> 70 retained) and numbers below branches indicate Bayesian posterior probabilities (> 70 % retained).

FIGURE 3. Holotype of Leptobrachium tengchongense sp. nov. (SYS a 004603) in preservative: (A) dorsal view; (B) ventral view; (C) plantar view of the left foot; (E) posteroventral surface of thigh, note the imperceptible femoral gland; (D) volar view of the right hand; (F) ventral surface of right chest, note the indistinct axillary gland. Photos by J. H. Yang.

FIGURE 4. (A) Holotype of Leptobrachium tengchongense sp. nov. (SYS a 004603) in life, inserted picture depicts the iris coloration of holotype in life; (B) Leptobrachium huashen from Linjiapu substation of Gaoligongshan NNR, inserted picture depicts the iris coloration in life. Photos by J. H. Yang.

FIGURE 5. (A) Paratype of Leptobrachium tengchongense sp. nov. (SYS a 004625) in life from Datang Substation of GLGS, inserted picture depicts the microhabitat; (B) paratype of Leptobrachium tengchongense sp. nov. (SYS a 004628) in life from Cizhuhe Substation of GLGS, inserted picture depicts the microhabitat. Photos by J. H. Yang.