Zootaxa 2796: 15–28 (2011) ISSN 1175-5326 (print edition) www.mapress.com/zootaxa/ Article ZOOTAXA Copyright © 2011 · Magnolia Press ISSN1175-5334(online edition) A new species of Leptolalax (Anura: Megophryidae) from southern Vietnam JODI J. L. ROWLEY1,4, DUONG THI THUY LE2, DAO THI ANH TRAN2,3, HUY DUC HOANG2 1Australian Museum, 6 College St, Sydney, NSW, 2010, Australia. 2Faculty of Biology, University of Science-Ho Chi Minh City, 227 Nguyen Van Cu, District 5, Ho Chi Minh City, Vietnam 3Zoologisches Forschungsmuseum Alexander Koenig, Adenauerallee 160, D-53113 Bonn, Germany 4Corresponding author. E-mail: [email protected] Abstract We describe a new species of small megophryid frog from the Langbian Plateau in southern Vietnam, the southernmost record of Leptolalax from Vietnam. Leptolalax bidoupensis sp. nov.is distinguished from its congeners by a combination of a dark brownish red ventral surface with white speckling on entire ventral surface including throat, arms and legs, small size (23.6–24.6 mm in four adult males and 29.2–29.4 mm in two adult females), bicoloured iris (coppery red upper half, fading to pale silver ventrally), a mostly smooth skin texture with no skin ridges, and relatively short tibia (male TIB:SVL 0.44–0.46).The male advertisement call of the new species, consisting of 6–9 single-pulsed notes with a dominant fre- quency of 1.9–3.8 kHz, is also unique among Leptolalax species for which calls are known. From the morphologically similar L. applebyi and L.melicus,L. bidoupensis sp. nov. differs by 9.3% and 9.6% sequence divergence at the 16S mtD- NA gene. At present, the new species is known from montane evergreen forest between 1620–1730 m elevation, within an area of 1 km2. We suggest the species should be considered Data Deficient following IUCN’s Red List categories. Key words:Acoustics,Anura,Da Lat Plateau, Leptolalax bidoupensis sp. nov., Lam Dong Province, Langbian Plateau, Southeast Asia, Vietnam Introduction The genus Leptolalax (Dubois 1983) currently contains 29 species of small frogs of the forest floor, distributed in rocky streams in hilly evergreen forest throughout Southeast Asia, southern China and northeastern India (Frost 2010, Rowley et al. 2010a, 2010b, 2010c). There has been a rapid increase in the number of known Leptolalax spe- cies in recent years, resulting from a combination of increased field surveys in the region and the incorporation of acoustic and molecular data in delineating species boundaries (eg. Das et al. 2010; Rowley et al. 2010a, 2010b, 2010c; Sengupta et al. 2010). To date, eleven species of Leptolalax have been reported from Indochina; L. aereus,L. applebyi, L. bourreti, L. croceus, L. melicus, L. nahangensis, L. oshanensis, L. pelodytoides, L. pluvialis, L. sungi and L. tuberosus (Frost 2010; Rowley et al. 2010a, 2010b, 2010c). All but L. aereus and L. melicus have been reported from Vietnam (Nguyen et al. 2009; Rowley et al. 2010a, 2010c). Within Vietnam, Leptolalax are known throughout suitable hab- itat in northern and central Vietnam, with the southernmost record of Leptolalax to date from the Kon Tum Plateau in central Vietnam (Gia Lai Province; Nguyen et al. 2009). Here we describe a new Leptolalax species collected during recent herpetological surveys on the Langbian Pla- teau in southern Vietnam. Although important amphibian collections were made on the Langbian Plateau by Smith (1921, 1924), Leptolalax were not reported from the Plateau, and the new species represents the southernmost record of Leptolalax from Vietnam. The new species is most morphologically similar to L. applebyi and L. melicus, both recently discovered from the Kon Tum Plateau to the north, but can be distinguished from these and all other Leptolalax species on the basis of morphological, acoustic and molecular differences. Accepted by M. Vences: 9 Feb. 2011; published: 21 Mar. 2011 15 Material and methods We recorded morphological data from specimens fixed in 10% formalin and then stored in 70% ethanol. Specimens were deposited at the Australian Museum (AMS) and the North Carolina Museum of Natural Sciences (NCSM). Some specimens currently at the AMS will be deposited at the University of Science, Ho Chi Minh City (UNS) and have been cross-cataloged at both institutions. In these instances, voucher numbers are reported as UNS/AMS. Morphometric data were taken (to the nearest 0.1 mm) with digital callipers. Measurements include snout-vent length (SVL); head length from tip of snout to rear of jaws (HDL); head width at the commissure of the jaws (HDW); snout length from tip of snout to the anterior corner of eye (SNT); diameter of the exposed portion of the eyeball (EYE); interorbital distance (IOD); horizontal diameter of tympanum (TMP); distance from anterior edge of tympanum to posterior corner of the eye (TEY); tibia length with the hindlimb flexed (TIB), manus length from tip of third digit to base of inner palmar tubercle (ML), pes length from tip of fourth toe to base of the inner meta- tarsal tubercle (PL), length of adpressed first finger from tip to distal edge of the inner palmar tubercle (F1L), length of adpressed second finger from tip to distal edge of inner palmar tubercle (F2L), and length of adpressed third finger from tip to distal edge of inner palmar tubercle (F3L). Sex was determined by direct observation of calling in life, the presence of internal vocal sac openings and/or gonadal inspection. Mass was recorded in life (to the nearest 0.1 g), using Pesola scales. We obtained comparative morphological data from museum specimens of Leptolalax and photographs of these specimens in life (Appendix) and from the literature: L. aereus (Rowley et al. 2010c), L. alpinis (Fei et al. 1991, 2009, 2010), L. arayai (Matsui 1997), L. bourreti (Dubois 1983), L. croceus (Rowley et al. 2010a), L. dringi (Dubois 1986; Inger & Stuebing 2005), L. fuliginosus (Matsui 2006), L. gracilis (Günther 1872; Malkmus et al. 2002; Inger & Stuebing 2005), L. hamidi (Matsui 1997), L. heteropus (Boulenger 1900), L. kecil (Matsui et al. 2009), L. khasiorum (Das et al. 2010), L. kajangensis (Grismer et al. 2004), L. latera- lis (Anderson 1871; Humtsoe et al. 2008), L. liui (Fei et al. 1991, 2009, 2010), L. maurus (Inger et al. 1997), L. melanoleucus (Matsui 2006), L. melicus (Rowley et al. 2010b), L. nahangensis (Lathrop et al. 1998), L. oshanensis (Liu 1950; Fei et al. 2009, 2010), L. pelodytoides (Boulenger 1893, 1908), L. pictus (Malkmus 1992; Malkmus et al. 2002), L. pluvialis (Ohler et al. 2000), L. solus (Matsui 2006), L. sungi (Lathrop et al. 1998), L. tamdil (Sen- gupta et al. 2010), L. tuberosus (Inger et al.1999; Rowley et al. 2010a), L. ventripunctatus (Fei et al. 1991, 2009, 2010). Due to the undiagnosed diversity within genus, where available we relied exclusively on descriptions of specimens reported in the original species descriptions. We also examined colour photographs of the holotype of L. pluvialis (MNHN 1999.5675) in preservative. Advertisement calls were recorded with an Edirol R-09 WAVE/MP3 Recorder (44.1 kHz sampling rate and 24- bit encoding) with a Røde NTG-2 condenser shotgun microphone. Calls were recorded at a distance of approxi- mately 0.1–0.3 m and ambient temperatures were taken immediately after recordings using a Kestrel 3500 hand- held weather meter. Calls were analysed with Raven Pro 1.3© software (http://www.birds.cornell.edu/raven). Audiospectrograms in figures were calculated with fast-Fourier transform (FFT) of 256 points, 50% overlap and 172 Hz grid-spacing, using Hanning windows. Comparisons of advertisement calls among species within the genus Leptolalax are hindered by the limited nature of many published call descriptions, with call measurements and visual representations of calls insufficient for meaningful comparisons between species. Comparisons of anuran advertisement calls in general are also complicated by inconsistent use of terms and a lack of clear definitions in terminology, particularly with respect to the units of a call, a note or a pulse (Gerhardt & Huber 2002). Here we use the definitions of Duellman (1970), except that we define a single call as vocalisations produced during a single expiration (Brown & Richards 2008). Temporal and spectral parameters of calls were measured using the defini- tions of Cocroft & Ryan (1995), except for fundamental frequency, where the definition of Duellman (1970) was used. For each call recording, we measured the call duration (ms), intercall interval (ms), number of notes per call, note duration (ms), internote interval (ms), number of pulses per note, note repetition rate (notes/s) and dominant frequency (kHz). Comparative advertisement call characters for Leptolalax species were taken from references, with advertisement calls known for 18 of the 29 known species of Leptolalax (Jiang et al. 2002; Malkmus et al. 2002; Matsui 1997, 2006; Matsui et al. 2009; Xu et al. 2005; Rowley & Cao 2009; Rowley et al. 2010a, 2010b, 2010c; Sukumaran et al. 2010). To maintain consistency and facilitate meaningful comparisons, we have used the terminology defined above to compare calls, regardless of terms used in these references. We analyzed approximately 550 base pairs (bp) of mitochondrial 16S ribosomal RNA for four individuals of the new species and compared them with two Leptolalax applebyi (GenBank accession numbers HM133597– 16 · Zootaxa 2796 © 2011 Magnolia Press ROWLEY ET AL. HM133598) and three Leptolalax melicus (GenBank accession numbers HM133599–HM133601), the most mor- phologically similar and least geographically distant species. DNA was extracted using DNeasy tissue extraction kits (Qiagen). We used the primers 16SAR and 16SBR of Palumbi et al. (1991) to amplify the 16S rRNA gene. Standard PCR protocols were used and PCR products were purified using ExoSap-IT (USB Corporation, OH, USA). Purified templates were sequenced directly by Macrogen (Seoul, Korea). Sequences were validated using Sequencher 4.10 (Gene Codes, Ann Arbor, MI), aligned using the Clustal option in MEGA 4 and refined by eye. Kimuara Uncorrected pairwise sequence divergence was calculated using MEGA 4. DNA sequences for the new species were deposited in GenBank under the accession numbers HQ902880– HQ902883. Leptolalax bidoupensis sp. nov. Holotype: AMS R 173133, adult male, calling on clay bank 0.2 m from 1–4 m wide, medium-high gradient, rocky stream in montane evergreen forest in Bidoup-Nui Ba National Park, Lam Dong Province, Vietnam (12.19225º N, 108.71494º E, 1730 m, Figure 1). Collected at 23:55 h on 19 May 2008 by J. J. L. Rowley, Hoang D. H., Le T. T. D., and Tran T. A. D. Paratypes: UNS 00101/AMS R 173135, adult male, calling on tree root 0.2 m above 2–5 m wide, medium- high gradient, rocky stream in montane evergreen forest in Bidoup-Nui Ba National Park, Lam Dong Province, Vietnam (12.19106º N, 108.71703º E, 1641 m), collected at 20:25 h on 20 May 2008. UNS 00102/AMS R 173137, metamorph, in water of swampy area adjacent to a swift, rocky stream in cloud forest in Bidoup-Nui Ba National Park, Lam Dong Province, Vietnam (12.18644º N, 108.71486º E, 1627 m), collected at 19:45 h on 18 May 2008. AMS R 173134, adult female, on clay bank 0.2 m from 1–4 m wide, medium-high gradient, rocky stream in mon- tane evergreen forest in Bidoup-Nui Ba National Park, Lam Dong Province, Vietnam (12.19225º N, 108.71494º E, 1730 m), collected at 23:50 h on 19 May 2008, in close proximity to holotype. AMS R 173136, adult male, calling on leaf litter 0.2 m from 2–5 m wide, medium-high gradient, rocky stream in montane evergreen forest in Bidoup- Nui Ba National Park, Lam Dong Province, Vietnam (12.19106º N, 108.71703º E, 1641 m), collected at 21:40 h on 20 May 2008. NCSM 77320, adult female, in water of swampy area off swift, rocky stream in montane evergreen forest in Bidoup-Nui Ba National Park, Lam Dong Province, Vietnam (12.18644º N, 108.71486º E, 1627 m), col- lected at 19:50 h on 18 May 2008. NCSM 77321, adult male, sitting in upright posture (previously calling?) in leaf litter, 0.1 m from swift, rocky stream in montane evergreen forest in Bidoup-Nui Ba National Park, Lam Dong Province, Vietnam (12.18644º N, 108.71486º E, 1627 m), collected at 21:45 h on 18 May 2008. NCSM 77322, metamorph, on clay bank 0.5 m from 2–5 m wide, medium-high gradient, rocky stream in montane evergreen for- est in Bidoup-Nui Ba National Park, Lam Dong Province, Vietnam (12.19106º N, 108.71703º E, 1641 m), col- lected at 21:30 h on 20 May 2008. All specimens were collected by J. J. L. Rowley, Hoang D. H., Le T. T. D., and Tran T. A. D. Etymology. specific epithet is in reference to the type locality of Bidoup-Nui Ba National Park. Diagnosis. Assigned to the genus Leptolalax on the basis of the following: small size, rounded finger tips, the presence of an elevated inner palmar tubercle not continuous to the thumb, presence of macroglands on body (including supra-axillary, pectoral, femoral and ventrolateral glands), vomerine teeth absent, tubercles on eyelids, anterior tip of snout with vertical white bar (Dubois 1983; Lathrop et al. 1998; Delorme et al. 2006). Leptolalax bidoupensis is distinguished from its congeners by a combination of (1) a dark brownish red ventral surface with white speckling on entire ventral surface including throat, arms and legs, often forming distinct marbling on chest and belly, (2) small size (23.6–24.6 mm in four adult males and 29.2–29.4 mm in two adult females), (3) bicoloured iris (coppery red upper half, fading to pale silver ventrally), (4) a mostly smooth skin texture with no skin ridges, and (5) relatively short tibia (male TIB:SVL 0.44–0.46).The male advertisement call of the new species, consisting of 6–9 single-pulsed notes with a dominant frequency of 1.9–3.8 kHz, is also unique among Leptolalax species for which calls are known. Description of holotype. Head slightly longer than wide; snout bluntly rounded in dorsal view and in profile, projecting slightly beyond margin of the lower jaw; nostril closer to tip of snout than eye; canthus rostralis distinct, gently rounded; lores sloping; vertical pupil; eye diameter smaller than snout length; tympanum distinct, round, diameter smaller than that of the eye; tympanic rim elevated relative to skin of temporal region; vomerine teeth absent; pineal ocellus absent; vocal sac openings oval, located posteriolaterally on floor of mouth; tongue long, A NEW MEGOPHRYID FROG SPECIES FROM SOUTHERN VIETNAM Zootaxa 2796 © 2011 Magnolia Press · 17 moderate width, with slight notch at posterior tip; raised supratympanic ridge running from eye towards axillary gland. Tips of fingers rounded, not swollen; relative finger lengths I < II = IV < III; nuptial pad absent; subarticular tubercles absent; a large, round inner palmar tubercle distinctly separated from small, laterally compressed outer palmar tubercle; no finger webbing or lateral fringes. Tips of toes like fingers; relative toe length I < II < V < III < IV; subarticular tubercles absent, replaced by dermal ridges, distinct on second, third, fourth and fifth toes; large, oval inner metatarsal tubercle pronounced, outer metatarsal tubercle absent; basal webbing; weak lateral fringes. Tibia relatively short and stout, width approximately one third of length, tibia 46% of snout-vent length; tibiotarsal articulation reaches eye. Skin on dorsum mostly smooth, with fine, scattered tubercles concentrated on eyelids, ventrolateral surfaces and on upper surfaces on tibiotarsus; ventral skin smooth; pectoral gland oval, 1.2 mm diam- eter; femoral gland oval, 0.8 mm diameter, on posteroventral surface of thigh, closer to knee than to vent; supra- axillary gland raised, 1.2 mm diameter. Ventrolateral glands present, dorsolaterally compressed, forming an incom- plete line that is difficult to distinguish from white marbling. TABLE 1. Measurements (mm) of Leptolalax bidoupensissp. nov. Abbreviations defined in text. Tympana were indistinct in metamorphs. Males Females Metamorphs AMS R UNS AMS R NCSM AMS R NCSM UNS NCSM 173133* 00101/ 173136 77321 173134 77320 00102/ 77322 AMS R AMS R 173135 173137 SVL 23.6 24.3 24.6 24.6 29.4 29.2 17.3 17.1 HDL 9.2 9.2 9.6 9.6 10.1 10.5 5.4 5.7 HDW 8.9 8.8 9.2 9.0 9.6 9.8 5.5 5.5 SNT 3.6 3.5 3.6 3.6 3.8 3.8 2.3 2.3 EYE 2.5 2.6 2.8 2.8 2.8 2.7 1.6 1.7 IOD 2.9 3 2.9 2.9 3.1 3.1 2.1 2.4 TMP 1.4 1.5 1.9 1.5 1.8 1.6 – – TEY 1.3 0.4 0.9 1.0 0.6 1.5 – – TIB 10.4 10.7 11.2 11.3 12.2 11.3 7.6 7.9 ML 5.2 5.5 5.5 5.2 6.4 5.5 3.8 4.1 PL 9.9 10.3 10.3 9.8 11.9 10.9 6.8 7.2 F1L 2.0 2.0 2.3 2 2.3 2.3 1.1 1.4 F2L 2.3 2.5 2.5 2.3 2.5 2.5 1.3 1.6 F3L 3.8 4.1 4.3 4.3 5.0 4.5 2.7 2.9 TIB:SVL 0.44 0.44 0.45 0.46 0.41 0.39 0.44 0.46 HDL:HDW 1.04 1.05 1.04 1.07 1.06 1.07 1.00 1.02 HDL:SVL 0.36 0.37 0.39 0.39 0.34 0.36 0.31 0.33 Weight (g) 1.1 1.2 1.4 1.2 2.2 1.4 0.5 0.6 *holotype. Colour of holotype in life. Dorsal surface brown with distinct darker brown markings; V-shaped interorbital marking, W-shaped marking between axillae and inverted V-shaped, irregular marking above sacrum; fine, pale greyish-blue tubercles on upper eyelids, snout, lateral margins of dorsum and upper surfaces of legs; blackish brown patch on upper lip under eye; blackish brown line along canthus rostralis through eye, and continuing below supratympanic ridge, terminating above axilla, encompassing nare and most of tympanum; transverse dark brown bars on dorsal surface of limbs; large, black blotch on posterior flank joining with inverted V-shaped marking above sacrum and numerous, smaller black spots on sides from groin to axilla; elbow and upper arms without dark bars but with distinct copper colouration; fingers and toes with transverse barring. Dark brownish red ventral sur- face with white speckling on entire ventral surface including throat, arms and legs, forming distinct marbling on 18 · Zootaxa 2796 © 2011 Magnolia Press ROWLEY ET AL. chest and belly; ventral margin of throat bearing broken row of slightly larger white spots. Supra-axillary gland copper; femoral glands white; pectoral glands white, lined with dark brown. Iris bicoloured, with upper half cop- pery red, fading to pale silver in lower half; minute, black reticulations throughout. FIGURE 1. Collection site of Leptolalax bidoupensis sp. nov. from Bidoup-Nui Ba National Park (black star) and type locali- ties of Leptolalax applebyi (white circle) and Leptolalax melicus (black circles). Colour of holotype in preservative. Dorsum dark brown with slightly paler limbs. Ventral surface brown, with white speckling and marbling. Macroglands white. Patterns on dorsal surface less distinct (Figure 3). Measurements. Holotype: SVL 23.6, HDL 9.2, HDW 8.9, SNT 3.6, EYE 2.5, IOD 2.9, TMP 1.4, TEY 1.3, TIB 10.4, ML 5.2, PL 9.9, F1L 2.0, F2L 2.3, F3L 3.8, weight 1.1g Variation. Specimens vary slightly in colour in life. AMS R 173135 and NCSM 77320 have a paler brown dorsum compared to the holotype, and dorsal patterns lined with diffuse, paler brown. AMS R 173136 and NCSM 77321 also have paler brown dorsal surfaces, but with only indistinct, diffuse dorsal markings; in NCSM 77321 the only distinct dark markings are over the tympanum, below the supratympanic ridge and as a single patch on the flank; AMS R 173136 has larger and more numerous brownish black patches on the flanks. AMS R 173134 and UNS 00101/AMS R 173135 have more mottled dorsal patterns. The ventral surfaces of specimens in the type series vary in their degree of white speckling and in the presence or absence of white marbling on parts of the chest and belly; UNS 00101/AMS R 173135 and NCSM 77320–77321 have less intense white speckling on ventral surfaces and only slight white marbling on parts of their chest; AMS R 173133 has slightly less intense white speckling and no white marbling. Pectoral glands are more distinct in specimens with less marbling. Ventrolateral glands are most A NEW MEGOPHRYID FROG SPECIES FROM SOUTHERN VIETNAM Zootaxa 2796 © 2011 Magnolia Press · 19 easily detected in the holotype AMS R 173133, and more widely spaced and difficult to distinguish from white ventral patterning on all other specimens in the type series. UNS 00102/AMS R 173137 and NCSM 77322 are metamorphs with incompletely reabsorbed tails (tail length 2 mm in UNS 00101/AMS R 173137 and 8 mm in NCSM 77322), almost uniformly dark brown dorsal surfaces, dark brown ventral surfaces with white speckling, and indistinct tympana. Iris colouration varies slightly in the balance of coppery red versus pale silver (Figure 5A– C). FIGURE 2. (A) Lateral view of male holotype (AMS R 173133) of L. bidoupensis sp. nov.in life, and (B) ventral views of gravid female paratype (AMS R 173134) and male holotype (AMS R 173133) of L. bidoupensis sp. nov. in life. 20 · Zootaxa 2796 © 2011 Magnolia Press ROWLEY ET AL. FIGURE 3. (A) Dorsal, (B) ventral and (C) lateral view of preserved holotype (AMS R 173133) of L. bidoupensis sp. nov. Scale bars = 5 mm. Advertisement call. Call descriptions are based on the calls of the holotype, taken at 19.0 ºC ambient temper- ature. Calls were an average of 479 ms in duration and consisted of 6–9 (usually 7) singly pulsed notes, or clicks, of variable (1–13 ms) duration, repeated at a rate of 18–21 notes per second (Table 2, Figures 4A, C). Calls were highly amplitude modulated, with amplitude peaking towards the middle of the call. The dominant frequency was 2.3–2.6 kHz, and harmonics were weak or absent. Slight frequency modulation was present within each call, with lower amplitude notes, particularly the last note, often slightly higher or lower frequency. Calls were repeated at a rate of approximately 0.2 calls per second, and had an average intercall interval of 4 s. Call repetition rate, intercall interval, and the number of notes per call varied slightly within calling bouts and among individuals (Table 2, Fig- ure 4). To the human ear, the advertisement call of L. bidoupensis is a slow rasping, similar to an orthopteran, with each note distinctly discernable. The call is extremely faint and difficult to locate, even when compared to other species within the genus. A NEW MEGOPHRYID FROG SPECIES FROM SOUTHERN VIETNAM Zootaxa 2796 © 2011 Magnolia Press · 21 TABLE 2. Measurements of advertisement call parameters for Leptolalax bidoupensis sp. nov. Parameter values are given as means (and ranges). AMS R 173133* UNS 00101/ AMS R 173136 Non-vouchered 1 Non-vouchered 2 AMS R 173135 Number of calls 10 10 10 8 10 Number of notes 85 53 67 50 72 (cid:0) (cid:0) (cid:0) (cid:0) (cid:0) Call duration (ms) 479 (326 543) 320 (278 343) 334 (234 391) 394 (380 411) 401 (277 463) Call repetition rate (calls/s) 0.22 0.17 0.13 0.14 0.1 (cid:0) (cid:0) (cid:0) Intercall interval (ms) 4014 (3590 5537 (4393 7071 (4984 6689 9511 (cid:0) (cid:0) 4521) 7430) 13106) (5153 8492) (7560 12284) (cid:0) (cid:0) (cid:0) (cid:0) Notes/call 8.2 (6 9) 6.7 (6 7) 6.7 (5 8) 7 (7) 7.2 (5 8) (cid:0) (cid:0) (cid:0) (cid:0) (cid:0) Note duration (ms) 8.4 (1 13) 6.3 (2 9) 8.3 (4 11) 7.9 (4 12) 7.4 (4 11) (cid:0) (cid:0) (cid:0) (cid:0) (cid:0) Internote interval (ms) 55 (18 70) 49 (42 59) 49 (9 76) 56 (46 76) 55 (22 66) (cid:0) (cid:0) Note repetition rate (notes/s) 18 21 21 14 21 18 18 (cid:0) (cid:0) (cid:0) (cid:0) (cid:0) Dominant frequency (kHz) 2.1(2.3 2.6) 2.1 (2.1 3.6) 2.5 (1.9 3.4) 2.0 (1.9 2.3) 2.5 (2.3 3.8) Temperature (°C) 19.0 20.4 21.0 20.2 20.4 *holotype FIGURE 4. Advertisement call of Leptolalax bidoupensis sp. nov. (holotype AMS R 173133) recorded at ambient air temper- ature of 19.0º C. (A) 48 s waveform of relative amplitude over time for 11 calls, (B) waveform and (C) corresponding spectro- gram of single representative call containing eight notes, expanded from section shown in A, and (D) power spectrum (relative amplitude vs. frequency) of fourth note shown in B–C. 22 · Zootaxa 2796 © 2011 Magnolia Press ROWLEY ET AL. Sequence divergence. Uncorrected sequence divergences between L. bidoupensis (NCSM 77320–77321, AMS R 173133–173134) and L. applebyi (holotype AMS R 171703, paratype AMS R 171704) collected from approximately 350 km away were 9.3% at the 16S rRNA gene. Uncorrected sequence divergences between L. bid- oupensis and L. melicus (MVZ 258197–258199) collected from approximately 300 km away were 9.6% at the 16S rRNA gene. This degree of pairwise divergence in the 16S rRNA gene in frogs usually represents differentiation at the species level (Vences et al. 2005), and is greater than that between L. applebyi and L. melicus (6.6%). There was no intraspecific variation in this gene fragment for L. bidoupensis, L. applebyi, and L. melicus. Ecology. All specimens of the new species were found in montane evergreen forest between 1620–1730 m ele- vation. Males were observed calling on stream banks, less than 0.5 m from small (< 5 m wide) rocky streams in May and July. The new species was not heard or observed during surveys in the month of March, when conditions were cooler and drier. Conservation status. Leptolalax bidoupensis is known only from an area of approximately 1 km2 in Bidoup Nui Ba National Park. The actual distribution of the new species is unknown but probably extends to adjacent for- ested areas in the Langbian Plateau, including Chu Yang Sin National Park in Dak Lak Province, and Phuoc Binh National Park in Ninh Thuan Province, both of which contain montane evergreen forest at similar elevations and continuous with Bidoup-Nui Ba National Park. Given the available information, we suggest the species should be considered Data Deficient following IUCN’s Red List categories (IUCN 2001). Comparisons. Leptolalax bidoupensis is distinguished from all other species in the genus except for L. applebyi by having a dark brownish red ventral surface with white speckling on entire ventral surface including throat, arms and legs, forming distinct marbling on chest and belly (L. aereus,L. alpinis, L. arayai,L. bourreti,L. dringi, L. fuliginosus, L. gracilis, L. hamidi, L. khasiorum, L. lateralis, L. liui, L. nahangensis, L. oshanensis, L. pelodytoides,L. pictus, L. solus, L. sungi, L. tamdil and L. tuberosus have mostly white or pale grey/brown venters, with or without dark spots or mottling; L. croceus has a bright orange belly; L. melicus has an off-white to pale pink ventral surface with diffuse dark brown blotches and distinct white speckling on the chest, belly and throat; L. plu- vialis has a dirty white/grey venter with dark brown/grey marbling, and uniform pale dirty white/grey throat with pale speckling only around the margins; L. melanoleucus and L. ventripunctatus display large patches of distinct black and white marbling, L. heteropus has a grey venter, speckled with black; L. maurus has a black or dark grey brown venter, with indistinct small light areas, and L. kecil has a uniformly dark venter with large, dark orange pec- toral glands). The small size of Leptolalax bidoupensis (23.6–24.6 mm in four adult males and 29.2–29.4 mm in two adult females) further distinguishes it from the larger L. arayai (male 29.6 mm), L. bourreti (male 36.2 mm, females 42– 45 mm). L. dringi (males 28.7–30.3 mm, female 37.5 mm), L. fuliginosus (males 28.2–30.0 mm), L. gracilis (males 30–36 mm), L. hamidi (males 28.7–31.3 mm, females 36.1–42.8 mm), L. heteropus (male 33 mm), L. kajangensis (males 34–35 mm), L. nahangensis (male 40.8 mm), L. pelodytoides (male ~30 mm, female ~37 mm), L. pictus (males 31–34 mm, female 47 mm), L. sungi (males 48.3–52.7 mm, females 56.7–58.9), and L. tamdil (male 32.3 mm, female 31.8 mm). The remaining, small-sized congeners may have overlapping body-sizes for either or both sexes (L. aereus males 25.1–28.9 mm, females 27.1–38.6 mm; L. alpinis males 24.0–26.4 mm, females 31.7–32.5 mm; L. applebyi males 19.6–20.8 mm, females 21.7 mm; L. croceus males 22.2–27.3 mm; L. kecil males 19.3–20.5 mm, female 20.5 mm; L. khasiorum males 24.5–27.3 mm, females 31.2–33.5 mm; L. lateralis males 26.9–28.3 mm, female 36 mm; L. liui males 23.0–28.7 mm, females 23.1–28.1 mm; L. maurus male 26.1 mm, female 31.8 mm; L. melanoleucus males 26.6–28.8 mm, female 32.7 mm; L. melicus males 19.5–22.7 mm; L. oshanensis males 26.6–30.7 mm, female 31.6 mm; L. pluvialis males 21.3–22.3 mm; L. solus male 27.6 mm; L. tuberosus males 24.4–29.5 mm, female 30.2 mm; and L. ventripunctatus males 25.5–28.0 mm). In having a bicoloured iris, with the upper half coppery red and the lower half fading to pale silver, L. bid- oupensis differs from at least L. aereus, L. applebyi, L. croceus, L. kajangensis, L. kecil, L. maurus, L. melicus, L. nahangensis, L. sungi and L. tuberosus, all of which have uniform iris colouration under black reticulations (Table 3; Figure 5). Leptolalax bidoupensis is also differentiated from L. arayai, L. croceus, L. khasiorum, L. lateralis, L. maurus, L. solus, L. tamdil, L. tuberosus and L. ventripunctatus in having mostly smooth (versus tuberculate) skin texture with no skin ridges, and from L. pictus and L. pluvialis in having relatively short tibia (male TIB:SVL 0.44– 0.46, versus 0.55–0.61 in L. pictus and0.52–0.56 in L. pluvialis). Leptolalax bidoupensis can be further differentiated from the two most morphologically similar species, L. applebyi and L. melicus, by having slight lateral fringing on feet, and by being approximately 19% larger than L. A NEW MEGOPHRYID FROG SPECIES FROM SOUTHERN VIETNAM Zootaxa 2796 © 2011 Magnolia Press · 23 applebyi (Wilcoxin rank-sum test, W= 16, p = 0.0284; N=8) and 14% larger than L. melicus (Wilcoxin rank-sum test, W= 28, p = 0.0106; N=11). In life, L. bidoupensis also weighs more than either species (L. bidoupensis males 1.1–1.4 g, N=4; L. applebyi males 0.8–0.9 g, N=7; L. melicus males 0.7–0.75 g, N=2). TABLE 3. Iris colouration in life for species within the genus Leptolalax. Species Iris bi- Iris colouration Source coloured L. aereus* No Bronze with minute, black reticulations. Rowley et al. 2010c L. alpinis No? Copper/gold with black reticulations. Fei et al. 2010 (from figures on p. 169) L. applebyi* No Coppery gold to gold with black reticulations. Rowley & Cao 2009 L. arayai Yes Upper 100–120° bright red-brown, rest whitish grey to light Malkmus et al. 2002 greyish green, with black reticulations. L. bidoupensis sp. nov.* Yes Upper half coppery red, lower half fading to pale silver. Exten- Present paper sive black reticulations. L. bourreti ? ? – L. croceus* No Pale gold with distinct, black reticulations encircling the pupil. Rowley et al. 2010a L. dringi Yes Upper section reddish, rest pale golden grey. Black reticula- Inger & Stuebing tions. 2005 (from Fig. 19) L. fuliginosus* Yes Upper half reddish orange, lower half gold. Black reticulations. Matsui 2006 L. gracilis Yes Upper section reddish orange, rest pale golden grey. Black Inger & Stuebing reticulations. 2005 (from Fig. 20) L. hamidi Yes Upper half reddish, rest pale golden grey, with black reticula- Inger & Stuebing tions. 2005 (from Fig. 21) L. heteropus* ? ? – L. kajangensis* No Metallic gold with black reticulations. Grismer et al. 2004 L. kecil* No Dark red. Matsui et al. 2009 L. khasiorum* Yes Upper third bright orange, rest yellowish cream. Black reticula- Das et al. 2010 tions. L. lateralis ? ? – L. liui ? ? – L. maurus No Reddish brown. Malkmus et al. 2002 L. melanoleucus* Yes Upper half orange, lower half silver. Black reticulations. Matsui 2006 L. melicus* No Dark gold. Black reticulations. Rowley et al. 2010b L. nahangensis* No Gold with black reticulations. Lathrop et al. 1998 L. oshanensis No? Copper/gold with black reticulations. Fei et al. 2010 (from figures on p. 172) L. pelodytoides ? ? – L. pictus* Yes Upper 120° with segments of dark brown, rest golden-grey. Malkmus et al. 2002 Black reticulations L. pluvialis* No? Brown? Ohler et al. 2000 (from Fig. 2) L. solus* Yes Upper half dark red, lower half dark brown Matsui 2006 L. sungi* No Gold-green. Black reticulations only at edges Lathrop et al. 1998 L. tamdil* Yes Upper third bright orange, rest greyish cream, edged with black Sengupta et al. 2010 reticulations. L. tuberosus No Pale gold with minute black reticulations. Rowley et al. 2010a L. ventripunctatus ? ? – * documented for type specimens. 24 · Zootaxa 2796 © 2011 Magnolia Press ROWLEY ET AL.