Bioacoustics of Megophrys

Bioacoustic comparison: The male advertisement call of this new species differs from those of its three congeners (subgenus Panophrys ) found in mainland Southeast Asia, north of the Isthmus of Kra and neighbouring provinces of China for which calls have been described. The male advertisement call of Megophrys fansipanensis sp. nov. recorded at 15.3–18.3 °C differs from M. boettgeri (data from Wang et al . 2014 based on recordings made at 15.0–18.0 °C) by having an average call duration of 41.9 (34–49) ms, N =60 (versus average 54 ms, N= 76, in M. boettgeri ), and lacking clear harmonics (versus relatively clear harmonics at approximately 5 kHz in M. boettgeri ); from M. minor (data from Jiang et al . 2001 based on recordings made at 14 °C) by having a dominant frequency of 3.6–4.7 kHz, N =60 (versus 3.4–3.5 kHz, N =10, in M. minor ), a call duration of 34–49 ms, N =60 (versus 75–110 ms, N =14, in M. minor ), a call repetition rate of 3.92 (3.82–3.97) calls/s, N =60 (versus average 4.0 calls/s, N =14, in M. minor ), and lacking clear harmonics (versus relatively clear harmonics at approximately 7.2 kHz in M. minor ); from M. rubrimera (data from Tapley et al . 2017a , recorded at 21.0–22.9 °C) by having the following, nonoverlapping call parameters, an average dominant frequency of 3.8 (3.6–4.7) kHz, N =60 (versus average 3.3 [3.2– 3.4] kHz, N =60, in M. rubrimera ), an average call duration of 41.9 (34–49) ms, N =60 (versus average 73.3 [62–85] ms, N= 60, in M. rubrimera ), an average call repetition rate of 3.9 (3.8–4.0) calls/s, N =60 (versus an average of 3.3 [3.1–3.3] calls/s, N= 60, in M. rubrimera ), and distinctly pulsed calls (versus indistinct pulsation in M. rubrimera ).

Distribution and conservation status: Megophrys fansipanensis sp. nov. is only known from Mount Fansipan at elevations between 2200 and 2813 m asl ( Fig. 1 ). We did not detect this species during field work on Mount Fansipan at elevations of 1200 m asl ( June 2016 ), 1700 m asl ( June 2012 & 2016) and 1900 m asl ( June 2016 ; September 2017 ). The high elevation sites where this species occurs face the immediate threat of habitat degradation and pollution due to the impact of tourists; habitats were noticeably polluted with garbage and runoff from toilets near the stream ( Fig. 6D ). In addition, if Megophrys fansipanensis sp. nov. is restricted to a narrow, high-elevation band, then the species is likely to be particularly vulnerable to climate change. Megophrys fansipanensis sp. nov. may occur more widely, but it was not detected in habitat similar to that found on Mount Fansipan during recent surveys on Mount Ky Quan San, 25 km north of our collection sites. The current EOO is predicted to be 630 km 2 , qualifying Megophrys fansipanensis nov. sp. for listing as Endangered in accordance with the IUCN Red List of Threatened Species categories and criteria B1ab(iii). Comparisons: Megophrys fansipanensis sp. nov. can be distinguished from all species in the subgenus Panophrys found in mainland Southeast Asia, north of the Isthmus of Kra and neighbouring provinces of China on the basis of morphology, and from all species in the subgenus for which comparable data is available on the basis of molecular and acoustic data. Comparisons with each subgenus are discussed separately below. The following comparison is based on 13 males and two females of Megophrys fansipanensis sp. nov.

Description of holotype (measurements provided in Table 2 ): Sexually mature male ( Figs. 3 A–B, 4A–C, 5A & 5D). Head small, longer than wide; snout rounded in dorsal view, obtusely protruding in lateral view, rostral appendage absent; loreal region vertical and concave; canthus rostralis angular; dorsal region of the snout slightly concave; eye length 33% greater than the maximum diameter of the tympanum and subequal to the snout length; eye to tympanum distance shorter than the maximum tympanum diameter; tympanum circular. Pupil in life oval, vertically orientated when dilated; nostrils orientated laterally, equidistant to the eyes and the snout; internarial distance is less than the width of the upper eyelids, and less than the narrowest point between the upper eyelids; upper eyelid width and the shortest distance between the upper eyelids are subequal to one another. Vomerine ridges present, oblique and barely separated anteriorly, with small vomerine teeth; maxillary teeth present. Tongue moderately large and not clearly notched posteriorly. Forelimbs moderately long and thin, forearm length shorter than the hand length; fingers long and narrow, without lateral fringes, finger length formula FI<FII<FIV<FIII; interdigital webbing, and subarticular, supernumerary and palmar tubercles absent; thenar tubercle weak; finger tips slightly expanded relative to the adjacent digit width and flattened to oval pads; terminal grooves absent. Hindlimbs relatively long and thin; thigh length shorter than the shank length, and shorter than the foot length; toes long and rounded in cross section without dermal fringes, toe tips slightly dilated relative to the adjacent digit width and flattened to oval pads, terminal grooves absent; toes not webbed; outer metatarsal, subarticular and supernumerary tubercles absent; inner metatarsal tubercle very weakly defined. Skin of the dorsal surfaces of the body, limbs and dorsal and lateral surfaces of the head weakly granular; tympanum granular with borders slightly raised; very small palpebral horn present on the outer edge of the upper eyelids; supratympanic fold narrows as it passes above the tympanum and expands in width again posteriorly, terminating above the axilla; flanks with small scattered tubercles; thin dorsolateral fold on each side, extending from behind the supratympanic fold to approximately two-thirds of the distance to the groin; a weak, “V”-shaped parietoscapular ridge present, its two sides extending posteriorly from above the tympanum and meeting medially beyond the level of the axilla; a second inverted “V”-shaped sacral ridge present on the mid-dorsum which joins laterally with the dorsolateral folds; “V”-shaped ridges joined at their apices by a medial ridge; small tubercles arranged into distinct transverse rows on the dorsal surfaces of thighs, shanks and forearms. Large distinct tubercles present on the inguinal region; gular region, chest and ventral surfaces of the limbs smooth; pectoral glands distinct, small, slightly raised, positioned level with the axilla; femoral glands small, slightly raised, one positioned equidistant from the knee and cloaca on the posterior surface of each thigh; white skin asperities form a narrow band circummarginally on the upper and lower jaw; black asperities on the tubercles of the tympanic regions and posterior parts of the upper eyelids, absent elsewhere. Colour of holotype : In life ( Fig. 5A & 5D ): Dorsally dark tan; darker lines follow the opposing “V”-shaped parietoscapular-sacral ridges; dark brown loreal region, snout pale; darker brown triangular marking between the eyes with a lighter central blotch; vertical dark brown bar below the eyes; dark brown tympanum; dark tan hind and forelimbs possess dark bars; supratympanic fold longitudinally bicolored, cream above and dark brown below; tubercles in the inguinal region either a pale cream or dark brown in colour; gular and pectoral region dark greybrown, gular region with a medial dark brown stripe; abdomen lighter with dark blotches and speckling; ventral surface of the thighs speckled with brown; iris metallic orange with black reticulations throughout. In preservative ( Figs. 3 A–B & 4A–C): The majority of the dorsal and lateral surfaces of the head, body, forelimbs and hindlimbs brown; darker brown triangular marking with light central blotch between the eyes; darker brown “X”-shaped marking over the opposing “V”-shaped parietoscapular-sacral ridges; dorsolateral and supratympanic folds and flank tubercles brownish-cream; the front of the snout and the lateral canthus rostralis dark brown; wide vertical dark brown bar below the eyes; dark brown tympanum; two dark brown blotches on the anterior lateral surface of the forearms; dorsal surface of FII, FIII, FIV with dark brown blotches. The gular region, chest and the anterior part of the abdomen primarily creamy-grey, with brown speckling on the gular region; gular region with a medial dark brown stripe. Abdomen blotched with dark brown, large brown blotches incompletely encircle the abdomen; ventral surfaces of the thighs and shanks with pale brown mottling; ventral surfaces of the feet grey-brown; area surrounding the vent dark brown; forelimbs ventrally mottled and blotched with light and dark brown; ventral surface of the hands grey; pectoral and femoral glands white. Refer to Figures 3 A–B, 4A–C, 5A & 5D for further details of markings and colouration. Variation: Morphometric measurements of the type series and referred specimen are given in Table 2 . Paratypes and the referred specimen generally agree with the holotype morphologically, but with the following exceptions: Head width greater than the head length for 11 of the 15 specimens (VNMN 0 7033, AMS R186106¯10, AMS R186112, AMS R186113, AMS R186115, AMS R186116 & HLNP 20120620 00002); head width less than the head length for two specimens (VNMN 2018.01 & AMS R186105), and head length equal to the head width for two specimens ( AMS R186111 & AMS R186114 ) . Seven of the 15 specimens ( VNMN 2018.01 , AMS R186106 & AMS R186109 ¯13) lack dermal fringes on the toes whereas indistinct ridges were observed on the toes of five specimens ( VNMN 0 7033, AMS R186105 , AMS R186107 , AMS R186108 & HLNP 20120620 00002). It is not clear if the presence of indistinct ridges on the toes of these specimens is an artefact of specimen positioning during the fixation process. Relative finger lengths differ from the holotype as follows : FI<FIV<FIII<FII (AMS R186107); FI<FIVẼFIII<FII (AMS R186109); FI<FIIẼFIV<FIII (AMS R1 86108 & AMS R186112); and FIẼFII<FIV<FIII (AMS R186113). The tongue of the holotype is not clearly notched posteriorly, but several specimens in the series (VNMN 0 7033, AMS R186105¯6, AMS R186109 & AMS R186111¯3) appeared to have notched tongues posteriorly. The absence of a notch may be an artefact of fixation. Colouration in life is highly variable, one female specimen being a uniform brick red on the dorsal surface (VNMN 07033; Figs. 5B, 5C, 5E & 5H ). Most specimens in the series have opposing “V”-shaped parietoscapular and sacral ridges but two specimens (AMS R186113, Fig. 5F ; VNMN 0 7033, Fig. 5C ) have only the “V”-shaped parietoscapular ridge and lack sacral ridges. The underside of the thighs are blotched or speckled with brown on some specimens ( Fig. 5 H–J). Secondary sexual characters: All males had slightly raised nuptial pads covered with black microspinules; nuptial pads covering most of the dorsal surface of the base of FI and FII; nuptial pad oval shaped on FI and FII; a protruding fleshy projection posterior to the cloaca (a secondary sexual character of some male Megophrys , e.g., M . caudoprocta Shen, 1994 , M . koui and M . pachyproctus ) is absent on all male specimens. Female (AMS R186116) with large unpigmented ova; both examined females did not possess nuptial pads or dermal asperities. Molecular data: 16S sequences were generated from three adult specimens. Uncorrected p -distance between Megophrys fansipanensis sp. nov. and other Megophrys sequences available on GenBank were ±3.3%. Advertisement call: Call descriptions are based on the call groups of the holotype (VNMN 2018.01, N= 3) and two paratypes (AMS R186114 [ N= 12] & AMS R186115 [ N= 5]), and the total number of calls analysed ( N =60) was 20 calls per individual. Advertisement calls were recorded at 15.3–18.3 °C ambient temperature. Calls were an average of 42 ms (34–49 ms) in duration ( Table 3 , Fig. 7 ). Calls contained an average of 11.6 pulses (10–14). The average dominant frequency of all analysed calls was 3.8 kHz (3.6–4.7 kHz). Calls were repeated at a rate of 3.9 (3.8–4.0) calls per second in the analysed call groups and had an average intercall interval of 204.4 ms (180–290 ms). The number of calls within each call group ranged from 1 to 41, with an average of 22.8. Most call groups began with one to three calls at relatively low amplitude, with amplitude increasing with each call, after which amplitude remained relatively constant ( Fig. 7 ). Individual calls decreased in amplitude over time ( Fig. 7 ). Natural history: Megophrys fansipanensis sp. nov. is associated with upper montane bamboo forest, mixed bamboo forest and disturbed secondary broadleaf forest with a relatively open canopy. All individuals have been encountered at night and found in riparian habitat along 1–2 m wide streams with clear water and rocky stream beds. The majority of individuals were encountered in the month of June, fewer animals were observed in September. Males were calling from stream-side vegetation in June 2012 and June 2016 . A gravid female was found adjacent to a fast-flowing stream in June 2016 . Tadpoles were not observed at this time. During field work in September 2015 , only a single unvouchered male was encountered, no vocalisations were heard and tadpoles were not observed. AMS R186115 VNMN 2018.01 AMS R186114 Number of call groups measured 5 3 12 Number of calls measured 20 20 20 Call duration (ms) 41.2 (38–45) 43.8 (41–49) 40.8 (34–45) Intercall interval (ms) 213.7 (197–245) 197.7 (180–227) 201.8 (168–290) Call repetition rate (calls/s) 3.8 4.0 4.0 Calls/call group 22.2 (18–25) 34.7 (24–41) 11.4 (1–22) Pulses/call 11.1 (10–12) 10.8 (10–11) 12.9 (12–14) Dominant frequency (kHz) 3.6 (na) 3.9 (3.9–4.1) 4.0 (3.6–4.7) Temperature (°C) 15.3 18.3 15.3

Diagnosis: Megophrys fansipanensis sp. nov. differs from its congeners by a combination of the following morphological characters: (1) small size, adult male SVL 30.9–44.3 mm ( N =13), adult female SVL 41.7–42.5 mm ( N =2); (2) small palpebral horn on upper eyelid; (3) toes lacking distinct interdigital webbing and lateral dermal fringes indistinct or absent; (4) TYD/EL 53.0–80.0% in adult males ( N =13), 63.0–71.0% in adult females ( N =2); (5) subarticular tubercles absent; (6) palmar tubercles absent; (7) inner metatarsal tubercle present; (8) SHL/SVL 49.0–58.9% in adult males ( N =13), 55.8–58.5% in adult females ( N =2); (9) tympanum clearly defined; (10) vomerine ridges, vomerine and maxillary teeth present; and (11) an advertisement call with an average dominant frequency of 3.8 kHz at 15.3–18.3 °C.

Distribution and conservation status: This species is known from between 1898–2242 m asl at four localities, up to 92 km apart in the Hoang Lien Range (Sa Pa District, Lao Cai Province , HLNP; Tam Duong District, Lai Chau Province ; Mount Ky Quan San, Bat Xat District, Lao Cai Province and Che Tao Commune, Yen Bai Province , Vietnam ). The species’ EOO is currently predicted to be 3216 km 2 ( Fig. 8 ). Ongoing disturbance to the species’ habitat due to forest clearance for agriculture has been observed in the parts of Sa Pa District where the species was observed. In Bat Xat District the forest in which this species occurs is being negatively impacted by fuelwood collection for the tourism industry and by the grazing of livestock. Megophrys hoanglienensis sp. nov. is likely to qualify as Endangered in accordance with the IUCN Red List of Threatened Species categories and criteria B1ab(iii). AMS R186122 Number of call groups measured 5 Number of calls measured 20 Call duration (ms) 102.9 (96–108) Intercall interval (ms) 274.2 (178–565) Call repetition rate (calls/s) 2.6 Calls/call group 17 (11–21) Pulses/call 18.7 (12–22) Dominant frequency (kHz) 3.0 (2.8–3.0) Temperature (°C) 18.5 Comparisons: Megophrys hoanglienensis sp. nov. can be distinguished from all species in the subgenus Panophrys found in mainland Southeast Asia, north of the Isthmus of Kra and neighbouring provinces of China on the basis of morphology, and from all species in the subgenus for which comparable data is available on the basis of molecular and acoustic data. Comparisons with each subgenus are discussed separately below. The following comparison is based on 11 males and one female of Megophrys hoanglienensis sp. nov.

Description of holotype (measurements provided in Table 4 ): Sexually mature male ( Figs. 9 A–B, 10A–C, 11A–B). Head small, wider than long; snout mucronate in dorsal view, obtusely protruding in lateral view, rostral appendage absent; loreal region vertical and concave; canthus rostralis angular; dorsal region of the snout slightly concave; eye length 38.0% longer than the maximum tympanum diameter and subequal to the snout length; eye to tympanum distance greater than the maximum tympanum diameter; tympanum circular. Pupil in life oval, vertically orientated when dilated; nostrils orientated laterally, closer to the eyes than the snout; internarial distance subequal to the width of the eyelids, and subequal to the narrowest point between the upper eyelids; vomerine ridges present, oblique and barely separated anteriorly, with small vomerine teeth; maxillary teeth present. Tongue moderately large and feebly notched posteriorly. Forelimbs moderately long and thin; forearm length shorter than the hand length; fingers long and narrow without lateral fringes, finger length formula FI=FII=FIV<FIII; interdigital webbing absent; subarticular, supernumerary and palmar tubercles absent; thenar tubercle weak; finger tips slightly expanded relative to the adjacent digit width and flattened to oval pads; terminal grooves absent. Hindlimbs relatively long and thin; thigh length shorter than the shank length, but greater than the foot length; toes long and rounded without dermal fringes, toe tips slightly dilated relative to adjacent digit width and flattened to oval pads, terminal grooves absent; toes with rudimentary webbing; outer metatarsal tubercle, subarticular and supernumerary tubercles absent; inner metatarsal tubercle weakly defined. VNMN 2018.02 * Field tag JJLR 0 2663 VNMN 0 7034 ** Field tag JJLR 0 2391 AMS R 186118 ** Field tag JJLR 0 2392 AMS R 186119 ** Field tag JJLR 0 2607 AMS R 186120 ** Field tag JJLR 0 2639 AMS R 186121 ** Field tag JJLR 0 2666 AMS R 186122 ** Field tag JJLR 0 3820 AMS R 186123 ** Field tag JJLR 0 3821 HLNP 20120624 00003 ^ Field tag JJLR 0 3822 AMS R 186125 ** Field tag JJLR 0 3837 AMS R 186126 ** Field tag JJLR 0 3838 AMS R 186127 ** Field tag BX 0 0 3 Sex M M M M M F M M M M M M SVL 43.2 41.1 47.6 46 46.6 59.6 44.5 41.9 42 37.4 41.6 43.1 HW 17.2 16.0 17.8 18 19.7 22.8 18.7 17.2 17.5 15 16.9 18.1 HL 16.3 15.1 16.5 17.8 19.0 21.4 17.7 16.1 17.2 14.9 16.5 17.2 IFE 5.2 5.6 6.1 6.4 5.4 7.7 5.7 4.6 5.4 3.8 5.1 6.6 IBE 5.9 6.0 7.1 6.8 6.7 8.4 5.7 5.6 5.2 4.9 5.0 7.7 EL 4.2 5.1 5.2 4.8 5.9 6.5 4.6 4.7 4.8 3.9 4.7 4.7 TYD 2.5 2.8 3.4 3.6 3.2 3.9 2.7 2.9 2.8 2.4 3.4 3.1 TYE 2.9 3.3 3.1 3.2 3.1 4.5 3.5 4.0 3.2 3.2 2.6 2.9 SL 6.2 5.3 6.3 6.6 7.4 7.5 7.1 6.3 6.5 4.6 5.1 6.4 EN 2.6 3.2 2.4 3.2 2.8 3.5 3.1 2.6 2.7 2.1 2.1 2.9 SN 3.1 2.6 3.7 4.0 4.1 3.7 3.3 3.0 2.9 2.6 3.6 3.4 IUE 5.0 5.5 5.4 5.5 5.9 7.8 5.3 4.8 4.6 4.4 7.6 5.1 IN 4.4 4.8 5.9 5.6 6.3 7.6 5.9 4.9 4.6 3.3 4.8 5.3 UEW 5.2 4.3 5.4 5.4 5.2 7.0 5.5 5.1 5.2 3.7 4.0 4.6 FAL 9.6 9.5 10.4 10.4 11.5 13.2 10.7 10.6 10.5 8.8 8.8 9.2 HAL 11.8 11.1 13.7 13.8 13.9 16.4 12.1 12.0 12.6 11.1 12.2 13.4 FIL 5.1 4.2 8.1 6.8 6.1 6.6 6.6 5.1 5.6 5.3 5.2 5.8 FIIL 5.1 4.9 4.9 5.7 5.2 6.4 5.9 5.2 5.7 4.8 5.5 5.5 FIIIL 8.5 7.7 8.5 9.6 9.3 11.4 8.8 8.2 8.8 7.6 8.7 9.4 FIVL 5.1 4.8 5.9 6.3 5.4 6.9 5.8 6.0 5.4 5.6 4.7 5.7 SHL 26.0 23.5 21.1 27.5 28.1 34.3 24.5 24.3 24.4 23.0 26.0 26.2 TL 25.4 23.0 27.8 27.0 28.7 33.5 25.4 24.6 25.1 21.6 23.4 24.7 FOL 23.0 20.4 25.1 25.2 25.3 32.3 24.5 22.0 23.4 19.8 21.4 22.8 IMT 3.7 3.5 3.8 3.9 2.4 3.8 3.2 2.5 3.3 2.6 2.4 3.0 Weight in life (g) 7.7 6.6 9.3 9.0 11.0 19.5 8.6 6.6 6.4 5.3 6.5 8.8 F1 GURE 10 . Megophrys hoanglienensis sp. nov. in preservative ( holotype VNMN 2018.02 ). (A) Lateral view of head, (B) palmar surface of right hand, and (C) plantar surface of left foot. 5 mm scale bar. Skin of dorsal surfaces of the body, limbs, and both dorsal and lateral surfaces of the head weakly granular; tympanum granular with borders slightly raised; very small palpebral horn present on the outer edge of the upper eyelid; supratympanic fold narrows as it passes above the tympanum, widening again before terminating above the axilla; flanks with small scattered tubercles; thin dorsolateral fold extending posteriorly from behind the supratympanic fold to the groin on each side; a weak, “V”-shaped parietoscapular ridge present, its two sides extending posteriorly from above the tympanum and meeting medially beyond the level of the axilla; a second weak inverted “U”-shaped sacral ridge present on the mid-dorsum which joins laterally with the dorsolateral folds; “V” and “U”-shaped ridges not joined at their apex, apex of ridges with black asperities; small tubercles present on the dorsal surface of the thighs, shanks and forearms. Large distinct tubercles present on the flanks, inguinal region and dorsal surface of the hindlimbs; chest and ventral surfaces of the forelimbs and shanks smooth; pectoral glands obvious, small, slightly raised, positioned level with the axilla; femoral glands small, slightly raised, one positioned equidistant from the knee and the cloaca on the posterior surface of each thigh; white skin asperities forming narrow band circummarginally on the upper and lower jaw; black asperities on the posterior parts of the upper eyelids, on the dorsum and area directly above the vent, and on tubercles of the tympanic regions, absent elsewhere. Colour of holotype : In life ( Figs. 11 A–B): Dorsally dark tan; with light tan blotches; darker brown triangular marking between the eyes with a lighter central blotch; cream supratympanic folds with black lower margin; inguinal regions light tan; dark brown hindlimbs with darker blotches; posterior surface of the thighs and shanks very dark brown; forearms light brown with a single dark bar; dorsal surface of the fingers with dark brown bars; gular, pectoral and abdomen region mottled grey-brown and cream, gular region with a medial dark brown stripe; ventral surface of thighs unmarked; pectoral glands off-yellow; iris metallic gold. In preservative ( Figs. 9 A–B & 10A–C): Markings are the same as in life, colouration differences are as follows: The majority of the dorsal and lateral surfaces of the head, body, forelimbs and hindlimbs brown; dorsolateral and supratympanic folds and flank tubercles brownish-cream; tip of the snout cream, lateral canthus rostralis dark brown; wide vertical dark brown bar below the eyes; dark brown band running from the posterior edge of the eyes, through the tympanum, and terminating where the forelimbs join the body; lower margin of the supratympanic folds dark brown, sharply contrasting with creamy grey upper margin; gular region, chest and anterior part of the abdomen primarily creamy-grey with brown speckling, and a dark brown medial bar in the gular region; dark brown blotches anterior to each of the pectoral glands, pectoral glands light cream in colour; abdomen speckled with dark brown; ventral surfaces of the thighs and shanks with diffuse pale brown mottling; ventral surfaces of the feet grey brown with cream digit tips; area surrounding the vent dark brown; forelimbs ventrally mottled with dark brown bar running longitudinally on the outer ventral surface of the forearms; grey colouration of ventral side of the hands with darker mottling and cream digit tips; femoral glands cream. Variation: Morphometric measurements of the type series and referred specimens are provided in Table 4 . Nuptial pads were not visible on one male specimen (AMS VNMN 07034) although sex was confirmed via inspection of the gonads. Two (AMS R186120 & AMS R186121) of the 12 specimens possessed rudimentary webbing between the toes. Four (VNMN 0 7034 & AMS R186118¯20) of the 12 specimens with indistinct lateral fringes on the toes. It is not clear if the presence of indistinct fringes on the toes of these specimens is an artefact of the positioning during the fixation process. Relative finger lengths are highly variable, and only one specimen (AMS R186119; FIẼFII<FIV<FIII) had the same relative finger length formula as the holotype ; the remaining specimens differed from the holotype specimen in having the following relative finger lengths; FI<FIII<FIV<FII (VNMN 07034), FI<FII<FIV<FIII (AMS R186118, AMS R1 86120, AMS R186121 & AMS R186123), FII<FI<FIV<FIII (AMS R1 86122, AMS R186125 & AMS R186127), FIV<FI<FII<FIII (HLNP 2 0 120624 00003), and FIV<FI<FII<FIII (AMS R186126). Colour in life is variable from tan to dark brown ( Fig. 11 A–H). Secondary sexual characters: All sexually mature males had slightly raised nuptial pads covered with black microspinules, nuptial pads covering most of the dorsal surface of the base of FI and FII; nuptial pad oval shaped on FI and FII; the protruding fleshy projection posterior to the cloaca (a secondary sexual character of some male Megophrys , e.g., M . caudoprocta , M . koui and M . pachyproctus ) is absent on all male specimens. The examined female did not possess nuptial pads. Molecular data: Sequences generated from nine adult specimens in this study ( Table 1 ) were most similar to “ Megophrys sp. 5” ( sensu Chen et al . 2017 ) collected in Sa Pa District, Lao Cai Province , Vietnam ; 16S rDNA sequences differed by only 0.3% uncorrected p -distance between “ Megophrys sp. 5” and our specimens (from Lao Cai & Lai Chau provinces), and by 1.1% for specimens collected approximately 58 km away in Yen Bai Province , which is greater than the intraspecific variation reported elsewhere within the subgenus Panophrys (e.g., Li et al . 2014 reported an intraspecific p -distance of 0.6% between populations of M. obesa Wang, Li & Zhao, 2014 ), but is lower than intraspecific variation within Megophrys sensu lato (e.g., 3.1% mean uncorrected p -distance in M. elfina ; Poyarkov et al . 2017 ). Uncorrected p -distance was 0.8% between specimens of Megophrys hoanglienensis sp. nov. collected from Lao Cai and Lai Chau provinces. The smallest interspecific p- distance in our study was between Megophrys hoanglienensis sp. nov. and Megophrys fansipanensis sp. nov. at 3.3% where the two new species occur in syntopy supporting their position as distinct sister taxa. Advertisement call: Call descriptions are based on the calls of one paratype (AMS R186122), five call groups and 20 calls in total were analysed. Advertisement calls were recorded at 18.5 °C ambient temperature. Calls were an average of 102.9 ms (96–108 ms) in duration ( Table 5 ; Fig. 13 ). The average number of pulses per call was 18.7 (12–22). The average dominant frequency of all the analysed calls was 3.0 kHz (2.8–3.0 kHz). Calls were repeated at a rate of 2.6 calls per second for the call groups analysed, and had an average intercall interval of 274.2 ms (178– 565 ms). The number of calls within each call group ranged from 11 to 21, with an average of 17. Most call groups began at a relatively low amplitude, increasing with each call up to approximately a quarter of the duration, after which amplitude varied but did not follow a trend or pattern ( Fig. 13A & B ). Individual calls began with a medium relative amplitude but peaked early before declining towards the end, and the interval between the first and second pulse was longer than those between any of the following pulses ( Fig. 13C ). Natural history: Megophrys hoanglienensis sp. nov. is associated with disturbed secondary broadleaf forest with a relatively open canopy. All individuals have been found at night in riparian habitat along clear water, 2–5 m wide streams with a gravel or rocky substrate. Males were calling from streamside vegetation in June 2012 and June 2016 but not during the surveys undertaken in May or September 2017 . A non-gravid female was found adjacent to a wide, low-gradient, shallow stony stream in June 2016 . Tadpoles were not observed at any of the locations where this species has been found (surveys undertaken in June 2012 and 2016, and in September 2015 and 2017). Megophrys hoanglienensis sp. nov. is sympatric with M. jingdongensis at 1990 m asl on Mount Fansipan, and with M. fansipanensis sp. nov. on Mount Fansipan at 2242 m asl. Megophrys hoanglienensis sp. nov. is sympatric with M. jingdongensis and M. rubrimera on Mount Ky Quan San at 2153 m asl ( Tapley et al. 2018b ).

Diagnosis: Megophrys hoanglienensis sp. nov. differs from its congeners by a combination of the following morphological characters: (1) medium adult size, adult male SVL 37.4–47.6 mm ( N= 11); adult female SVL 59.6 mm ( N= 1); (2) small palpebral horn on upper eyelid present; (3) interdigital webbing and lateral fringes on the toes indistinct or absent; (4) snout mucronate in dorsal view; (5) TYD/EL 54.2–75.0% in adult males ( N =11), 60.0% in adult female ( N= 1); (6) subarticular tubercles absent; (7) palmar tubercles absent; (8) inner metatarsal tubercle present; (9) HW/SVL 37.4–42.5% in adult males ( N= 11), 38.3% in adult female ( N= 1); (10) SHL/SVL 44.3– 62.5% in adult males ( N =11), 57.5% in adult female ( N= 1); (11) tympanum clearly defined; (12) TYE/SVL 6.3– 9.5% in adult males ( N= 11), 7.6% in adult female ( N= 1); (13) vomerine ridges, vomerine and maxillary teeth present; (14) an advertisement call with a dominant frequency of 2.8–3.0 kHz at 18.5 °C.

Bioacoustic comparison: The male advertisement call of Megophrys hoanglienensis sp. nov. recorded at 18.5 °C differs from those of its four congeners (subgenus Panophrys ) found in mainland Southeast Asia, north of the Isthmus of Kra and neighbouring provinces of China for which calls have been described. The male advertisement call of Megophrys hoanglienensis sp. nov. differs from M. boettgeri (data from Wang et al . 2014 , recorded at 15.0– 18.0 °C) by having an average call duration of 102 (96–108) ms, N =20 (versus average 54 ms, N =76, in M. boettgeri ), and lacking clear harmonics (versus relatively clear harmonics at approximately 5.0 kHz in M. boettgeri ); from Megophrys fansipanensis sp. nov. (recorded at 15.3–18.3 °C; this study) by having an average call duration of 102 (96–108) ms, N =20 (versus 41.9 [34–49] ms, N= 60, in Megophrys fansipanensis sp. nov. ), an average dominant frequency of 3.0 (2.8–3.0) kHz, N =20 (versus average 3.8 [3.6–4.7] kHz, N= 60, in Megophrys fansipanensis sp. nov. ), and an average of 18.7 (12–22) pulses per call, N =20 (versus 11.6 [10–14] pulses per call, N =60, in Megophrys fansipanensis sp. nov. ); from M. minor (data from Jiang et al . 2001 , recorded at 14.0 °C) by having the following call parameters, a dominant frequency of 2.8–3.0 kHz, N =20 (versus 3.4–3.5 kHz, N= 13, in M. minor ), a call repetition rate of 2.6 calls/s, N =20 (versus average 4.0 calls/s, N =14, in M. minor ), and lacking clear harmonics (versus relatively clear harmonics at approximately 7.2 kHz in M. minor ); from M. rubrimera (data from Tapley et al . 2017a , recorded at 21.0–22.9 °C) by having an average dominant frequency of 3.0 (2.8–3.0) kHz, N =20 (versus average 3.3 [3.2–3.4] kHz, N =60, in M. rubrimera ), an average call duration of 102 (96–108) ms, N =20 (versus average 73.3 [62–85] ms, N= 60, in M. rubrimera ), and a call repetition rate of 2.6 calls/s (versus an average of 3.3 [3.1–3.4] calls/s, N =60, in M. rubrimera ).