Megophrys (Xenophrys) robusta Boulenger, 1908

Mahony, Stephen, Kamei, Rachunliu G. & Teeling, Emma C., 2018, Cryptic diversity within the Megophrys major species group (Amphibia: Megophryidae) of the Asian Horned Frogs: Phylogenetic perspectives and a taxonomic revision of South Asian taxa, with descriptions of four new species, Zootaxa 4523 (1), pp. 1-96 : 30-37

Megophrys (Xenophrys) robusta Boulenger, 1908

( Figures 11 & 12 ; Table 1)

[?] Xenophrys gigas Jerdon 1870:85 (partim: “ Sikim ”). In: Notes on Indian herpetology. Proceedings of the Asiatic Society of Bengal, March, 1870: 66–85.

Megalophrys robusta Boulenger 1908:418 , pl. xxiv. In: A revision of the oriental pelobatid batrachians (genus Megalophrys ). Proceedings of the Zoological Society of London, 1908: 407–430 + pl. xxii–xxv + fig. 71.

Lectotype (by present designation). Adult female ( BMNH 1947.2 .25.19 [RR 1908.4.8.8]: Figure 11 ), from “Darjeeling” (~ 27°03'0"N, 88°16'0"E), West Bengal, India, collector J. Gammie, presented by the Indian Museum, collection date unknown.

Paralectotypes (by implication). Two adult females ( ZSIC 9681 , ZSIC 10777 ), details are the same as for the lectotype .

Examined specimens. BMNH 1947.2 .25.19 (lectotype: Figure 11 ) ; ZSIC 9681 , ZSIC 10777 (paralectotypes); three adult males ( SDBDU 2011.1057 ; SDBDU 2011.1062 : Figure 12A ; SDBDU 2011.1064 : Figure 12F ), and one adult female ( SDBDU 2011.1063 : Figure 12C & F ), from near Latpanchar town (26°54'34.2"N, 88°23'54.54"E, 1135 m asl.), Kurseong sub-division, Darjeeling district , West Bengal, Northeast India, collected by the Systematics Lab members on 0 5 and 0 6 June 2011; a subadult male ( SDBDU 2011.415 : Figure 12E ), and an adult male ( SDBDU 2011.416 : Figure 12E ), from Stream 1 (26°54'34.7"N, 88°23'52.7"E, 1030 m asl.), Mahananda Wildlife Sanctuary , Latpanchar township , Kurseong sub-division, Darjeeling district , West Bengal, Northeast India, collected by SDB and RGK on 19 and 20 May 2011; two adult males ( SDBDU 2009.1270 ; SDBDU 2009.1284 : Figure 12B ), and one juvenile ( SDBDU 2009.1245 : Figure 12D ), from Sessa village (27°6'4.02"N, 92°31'38.52"E, 1110 m asl.), West Kameng district , Arunachal Pradesh state, Northeast India, collected by the Systematics Lab members on 07–09 August 2009 .

Lectotype description (measurements in mm). Mature female (SVL 102.0) ( Figure 11 ). Head large, wider than long (HW 42.0, HL 40.9, IFE 16.9, IBE 30.1); snout broadly pointed in dorsal view, obtusely protruding beyond mandible in lateral view, without rostral appendage ( Figure 11C ); loreal region acute, concave, with well developed canthus rostralis; dorsal surface of snout concave; eye diameter larger than maximum tympanum diameter, and shorter than snout (EL 11.6, TYD 7.7, SL 13.2); eye–tympanum distance (TYE 8.2) slightly longer than tympanum diameter; tympanum distinctly oval, obliquely orientated ( Figure 11C ), upper ~40% concealed by supratympanic ridge; pupil indistinct; nostril positioned laterally, circular with raised posterior rim, closer to eye than to snout (EN 5.3, SN 7.4); internarial distance equal to narrowest point between upper eyelids, and greater than upper eyelid width (IN 11.9, IUE 12.0, UEW 10.0); pineal ocellus absent; vomerine ridges well developed, positioned between choanae, closer to choanae than to each other; vomerine teeth absent; maxillary teeth present; tongue large, rounded posteriorly (possibly artefact of preservation), medial lingual process absent.

Forelimbs moderately long, thin ( Figure 11A & B ), forearm slightly enlarged relative to upper forelimb, slightly shorter than hand length (FAL 25.4, HAL 26.3); fingers long, narrow ( Figure 11D ), finger length formula IV<I=II<III (FIL 12.5, FIIL 12.5, FIIIL 18.2, FIVL 12.2); interdigital webbing and lateral fringes on digits absent; subarticular, supernumerary and metacarpal tubercles absent; thenar tubercle weakly developed; digit tips slightly expanded, flattened, without pads or terminal grooves. Hindlimbs relatively long, thin ( Figure 11A & B ); thighs slightly shorter than shanks, and feet (TL 50.0, SHL 52.6, FOL 50.8); toes long, thin ( Figure 11E ), relative toe lengths I<II<V<III<IV; digit tips slightly dilated, flattened, without distinct pads; base of digits with thick rudimentary webbing; lateral fringes on toes absent; outer metatarsal, subarticular and supernumerary tubercles all absent; inner metatarsal tubercle present but barely distinguishable; ridge of thickened skin on ventral surface of digits absent.

Skin on dorsal surfaces of snout, head, and back weakly granular; throat, chest, and limbs smooth; flanks densely granular with ~15 small scattered tubercles (counted on left side); entire abdomen wrinkled (artefact of preservation), posterior thighs and cloacal region finely granular; tympanum smooth, slightly concave; sides of head finely granular; palpebral horn absent but with a broad fleshy bump in its place; supratympanic ridges narrow anteriorly, widening posteriorly, extends from posterior orbital border, curves down along upper and posterior border of tympanum, terminating above forelimb insertion; narrow, raised dorsolateral ridge on each side, extending posteriorly from behind supratympanic ridge to ~75% trunk length; weak, V-shaped parietoscapular ridge present, extending posteriorly from temporal region on each side, meeting medially beyond level of forelimbs, second short weak inverted U-shaped sacral ridge on posterior dorsum; pectoral glands weakly raised, level with axilla on chest; femoral glands not distinctly raised, on posterior surface of thighs, closer to knee than to cloaca; dermal asperities absent.

Colouration: In preservative ( Figure 11 ): Flanks, and dorsal surface of head, body, and limbs primarily plain brown; large flank tubercles lighter brown, each bordered below by small dark brown blotch; faint solid dark brown triangular marking on dorsal surface of head; wide, slightly oblique dark brown bar below each eye; gular region and chest pale brown with some dark speckling; ventral surfaces of thighs and shanks primarily light with faint mottling; outer tarsi with continuous dark brown blotch extending from base of foot to base of shank along outer edge; surfaces adjacent to cloaca dark brown; forelimbs brown above with two or three dark brown transverse stripes and blotches on forearms; dark brown patch extends from base of hand to elbow on ventral surface; ventral surfaces of hands and feet faded greyish-brown; pectoral and femoral glands white. In life: Not originally documented for lectotype (refer to Figure 12 for other examples).

Variation. Refer to Table 1 for morphometric variation within the type series and referred specimens, consisting of seven adult males, four adult females, a subadult, and a juvenile. The paralectotypes and referred specimens resemble the lectotype for most morphological characters with the following exceptions: Head width/ head length ratios differ between Darjeeling and Arunachal Pradesh specimens—all Darjeeling specimens have HWŽHL (this study, N =9; Deuti & Kumar 2008, N =5), Arunachal Pradesh specimens have HLŽHW (this study, N =3); two specimens (SDBDU 2009.1270, SDBDU 2009.1245) have relative finger length formula IV=I=II<III; on most specimens, vomerine ridges are positioned equidistant from choanae and each other; vomerine teeth vary in size considerably, from indistinct on approximately half of examined specimens, to moderately well-developed on SDBDU 2011.1057; rounded appearance of tongue on lectotype appears to be artefact of preservation, as tongue of other specimens are distinctly, though weakly bifurcate posteriorly; dorsolateral ridges typically extend posteriorly almost to above cloaca on most specimens; tympanic concealment by supratympanic ridge varies from ~10 to 40%; parietoscapular-sacral ridge configurations vary considerably, i.e., “> (”, “>--<”, or only V-shaped parietoscapular ridge present; dermal asperities absent on all females and subadults, but all males from Darjeeling possess black spinular asperities in dense thick circummarginal band along lower and upper jaw, covering entire surface below (and including) supratympanic ridges, more sparse on loreal region, absent from front of snout, sparse on posterior surface of upper eyelids, posterior dorsal surface of head and anterior body, increasing in density posteriorly on dorsum, sparse on dorsal and ventral surfaces of thighs, shanks, and tarsi, but more dense on dorsal surfaces near joints, sparse (or occasionally absent) on posterior surface of abdomen, present along all dorsal ridges, absent from flanks, anterior abdomen, chest and throat. Males from Arunachal Pradesh agree with the Darjeeling specimens regarding asperities cover, except that they have less dense asperities on upper jaw, none on supratympanic ridges except on tympanum and surface posterior to tympanum. Some specimens have weakly developed outer metacarpal tubercles visible; moderately well developed thenar tubercles present on some individuals. Flank tuberculation varies from sparse, small tubercles to large, dense tubercles. Dorsal surfaces of snout usually with small irregularly arranged dark brown blotches ( Figure 12A, B & D ); triangular marking on dorsal surface of head occasionally incomplete; parietoscapular-sacral ridges typically accompanied by dark brown marking that follows shape of dermal ridges, others densely mottled with brown blotches over entire dorsum; general colouration in preservation ranges from pale brown to dark brown dorsally, dorsal marking barely visible (or absent) on darker individuals; some specimens have 3–6 distinct light brown longitudinal stripes extending from gular region onto chest ( Figure 12E & F ); most have faint to distinct brown blotch extending from posterior border of orbit through tympanum ( Figure 12A & B ), and from anterior orbit laterally along edge of canthus rostralis to snout tip; all have distinctly dark brown lower edge to supratympanic ridge and brown vertical bars on sides of face, most distinct is broad bar extending from lower border of orbits to edge of upper lip [erroneously reported to be absent in M. robusta by Ohler et al. (2002)] ( Figures 11C , & 12A, C & E ); outer edge of gular region typically brown, with eight light patches.

Secondary sexual characters. Males: nuptial pads present, weakly raised, covered with black microasperities, covering almost entire dorsal surface of Finger I; nuptial pad on Finger II small, oval, positioned on base of digit on inner dorsal side (extending on to base of proximal phalange on some individuals); external vocal sac usually indistinct, though skin is loose on some individuals indicating large sub-gular sac that extends onto chest; large internal vocal slit is present on floor of mouth near rear of mandible on each side; forearms enlarged relative to upper forelimbs. Females: ova not pigmented; nuptial pads, vocal sac, vocal slits, all absent.

Morphological comparison. Megophrys robusta (adult males N =6, adult females N =4) differs from M. monticola and M. zhangi by its larger adult body size, male SVL 73.5–83.1 mm, female SVL 81.3–108.3 mm (vs. male SVL 38.2–49.5 mm, N =17, female SVL 40.5–56.1 mm, N =6; male SVL 32.5–37.2 mm, N =3, respectively); differs from M. mangshanensis by absence of distinct white upper lip stripe (vs. present). For comparisons with subsequent species covered in this study, refer to relevant morphological comparison sections for those species.

Systematic position. Megophrys robusta was found to be the sister taxon to the MMC clade, and was the second most ancestrally derived lineage in the MMSG after M. monticola ( Mahony et al. 2017; Figures 2 & 3 ; Appendix II, Figures 2 , 3A & 4B ). The inclusion of additional sequences from Chen et al. (2017) in ML analyses demonstrates that M. robusta is the sister taxon to the morphologically similar M. medogensis , however, the systematic position of this clade was not strongly supported ( Figure 4 ; Appendix II, Figures 5 & 6 ). Refer to Appendix I, Table 6 for uncorrected p -distances for the 16S rRNA gene between M. robusta and other MMSG species.

Etymology. The specific epithet “ robusta ” is Latin for “hard” or “strong”, presumably a reference to the heavy set habitus of this species.

Suggested common name: Robust Horned Frog.

Distribution. Specimens examined in this study demonstrate that M. robusta ranges at least from the Darjeeling Hills, northern West Bengal state, east through Bhutan to at least the West Kameng district of western Arunachal Pradesh state, in India ( Figure 8A ). Specimens examined in this study were collected at elevations from 1030 to 1135 m asl. Daniel (1962) reported two specimens (as M. major ) collected from Darjeeling itself (~ 2200 m asl.) and from nearby Darjeeling (~ 1675 m asl.), Darjeeling Sadar division, Darjeeling district, West Bengal. Rai and Anders (2002) provided a number of localities in Ilam, Pachthar and Taplejung provinces in eastern-most Nepal (bordering Darjeeling and Sikkim) from 1600 to 2400 m asl., indicating that the Arun River forms the western boundary for the species. They provided a description of M. robusta and figures, which corresponded well with the species as defined here. This species is also found in neighbouring Sikkim state, in Northeast India ( Subba et al. 2016; Deuti et al. 2017), and may extend north along river valleys into bordering southern Tibet, China. Refer to the Remarks section for discussions on additional localities published elsewhere.

Habitat and natural history. All but one of the specimens collected during this study were collected after dusk from the banks of moderate sized (3–10 m wide), typically shallow rocky hill streams surrounded by at least patches of mature forest, and dense vegetation ( Figure 9A ). In Darjeeling district during late May to early June, calls provisionally attributed to this species (a loud typical Megophrys (Xenophrys) call) were occasionally heard from streams during both day and night surveys. These calls consisted of a short burst of notes followed by a long silence. At Sessa, similar calls were heard during night surveys in early August, and were used on two occasions to locate individuals. However, in both instances the suspected calling males were found within 1–2 m of a similar sized, and considerably more abundant sympatric MMC species (described below) on the banks of the Sessa River, so calls could not be confirmed to be from M. robusta . At both localities, adult male individuals were always widely dispersed along stream banks and were typically found perched on top of large rocks in the open. A female (SDBDU 2011.1063, SVL 81.3 mm: Figure 12C & F ) was collected from a roadside embankment amongst dense vegetation ~ 50 m from the nearest stream. Its ovaries contained small, pigmentless ova. Daniel (1962) reported that a large female (SVL 101 mm) collected in June contained enlarged ova (diameter 2 mm), however, he stated a female specimen (SVL 80 mm) “collected in September had the ovaries dormant”. Though the latter female may have been sexually immature, the available evidence suggests that the breeding season for this species extends at least from mid-May until August. A male specimen (SDBDU 2011.415, SVL 69.1 mm: Figure 12E ) collected in late May had distinctly enlarged testes, but had not yet developed nuptial pads and the internal vocal slits were small indicating that this was not yet sexually mature. SDBDU 2009.1284 (SVL 73.5 mm [ Figure 12B ]) was the smallest sexually mature male. Rai and Anders (2002) provided a number of additional observations that are of interest, however, their description of the call (as “a sharp whistling call, sounding like “ khui-khui ”) does not correspond with the authors’ experience, where calls were distinctly of a lower frequency sound more similar to the “caws” of a crow.

Remarks. Boulenger (1908) described this species based on a syntype series of five specimens collected by Dr. J. Gammie from Darjeeling (four in the Indian Museum [now ZSIC], one in the BMNH [now NHMUK]–the figured specimen). He provided measurements of two ZSIC specimens, a female (SVL 114 mm), and a ‘halfgrown’ individual (SVL 54 mm). Dutta (1997) erroneously claimed (and followed by Bordoloi & Borah 2001) that M. robusta was a replacement name for X. gigas Jerdon, 1870 (now M. major ). Dutta (1997) also only mentioned ZSIC as the location of type specimens. Chanda et al. (2000), apparently also following Dutta (1997), did not mention M. robusta in the list of type specimens in ZSIC, but instead assigned four specimens (ZSIC 9670, ZSIC 9681, ZSIC 10777, ZSIC 10779) as syntypes of Xenophrys gigas . ZSIC 9681 and ZSIC 10777 clearly represent M. robusta . ZSIC 9670 and ZSIC 10779 were not examined in this study. The ZSIC also recognised (according to a jar label) two additional specimens, ZSIC 9650 (an adult female M. monticola , SVL 51.1 mm) and ZSIC 9668 (not examined in detail but represents either an adult M. monticola or juvenile M. robusta ) as ‘types’ of X. gigas . Our study demonstrates that none of these ZSIC specimens are the types of X. gigas (see X. major Remarks section). Thus it remains unclear which four of these six specimens represent the Indian Museum (ZSIC) syntypes mentioned by Boulenger (1908) for M. robusta . For this reason, only BMNH 1908.4.8.8 can currently be verified as a syntype of M. robusta and is therefore designated above as the lectotype of this taxon, thus providing nomenclatural stability by restricting the name to a verified original syntype.

Megophrys robusta has been reported from several localities further east in Arunachal Pradesh state, e.g., Lower Subansiri district ( Bordoloi et al. 2000; Bordoloi & Borah 2001) and the Upper Siang district and Changlang districts ( Pawar & Birand 2001). Pawar and Birand (2001) did not provide a description of the species they observed and did not appear to have collected voucher specimens, so these localities should be considered anecdotal pending collection of verifiable specimens from the Upper Siang and Changlang districts. Several of the morphological characters described by Bordoloi and Borah (2001) indicated that they may have misidentified an MMC species, e.g., “belly with faint darker spots”. They however did not provide further defining and useful taxonomic characters to help identify which species they collected. Therefore, the identities of Bordoloi and Borah’s (2001) specimens require verification before Lower Subansiri district can be included in the verified distribution of M. robusta .

Dutta (1997) reported M. robusta from the Garo Hills in Meghalaya state, presumably misidentifying the large unnamed MMC species (formally described below), so this locality also requires verification based on the reexamination of voucher specimens. Sangma and Saikia (2015) provided an account and figure of a specimen they referred to as M. robusta from Tura Peak, West Garo Hills district in Meghalaya state. This specimen clearly possesses a whitish upper lip stripe (absent on M. robusta ), demonstrating that it is one of the three MMC species present in Meghalaya. An early report of M. robusta from the East Khasi Hills district, Meghalaya by Hora (1923) was based solely on nondescript tadpoles, and thus this locality should not be included in the species distribution.

Khan’s (2008) inclusion of M. robusta on a checklist for Bangladesh (from “NE and N of hills”) herpetofauna is likely erroneous based on geographical distance from verified populations. Furthermore, within the political borders of Bangladesh the maximum elevation barely exceeds 1000 m asl., less than the lowest recorded elevation verified for this species. For the distribution of M. robusta, Sen and Mathew (2008) listed, aside from Arunachal Pradesh state, the Northeast Indian states of Assam, Meghalaya and Nagaland, and the countries of Bangladesh, China (and specifically Hong Kong), Myanmar and Vietnam without accompanying justification, hence these uncorroborated localities are at best considered erroneous.

Figures

FIGURE 2. BI concatenated gene tree based on the 47 taxa concatenated mt+nuDNA alignment (Dataset C), with ML node values provided. Posterior probability/bootstrap values>95% in both analyses are represented by a black spot on the node, values <95% in one or both analyses are provided next to relevant nodes, where * is>95%, and – is given for nodes that were not resolved on the ML tree. Taxon names in pink represent the eastern clade, blue represents the western clade (southern subclade), and green represents the western clade (northern subclade). Revised names for taxa based on the taxonomic review are provided in parentheses.

FIGURE 3. Concatenated gene trees based on the 29 taxa concatenated nuDNA alignment (Dataset D): A. RAxML tree; B. MrBayes tree. Posterior probability/bootstrap values are provided next to associated nodes. Taxon names in pink represent the eastern clade, blue represents the western clade (southern subclade), and green represents the western clade (northern subclade). Refer to Appendix I, Table 2 for revised names for taxa based on the taxonomic review.

FIGURE 4. RAxML tree for the combined mt+nuDNA dataset expanded to include the Chen et al. (2017) sequences (Dataset J). Bootstrap support values are provided adjacent to the associated nodes. Refer to Appendix I, Table 2 for revised names for taxa based on the taxonomic review.

FIGURE 5. DensiTree cloudogram representations of *BEAST analyses, removing the first 10% of trees as burn-in, produced from the four nuDNA gene alignments of Dataset E: A. original unphased; B. phased. Dark blue branches represent the most frequently occurring topology, light blue branches represent the consensus trees with intensity proportional to the frequency of the topology in the dataset, and green areas represent all topologies in the dataset. Refer to Appendix I, Table 2 for revised names for taxa based on the taxonomic review.

FIGURE 6. Megophrys monticola lectotype: adult female (BMNH 1947.2.25.13) in preservation: A. dorsal view; B. ventral view; C. profile view of head; D. ventral view of hand; E. ventral view of foot.

FIGURE 8. Distribution map of the western MMSG taxa: A. All taxa (excluding M. monticola) from Northeast India and surrounding countries: diamonds represent type localities, circles represent referred localities based on molecularly sampled specimens: orange = M. zhangi; yellow = M. robusta; black = M. medogensis; dark blue = M. major; pink = Megophrys flavipunctata sp. nov.; dark green = Megophrys oreocrypta sp. nov.; light blue = Megophrys himalayana sp. nov.; light green = Megophrys periosa sp. nov.; pale blue = M. cf. periosa; purple = M. glandulosa. B. Inset rectangle in Figure A enlarged to show the distribution map for M. monticola from Darjeeling and Sikkim region, Northeast India, showing distribution of named mitochondrial haplotypes: stars represent the type localities, circles represent referred localities confirmed from sequence data: red = M. “sanu” comb. nov., light blue = M. “katabhako” comb. nov. Blue lines represent major rivers; grey lines represent international borders.

FIGURE 9. Habitat images for collection localities of MMSG species in Northeast India. A. Sessa River, near Sessa village (1110 m asl.), West Kameng dist., Arunachal Pradesh, collection locality of Megophrys robusta and Megophrys periosa sp. nov.; B. and D. Mawphlang Sacred Grove (~1810 m asl.), near Mawphlang village, East Khasi Hills dist., Meghalaya, type locality of Megophrys flavipunctata sp. nov.; C. Senchal Wildlife Sanctuary (2220 m asl.), Rambi (East Range), Darjeeling Sadar subdiv., Darjeeling dist., West Bengal, collection locality of M. monticola; E. Sessa River near Elephant village (370 m asl.), West Kameng dist., Arunachal Pradesh, off which smaller side streams were the type locality of Megophrys himalayana sp. nov.; F. stream near Pangin town (450 m asl.), East Siang dist., Arunachal Pradesh, type locality of Megophrys periosa sp. nov.

FIGURE 11. Megophrys robusta lectotype: adult female (BMNH 1947.2.25.19: SVL 102.0 mm) in preservation: A. dorsal view; B. ventral view; C. profile view of head; D. ventral view of foot; E. ventral view of hand.

FIGURE 12. Megophrys robusta: A–F. referred specimens from West Bengal state and Arunachal Pradesh state: A. adult male topotype (SDBDU 2011.1062: SVL 79.8 mm) in life; B. adult male (SDBDU 2009.1284: SVL 73.5 mm) from Arunachal Pradesh state, in life; C. adult female topotype (SDBDU 2009.1063: SVL 81.3 mm) in life; D. juvenile (SDBDU 2009.1245: SVL 29.2 mm) from Arunachal Pradesh state, in life; E. ventral view of topotype males, adult left (SDBDU 2011.416: SVL 83.1 mm), subadult right (SDBDU 2011.415: SVL 69.1 mm) in preservation; F. ventral view of a an adult pair of topotype specimens taken immediately after euthanisation, left male (SDBDU 2011.1064: SVL 79.3 mm) and right female (SDBDU 2011.1063: SVL 81.3 mm).