Leaping towards a saltatorial lifestyle? An unusually long-legged new species of Rhombophryne (Anura, Microhylidae) from the Sorata massif in northern Madagascar Author Scherz, Mark D. Author Rakotoarison, Andolalao Author Hawlitschek, Oliver Author Vences, Miguel Author Glaw, Frank text Zoosystematics and Evolution 2015 91 2 105 114 http://dx.doi.org/10.3897/zse.91.4979 journal article http://dx.doi.org/10.3897/zse.91.4979 1860-0743-2-105 79F1E7CE3ED34A6CAA15DC59AB3046B8 Taxon classification Animalia Anura Microhylidae Rhombophryne longicrus sp. n. Figs 3, 4 Holotype. ZSM 1630/2012 (FGZC 3653), an adult female with immature oocytes, collected in the montane forest of the Sorata Massif, north Madagascar (ca. 13.675°S , ca. 49.4392°E , ca. 1580 m; datum = WGS84) on 28 November 2012 by A. Rakotoarison, A. Razafimanantsoa, T. Rajoafiarison, F. M. Ratsoavina, O. Hawlitschek and F. Glaw. Paratype. UADBA-A 60271 (FGZC 3651), an adult male with the same collection data as the holotype. Diagnosis. A microhylid assigned to the genus Rhombophryne on the basis of overall morphology, including the possession of maxillary and vomerine teeth, absence of expanded toe pads, and absence of an enlarged prepollex. Confirmed as a member of the genus Rhombophryne on the basis of its phylogenetic relationships as assessed by mitochondrial DNA, as there are no known morphological characters by which Rhombophryne may be distinguished from Plethodontohyla . Rhombophryne longicrus sp. n. is distinguished from all other Madagascan frog species by the following set of characters: SVL 23.8-27.9 mm, head wider than long, horizontal tympanum diameter 47% of eye diameter, absence of superciliary spines, weak supratympanic fold, dark supratympanic region and nostril, tibiotarsal articulation reaching the nostril, total hindlimb length 183-185% of SVL, second finger shorter than fourth, and fifth toe shorter than third. It is also separated by a pairwise genetic distance of at least 6.8% in the 16S mitochondrial gene from all other known species of the genus Rhombophryne . Within the genus, Rhombophryne longicrus sp. n. may be distinguished from all Rhombophryne species, except Rhombophryne laevipes , Rhombophryne minuta , and Rhombophryne vaventy , by tibiotarsal articulation reaching the nostril (versus not exceeding the eye); from Rhombophryne coronata , Rhombophryne ornata , Rhombophryne serratopalpebrosa , Rhombophryne tany , and Rhombophryne vaventy by the absence of superciliary spines (versus presence); from Rhombophryne alluaudi , Rhombophryne laevipes , Rhombophryne matavy , Rhombophryne testudo , and Rhombophryne vaventy by its smaller size (SVL 23.8-27.9 mm versus 32-53 mm); from Rhombophryne minuta by its larger size (SVL 23.8-27.9 mm vs. up to 17.1 mm); from Rhombophryne testudo by the absence of barbels on the throat and tympanum smaller than eye; from Rhombophryne alluaudi , Rhombophryne coronata , Rhombophryne serratopalpebrosa , Rhombophryne tany , and Rhombophryne vaventy by its weak, almost absent supratympanic fold; from Rhombophryne coudreaui and Rhombophryne vaventy by smooth dorsal skin (versus granular/tubercular); from Rhombophryne mangabensis by lack of paired dark dorsal tubercles; from Rhombophryne laevipes , Rhombophryne mangabensis , Rhombophryne ornata , Rhombophryne testudo , and Rhombophryne vaventy by absence of dark crossbands on hindlimbs; and from Rhombophryne coronata , Rhombophryne minuta , Rhombophryne testudo , and Rhombophryne vaventy by dark supratympanic region. Osteologically, a micro-CT scanned specimen of Rhombophryne longicrus sp. n. tentatively differs from Rhombophryne ornata (3 specimens: ZSM 1815/2010, 1816/2010, and 2859/2010), Rhombophryne serratopalpebrosa (1 specimen: MNHN 1975.24), Rhombophryne tany (1 specimen: ZSM 1814/2010), and Rhombophryne vaventy (1 specimen: ZSM 375/2005) (as described in Scherz et al. 2014 , 2015 ) by its relatively larger nasals (nasal length at longest point 18.5% of skull length versus 11.1-16.4%), which are in contact with the sphenethmoid (versus not contacting any other bones), its relatively longer and less broad skull (skull length 81.6% of skull width versus 66.5-79.4%), and its relatively longer brain case (frontoparietals+sphenethmoid length 74.0% of skull length versus 63.3-71.5%; length of frontoparietals+sphenethmoid 197.7% of width of frontoparietals anterior to prootic versus 173.4-185.6%). A thorough osteological treatment of this genus is needed to confirm further differences and their values. Rhombophryne species can be confused with Plethodontohyla species. Rhombophryne longicrus sp. n. differs from them in the following ways: absence of a sharp dorsolateral colour border and expanded finger and toe pads (versus presence in Plethodontohyla notosticta , Plethodontohyla guentheri , Plethodontohyla mihanika , and Plethodontohyla inguinalis ), absence of inguinal spots (versus presence in Plethodontohyla mihanika , Plethodontohyla inguinalis , Plethodontohyla ocellata , and Plethodontohyla bipunctata ), tibiotarsal articulation reaching the nostril (maximally reaching to the mid-eye in all Plethodontohyla except Plethodontohyla mihanika ), absence of crossbands on legs (versus presence in Plethodontohyla fonetana , Plethodontohyla inguinalis , Plethodontohyla notosticta , Plethodontohyla guentheri , and Plethodontohyla mihanika ), and smooth skin (versus granular to rough in Plethodontohyla tuberata ). Description of the holotype. Adult female in an excellent state of preservation. A ventral incision was made in order to check the sex and access the stomach contents. The incision runs laterally and posteroventrally anterior to the pubis and up the middle of the venter. Body gracile; dorsal and ventral skin smooth. Head wider than long (HW 122.5% of HL), snout rounded in dorsal view, squarish in lateral view; nostrils weakly protuberant, directed laterally, equidistant between eye and snout; canthus rostralis concave; loreal region concave; tympanum indistinct, oval, horizontally 47% of eye diameter; pupil round; supratympanic fold weak, almost absent ; tongue unlobed, posteriorly free; vomerine teeth present in a straight row with a small medial gap (<1 mm; see Osteology below); choanae small, oval. Arms slender and long; fingers without webbing, long, without distinct subarticular tubercles, relative lengths 1<2<4<3, second finger much shorter than fourth, without enlarged terminal discs; inner metacarpal tubercle present; nuptial pads absent. Legs exceptionally long and slender (HIL 185% of SVL), tibiotarsal articulation reaching the nostril when hindlimb is adpressed along body; toes long, unwebbed, with indistinct subarticular tubercles, relative toe lengths 1<2<5<3<4, third toe much longer than fifth; inner metatarsal tubercle present and indistinct. Colouration of the holotype: (Fig. 3a, c). In life, snout anterior to eyes, above eyes, side of head, and upper arms bronze to tan in colour; tip of snout darker, lightening posteriorly; area around nostril black; supratympanic region dark brown, fading below to the tan of the lateral side of the head. Body laterally light brown, becoming increasingly yellowish brown dorsolaterally until tan border with dark dorsal marking; this marking is flecked with additional tan spots, and extends from a black horizontal bar between the eyes to the legs, where dark ashy grey dominates; border between dorsal and dorsolateral colouration almost symmetrically emarginated. Hands and feet tan with dark flecks. Ventral skin pinkish and slightly translucent; chin dark relative to rest of venter, posteriorly lightening with few darker patches interspersed with translucent areas lacking pigment. Anteroventral surface of legs with dark pigment, becoming less pigmented more ventrally; ventral surface mottled pinkish and light brown. Figure 3. Rhombophryne longicrus sp. n. in life. Holotype ZSM 1630/2012 in (a) dorsolateral and (c) ventral view. Paratype UADBA-A 60271 in (b) dorsolateral and (d) ventral view. After three years in 70% ethanol, all browns have faded to shades of grey. Dorsal areas that were lightest in shade are whitish, particularly between the eyes anterior to the dark inter-ocular bar. Ventrally, all areas that lack pigmentation and were pink in life are cream in preservation. Chin the same colour as the snout. Osteology of the holotype: (Fig. 4, Suppl. material 1). All bones of the skull paired except the parasphenoid and sphenethmoid. Vomer divided into pre- and postchoanal portions; prechoanal part small, longer than broad, subtriangular; postchoanal part overlapping neopalatine, bearing ventral serrations (vomerine teeth), separated medially from its counterpart by a gap of 0.7 mm. Postchoanal vomer+neopalatine in dorsal contact with anterior end of cultriform process of parasphenoid, and through it with the sphenethmoid; laterally not in contact with maxilla. Teeth present on maxilla and premaxilla. Premaxilla medially not fused to counterpart, anterodorsal alary processes rising dorsolaterally, pars palatina with two well-defined processes, the medial (palatine) process thin, lateral process broad; pars dentalis bearing small teeth. Septomaxilla roughly spiralling upward from posterior ramus to lateral ramus to anterior ramus to medial ramus. Nasal medially in contact with the sphenethmoid posteriorly, possessing a pointed maxillary process extending ventrolaterally toward the maxilla, lacking an anterolateral ramus. Maxilla long, bearing small, poorly resolved teeth, possessing a horizontal pars palatina along its lingual margin; in broad lateral contact with anterior ramus of pterygoid; posteriorly without clear distinction from quadratojugal. Pterygoid broad and triradiate, with anterior, medial, and posterior rami; the ventrolateral edge of its anterior ramus, posterior margin of its medial ramus, and lateral face of its posterior ramus sculpted inward; medial ramus much shorter than posterior ramus; posterior ramus not in contact with quadratojugal. Quadratojugal L-shaped, anterior process without clear distinction from posterior of maxilla; posteriorly possessing a ventral bulbous process with a concave posterior face; dorsally without clear distinction from squamosal. Squamosal thin and distally bifurcated, extending anterodorsomedially from quadratojugal to level of otic capsule passing anterior to columella; otic ramus longer than zygomatic ramus. Columella with a long shaft that exceeds the level of the squamosal; dorsal edge of columella straight even to end of footplate; columellar footplate broad and concave. Frontoparietal medial and lateral edges straight and parallel, lateral edge curved ventrally to form dorsolateral border of brain case; possessing paired bumps at the transverse level of the columellae; posterolateral sutures with prootics and exoccipitals not clear from micro-CT scans; anterior process contacting sphenethmoid. Parasphenoid T-shaped; cultriform process broadening anteriorly, contacting sphenethmoid at its anterior end; broad posterior alary processes perpendicular to cultriform process, in dorsal contact with prootics anteriorly and exoccipitals posteriorly. Figure 4. Osteology of the holotype of Rhombophryne longicrus , ZSM 1630/2012. Skull in (a) lateral, (b) dorsal, and (c) ventral view. Skeleton in (d) dorsal and (e) ventral view. Note: figures display only calcified structures; cartilages are omitted due to limitations of micro-CT scanning. Abbreviations: angspl = angulosplenial, col = columella, exoc = exoccipital, fpar = frontoparietal, max = maxillary, mmk = mentomeckelian, pmax = premaxilla, povom = postchoanal vomer+neopalatine, proot = prootic, prsph = parasphenoid, prvom = prechoanal vomer, pter = pterygoid, qj = quadratojugal, smax = septomaxilla, spheth = sphenethmoid, sq = squamosal. Mandible slim, edentate. Mentomeckelian small, in narrow medial contact with counterpart (possibly artefactual), and in dorsolateral contact with dentary. Dentary long and thin with a sculpted outer face and smooth inner face, overlapping angulosplenial for much of its length. Angulosplenial broadening posteriorly, with a posterior dorsomedial crista; possessing a lateral channel running from the posterior into the sculpted outer edge of the dentary. Posterolateral processes of hyoid shovel-like, a medial crista running along posteromedial process, the base of which is broad and flat with a rounded anteromedial edge and sharp anterolateral and posteromedial corners; parahyoid absent. Humerus long, slim and straight; crista lateralis weak, crista ventralis short (~30% of humerus length), crista medialis absent. Radioulna broadening distally. Finger phalangeal formula 2-2-3-3. Terminal phalanges of fingers 2, 3, and 4 with distal knobs. Prepollex 31% of first finger. Pectoral girdle composed of paired coracoids, clavicles, scapulae, cleithra and suprascapulae. Sternal characters not visible in CT render. Coracoids in medial contact; medially dorsoventrally flattened, laterally rounded, posterior surface straight, anterior surface strongly concave. Clavicle thin and curved approximately parallel to the anterior edge of the coracoid, its lateral end broadened, posteriorly in contact with ventral edge of scapular pars acromialis. Scapula thick, hourglass shaped, its posterior edge less strongly curved than its anterior edge, medioventrally bifurcated; pars acromialis distally rounded, in contact with the lateral end of the clavicle, its anterior surface concave; pars glenoidalis curved ventrally, in contact with lateral face of coracoid, posterior face concave; dorsal edge of scapula approaching cleithrum. Cleithrum thin and long, not possessing any cristae, anteriorly thicker than posteriorly. Suprascapula with highest X-ray absorption ventrally and posteriorly suggesting possible ossification in these areas. Toe phalangeal formula 2-2-3-4-3; terminal phalanges without distinct distal knobs. Leg bones long and thin. Femur without any crests. Tibiofibula slightly longer than femur. Tibiale and fibulare proximally and distally fused, articulating distally with metatarsals V and IV, tarsals 1-3, and the centrale. Prehallux present, short. Ilium, ischium, and pubis forming ossified acetabulum, each composed of paired, medially fused elements. Iliac shafts oval in cross-section, dorsal-ventral diameter larger, possessing a weak dorsal tubercle posterior to shallow oblique groove. Iliosacral articulation type IIA sensu Emerson (1979) . Eight presacrals present; no vertebrae fused. Posterior articular processes round. Transverse processes of presacrals II-IV broader than those of V-VIII . Neural spines decreasing in size from presacral II to absent by V. Sacrum wide, with broad diapophyses articulating with the ilia; anterior edge of each diapophysis roughly perpendicular to body axis, posterior edge oblique. Urostyle long and thin, with a dorsal ridge along a third of its length, beginning at its anterior end; articulation with sacrum bicondylar. Measurements. Holotype (paratype in brackets), measurements in mm: SVL 28.0 (23.8), HW 9.9 (9.7), HL 8.0 (7.0), ED 3.3 (2.8), END 2.0 (2.0), NSD 1.9 (1.7), NND 3.0 (2.2), TDH 1.5 (1.3), TDV 1.8 (1.4), HAL 8.4 (7.0), UAL 5.7 (4.7), LAL 6.8 (5.7), FORL 20.9 (17.4), THIL 13.2 (11.7), THIW 3.9 (3.4), TIBL 14.6 (11.7), TIBW 2.97 (2.64) TARL 8.6 (7.3), FOL 14.8 (12.5), FOTL 23.4 (19.8), HIL 51.2 (43.2), IMCL 1.0 (0.9), IMTL 1.3 (1.0). Variation. Only two specimens are known. The paratype is male, and smaller than the holotype (SVL 23.8 mm). It agrees in all aspects of its morphology with the holotype, but differs strongly in colouration (see Fig. 3b, d). In life, the dorsum has a yellow-brown base colour, with distributed black or dark brown flecks. A black inter-ocular bar is present, behind which the skin fades from brown to the base colour; the back does not possess the dark marking of the holotype, but instead two darker areas with a few black flecks lie in the suprascapular region. The lateral skin fades to grey ventrally, also speckled with black. A dark line runs from the preocular region to the axial pit through the supraocular and supratympanic regions. The nostril is surrounded by black, and the tympanum has a dark fleck on it. The legs are grey at the hip, but this lightens to the yellowish brown of the dorsum further away; the arms are dorsally yellow, the hands possessing a few black spots. Ventrally, the pink areas of the holotype are orange in life in the paratype, particularly over the pectoral girdle and beneath the chin. The venter is marked with many irregular black flecks. The arms are ventrally orange, bordered posteriorly in black. Etymology. The species epithet is an invariable noun in apposition to the genus name, derived from the Latin words longus (meaning long), and crus (meaning leg), and refers to the unusually long legs of this species. Distribution. This species has only been found at high altitude in the montane forests of the Sorata massif in north Madagascar. Its distinctiveness leads us to hypothesize that it has never been found elsewhere and misidentified, so it may be microendemic to this small area. Additional surveys are required in areas in and around Sorata to identify its full distribution. Ecology. Both specimens described here were captured in the early evening on the ground along a path through primary montane forest. The stomachs of both specimens contained remains of several small insects (mostly Coleoptera ) and a spider (possibly belonging to the family Salticidae ), mixed with moss, suggesting an opportunistic diet of arthropods. Calls of this species are unknown. The female holotype had more than twenty immature oocytes with the largest having diameters ranging from 1.3 to 1.6 mm. As a member of the Cophylinae , it is likely that Rhombophryne longicrus lays its eggs away from running water or large water bodies, and has endotrophic tadpoles. Conservation status. The forests of Sorata are currently unprotected. All locally endemic species are threatened by uninhibited deforestation and forest degradation. The greatest pressure on forests is at their edges. High altitude species like Rhombophryne longicrus may therefore be the least threatened by this. However, a sustained rate of deforestation will increase the threat level to species at ever-higher altitudes. It is conceivable that a restriction of this species to high altitudes may mean that it is susceptible to climate change ( Raxworthy 2008 , Raxworthy et al. 2008 ). We consider this threat far less serious than that of deforestation. Batrachochytrium dendrobatidis has now been confirmed from numerous localities in Madagascar ( Bletz et al. 2015 ). So far no negative impacts on native frogs have been observed. The water-independent lifestyle of Rhombophryne species suggests that they are probably at relatively low threat from chytridiomycosis. While this species is, at present, known from just two specimens collected on one expedition, the fact that it has not been collected by previous expeditions suggests it may be scarce, seasonal, or have a scattered distribution. Even if it were distributed throughout the forests of the Sorata massif, its distribution would still only constitute an area of ~250 km2 (as calculated in Google Earth® Pro 6.1.0.500, Google Inc., Mountain View, CA). Thus, because of its potentially limited range inside an unprotected forest, the on-going and intensifying threat of deforestation, potential threat by climate change, and potential scarcity or seasonality, it qualifies as Endangered B1ab(iii) under the IUCN Red List Criteria (2012).