Pseudophilautus stellatus

Mendis Wickramasinghe, L. J., Vidanapathirana, Dulan Ranga, Airyarathne, Sameera, Rajeev, Gehan, Chanaka, Amila, Pastorini, Jennifer, Chathuranga, Gayan & Wickramasinghe, Nethu, 2013, Lost and found: One of the world's most elusive amphibians, Pseudophilautus stellatus (Kelaart 1853) rediscovered, Zootaxa 3620 (1), pp. 112-128 : 115-122

Pseudophilautus stellatus

Polypedates stellata Kelaart, 1853

Philautus variabilis (Günther 1859; Kirtisinghe 1957)

Philautus stellatus (Bossuyt & Dubois 2001; Manamendra-Arachchi & Pethiyagoda 2005, 2006) Pseudophilautus stellatus (Yu et al. 2010)

Neotype. NMSL 2013.09.0 1 NH Adult male 55.3 mm SVL ( Figure 2 & 3 ). Sripada World Heritage Site, (Peak Wilderness), Ratnapura District, Sabaragamuwa Province, Sri Lanka (06° 48' 30.89" N 080° 29' 19.18" E). Alt. 1679 m ( Figure 1 ). Collected by L. J. Mendis Wickramasinghe, Dulan Ranga Vidanapathirana & Sameera Ariyarathne on November 22, 2009.

Others. DWC 2013.01.14, adult female, 48.9 mm SVL; DWC 3013.01.15, adult male, 39.6 mm SVL. Date, locality and collectors same as neotype.

Diagnosis. Pseudophilautus stellatus is assigned to the genus Pseudophilautus as it was well nested within this taxon in our molecular phylogenetic tree ( Figure 4 ). Pseudophilautus stellatus can be distinguished from known congeners by the following combination of characters: Body large size (SVL 55.3); snout rounded in lateral, dorsal and ventral aspects; lingual papilla absent; vomerine teeth present; 3rd and 4th fingers with bifid distal subarticular tubercles; tympanum indistinct; supratympanic fold absent.

Description of neotype. Body large size (SVL 55.3 mm); head large (HL/SVL 0.4), about as wide as long (HW/HL 1.1), concave above; snout rounded in lateral, dorsal and ventral aspects (ES/DFE 0.8, SN/IN 0.7), its length longer than horizontal diameter of eye (ES/ED 1.5); internasal space concave; canthus rostralis rounded, loreal region concave; interorbital space concave, upper eyelid smaller than interorbital distance (IO/UEW 1.7); internasal distance equal to upper eyelid width (IN/UEW 1.1); distance between front of eyes 2/3 the distance between back of eyes (DBE/DFE 1.5); nostrils vertically elliptical without flap of skin laterally, closer to tip of snout; pupil horizontally elliptical; tympanum indistinct; pineal ocellus and vomerine teeth present, small (Left=4, Right=6), odontophores oblique and widely separated, between choanae with an angle of 50 0 relative to body axis; tongue large, lanceolate, lingual papilla absent, and conical tubercles absent on tongue.

Arm short, robust and strong (LAL/FEL 0.5, UAL/FEL 0.4); forearm shorter than hand length (LAL/HNL 0.7), longer than upper arm (LAL/UAL 1.3); fore arm distinctly enlarged; fingers robust and strong, relative length of fingers I <II <IV <III (FL-1/FL-3 0.5, FL-2/FL-3 0.6, FL-4/FL-3 0.9) ( Table 1 ); tips of finger disks semi circular, enlarged, discs present on all fingers, with distinct basal and circum marginal grooves; dermal fringe present on inside of all fingers, prominent dermal fringe present on exterior edge of finger IV and hand, and ulnar fold; rudimentary webbing present on all fingers, webbing formula I2–2II2 +–3+ III2–2 IV; distal subarticular tubercles prominent, fingers I and II with rounded distal subarticular tubercles, but III and IV fingers with bifid distal subarticular tubercles, comparatively larger distal subarticular tubercle on finger IV; inner palmar tubercle, large, single, oval, prominent, larger than the distal subarticular tubercles; outer palmar tubercle present, but indistinct; supernumerary tubercles present, but indistinct and widespread on palm and on all fingers; prepollex absent ( Figure 5 A).

Femur 2 1/4 times longer than fourth toe length (FEL/TL-4 2.2); foot length longer than thigh (FOL/FEL 1.4); toes strong, relative length of toes I <II <III <V <IV (TL-1/TL-4 0.4, TL-2/TL-4 0.5, TL-3/TL-4 0.8 (0.78), TL-5/ TL-4 0.8 (0.83), tips of toes rounded, enlarged, discs present on all toes with distinct basal and circum marginal grooves; webbing formula I2 +–2- II1 +–2- III1–2IV1 -–1+V; lateral dermal fringe on inside of all toes and prominent undulating fringe on postaxial edge of toe V, metatarsal fold, and tarsal fold present but indistinct; tarsal fringe present; distal subarticular tubercles prominent, rounded and single, all equal in size; penultimate subarticular tubercles present on toes III, IV and V; supernumerary tubercles present, but indistinct, wide spread and concentrated on foot and all toes; inner metatarsal tubercle oval prominent and large, its length 1/2 the length of toe I (IML/TL-1 0.5); outer metatarsal tubercle absent ( Figure 5 B).

Skin on dorsal and lateral snout, head and entire dorsum weakly shagreened; upper flank shagreened to weakly areolate; lower flank weakly areolate to granular; supratympanic fold absent; upper arm, forearm, and hand weakly shagreened; inner thigh dorsally and outer thigh weakly shagreened; leg, tarsus and foot weakly shagreened; a single prominent large blunt tubercle on heel.

Skin on ventral side of body: Throat and chest weakly granular; belly granular ( Figure 6 A); upper arm weakly granular, forearm granular; thigh granular, leg smooth, tarsus weakly granular.

Colour in life. Body colour is bright green with intermittent pinkish white spots; dorsal part of head and dorsum prominent pinkish white spots outlined in dark brown on bright green, intermittent with smaller blurred dark brown blotches ( Figure 2–3 & 6 B); flank with transverse dark brown bands on white ( Figure 6 C); loreal and tympanic regions and tympanum small brown blotching on bright green; forelimb prominent pinkish white spots outlined in dark brown on bright green, as in dorsum, absent on upper arm; hind limb prominent pinkish white spots outlined in dark brown on bright green ( Figure 6 D), posterior part of femur barred brown; throat, vocal sacs, chest and belly all pinkish white.

Colour in alcohol. Colour of spots and strips faded a little from above, but the white spots remain, and the overall green colour changes to a purplish brown.

The following combination of characters does not match any known Pseudophilautus species described from the island to date. A fully grown female has the described size of 57 mm (=2 ¼’), but an adult male is comparatively smaller with an SVL of 51 mm (= 2’), very broad fingers, large discs, body colours and patterns.

Molecular analysis. The aligned nucleotide sequences span a total of 938 base positions (bp). The analysed dataset consists of parts of the 12S (380 bp) and 16S (558 bp) ribosomal RNA genes. Pseudophilautus stellatus could not be sequenced for the last 32 bp of the aligned 12S fragment. The new mtDNA sequences generated for P. stellatus have been deposited in GenBank (Accession Nos. JN862535 for 12S and JN862536 for 16S). The aligned sequences yielded 490 constant and 379 parsimony-informative characters. The alignment included a total of 69 indels (37 for 12S and 32 for 16S). However, gaps were treated as missing data in MP and NJ analyses.

The maximum parsimony (MP) heuristic search with all characters weighted equally resulted in six trees, 1784 steps in length with consistency indices of 0.373 and retention indices of 0.660. The distance matrices constructed using Kimura 2-parameter corrections, and subsequently analysed by neighbor-joining methods, reconstructed the tree shown in.

In maximum parsimony and neighbor-joining (NJ) analyses, phylogenetic relationships among clades and their bootstrap (BP) supports are very similar. Arrangements among the 3 clades are strongly supported by BP analyses (100%) using either MP or NJ searches. Clades containing individuals of one species have 93-100% BP support ( Figure 4 ).

In MP and NJ analyses, P. stellatus groups within the genus Pseudophilautus . The three species P. f e m o r a l i s, P. poppiae and P. m oo re or u m form a subclade with strong BP support (99% NJ/98% MP). Another well-supported (98%/94%) subclade includes P. frankenbergi and an unknown species. P. stellatus is falling between those two subclades ( Figure 4 ). The sister-group relationship of P. stellatus with the femoralis - poppiae - mooreorum subclade is only weakly supported with BP values of 50 and 55%. However, the grouping of all 6 Pseudophilautus species gets 91% BP support in NJ and 68% in MP analyses. Regardless of the branching order, P. stellatus is clearly different from the other five Pseudophilautus species in this clade, as the long branch in the phylogram nicely illustrates ( Figure 4 ).

Natural history. Typical habitat: These, unusually striking, nocturnal and slow moving amphibians were observed in an area of about 2 km 2 in the Peak Wilderness of the Central Highlands of Sri Lanka ( Figure 1 ). They were commonly found in the canopy of the cloud forest, 1-10 m above the forest floor, perched on large sized leaves well camouflaged. The species was observed on misty/foggy days, and were found to be highly seasonal, and very rare. On subsequent visits, 78 individuals were observed within an area which extends to the Ratnapura District at elevations of 1540 m asl by the team within a period of four months, with no overlapping transects.

Threats. Amongst the most important threats noted in this region, is the forest dieback phenomena ( Figure 7 A&B), possibly due to pollution and/or climate change, which has never been documented in this region before. With decrease in the canopy cover, alien invasive species, such as Clusia rosea , and Pteribium revolutum, are widely distributed in lower areas and is slowly spreading to higher elevations, which can potentially become a threat in the future. The Peak Wilderness is now under severe anthropogenic pressure, because of its religious importance, and is visited every year by a large number of pilgrims who pollute the environment there, especially the streams. As a result, a large amount of garbage gets collected, and the natural forest gets overexploited. Illegal gem mining on either sides of the riverbank within the forest has become another potential threat to the amphibian diversity. Tea plantations in the surrounding areas are slowly expanding and illegal tree felling to cultivate tea, has become a major threat in the area.

Figures

FIGURE 1. Current distribution map of Pseudophilautus stellatus (blue circle denotes the location of neotype).

FIGURE 2. Dorsolateral view of neotype.

FIGURE 3. Front view of neotype.

FIGURE 4. Neighbor-joining tree based on sequences of part of the mitochondrial 12 S and 16 S ribosomal genes with bootstrap values obtained from neighbor-joining (higher nodes) and maximum parsimony analysis (lower nodes).

FIGURE 5. (A – B). Ventral side of Pseudophilautus stellatus neotype: A. left hand with large discs of a pinkish white colouration; B. left foot with large discs.

FIGURE 6. (A – D). Pseudophilautus stellatus, neotype: A. ventral side, belly area; B. dorsal skin colouration, showing bright green with irregular white spots; C. flank, showing transverse darker bands; D. limbs, barred brown and spotted white.

FIGURE 7. Forest die back in the Peak Wilderness area, habitat of Pseudophilautus stellatus.

Tables

TABLE 1. Morphometric measurements of neotype and other specimens of Pseudophilautus stellatus.

Characters	Neotype male NMSL 2013.09.0 1 NH	Female DWC 2013.01.14	Male DWC 2013.01.15	Mean	SD	Range
SVL	55.3	48.9	39.6	48.0	7.9	39.6—55.3
DB	2.2	1.6	1.4	1.7	0.4	1.4—2.2
DBE	18.9	18.3	14.7	17.3	2.3	14.7—18.9
DFE	12.0	11.6	8.9	10.8	1.7	8.9—12.0
DL	2.7	2.2	1.9	2.3	0.4	1.9—2.7
DW	3.9	3.4	3.0	3.4	0.4	3.0—3.9
ED	6.1	5.9	4.8	5.6	0.7	4.8—6.1
EN	5.5	5.4	4.1	5.0	0.8	4.1—5.5
ES	9.0	8.8	6.7	8.2	1.3	6.7—9.0
FEL FL-1	27.6 5.2	27.8 4.9	21.0 3.6	25.5 4.6	3.9 0.9	21.0—27.8 3.6—5.2
FL-2 FL-3	6.2 10.4	6.1 9.5	4.2 8.4	5.5 9.4	1.1 1.0	4.2—6.2 8.4—10.4
FL-4	9.3	8.8	7.2	8.4	1.1	7.2—9.3
FOL	38.1	38.4	29.9	35.4	4.8	29.9—38.4
GK	23.5	23.0	15.9	20.8	4.2	15.9—23.5
HD	10.2	9.8	7.4	9.2	1.5	7.4—10.2
HL	22.0	21.4	15.9	19.8	3.3	15.9—22.0
HW IML	24.0 2.5	23.3 2.3	13.4 1.3	20.2 2.0	6.0 0.6	13.4—24.0 1.3—2.5
IN IO	5.1 7.6	4.3 6.9	3.9 5.6	4.4 6.7	0.6 1.1	3.9—5.1 5.6—7.6
KT	23.0	24.2	18.5	21.9	3.0	18.5—24.2
LAL	12.3	12.0	9.8	11.4	1.4	9.8—12.3
MBE	9.6	8.1	6.9	8.2	1.4	6.9—9.6
MFE	15.9	14.6	11.2	13.9	2.4	11.2—15.9
MN	19.9	18.8	14.0	17.6	3.1	14.0—19.9
HNL	16.7	15.6	12.1	14.8	2.4	12.1—16.7
SN	3.5	3.2	2.9	3.2	0.3	2.9—3.5
TAS	16.9	16.0	12.3	15.1	2.5	12.3—16.9
TBL	27.2	27.4	21.1	25.2	3.6	21.1—27.4
TL-1	5.0	4.8	3.5	4.4	0.8	3.5—5.0
TL-2	5.9	6.0	4.7	5.5	0.7	4.7—6.0
TL-3	9.6	9.5	7.2	8.8	1.3	7.2—9.6
TL-4	12.4	12.9	11.7	12.3	0.6	11.7—12.9
TL-5	10.3	9.6	8.4	9.4	1.0	8.4—10.3
UAL	9.7	8.4	6.6	8.3	1.6	6.6—9.7
UEW	4.5	4.6	3.5	4.2	0.6	3.5—4.6
VKL	22.6	23.3	18.5	21.4	2.6	18.5—23.3