Phrynobatrachus bibita Goutte, Reyes-Velasco & Boissinot

Goutte, Sandra, Reyes-Velasco, Jacobo & Boissinot, Stephane, 2019, A new species of puddle frog from an unexplored mountain in southwestern Ethiopia (Anura, Phrynobatrachidae, Phrynobatrachus), ZooKeys 824, pp. 53-70 : 53

Phrynobatrachus bibita Goutte, Reyes-Velasco & Boissinot sp. n.

Material.

Holotype. A male (SB440; Figure 2), collected on 17 June 2018, at night, in Bibita Mountain, southwestern Ethiopia (6.8229N, 35.0938E, datum = WGS84; 1972 m a.s.l.). The specimen was collected while in amplexus (female paratopotype SB439), on vegetation, in overgrown pond in primary forest. Paratopotypes. Nine specimens: eight females collected in the same pond as the holotype on 16 June 2018 (SB418-SB420, SB424-SB427) and on 17 June 2018 (SB439); another male (SB421) collected at night in the water (probably calling), at same locality as the holotype on 16 June 2018.

Diagnosis.

Small species (SVL = 16.8 ± 0.1 mm for males, 20.3 ± 0.9 mm for females) attributed to the genus Phrynobatrachus by the presence of tarsal and outer metatarsal tubercles (Suppl. material 2: Figure S1A). Body slender, with long legs (tibia length/SVL = 0.6 in both sexes) rather long snout for the genus and very elongated fingers (hand length/SVL = 0.3 in both sexes) and toes (foot length/SVL = 0.6 in both sexes) in comparison to its congeners. Webbing absent between fingers and minimal between toes. Tympanum not visible. Canthus rostralis marked and concave from nostril to eye. Snout pointed. Nostrils not visible from above. Eyelid spine absent. Throat of adult males white with light grey freckles on the anterior third, without any spinulae. Femoral glands hardly distinguishable but present in adult males. Two ridges in the scapular region and two short, oblique ridges behind the eyes. These four ridges may be all disjointed, the two scapular ridges may be jointed to form a chevron shape, or the ridges may be jointed laterally in an hourglass shape.

Comparisons .

This species can easily be distinguished from other Ethiopian Phrynobatrachus : the body is slenderer, the hind limbs, fingers and toes are longer than all four other described Ethiopian Phrynobatrachus , P. bullans , P. natalensis , P. minutus , and P. inexpectatus . Additionally, the tips of fingers and toes are more enlarged than in any of these species, particularly in females. It is similar in size as P. minutus , slightly larger than P. inexpectatus and much smaller than P. bullans and P. natalensis . It can be further distinguished from P. natalensis by a more marked canthus rostralis and a completely hidden tympanum. Adult males of P. bibita sp. n. can be distinguished from P. bullans , P. natalensis , P. minutus , and P. inexpectatus by the white coloration of their throat ( Largen 2001, Crutsinger et al. 2004). The new species is also distinct from the three junior synonyms of P. natalensis described from Ethiopia: Arthroleptis bottegi Boulenger, 1895, described from Auta, Bale province, possesses a distinct tympanum, a rounded snout, and is much larger than P. bibita (SVL of female holotype = 31 mm vs. 20.3 ± 0.9 mm in female P. bibita ). Phrynobatrachus sciangallarum Scortecci, 1943, described from a presumably adult male from Murle, Gemu-Gofa province, differs from P. bibita by the presence of a considerably darkened throat and the lack of femoral glands. Additionally, it has more extensive foot webbing than P. bibita ( Largen 2001). Arthroleptis-Phrynobatrachus zavattarii Scortecci, 1943, described from Caschei, Gemu-Gofa province, only differs from P. sciangallarum by the extent of the foot webbing, and all measurements of the type specimens fall within the range of P. natalensis ( Largen 2001). Blackburn (2014) provided color photographs of the holotypes of these last two junior synonyms of P. natalensis . Based on those photographs, it appears that P. bibita has a slenderer head and more elongated fingers than these individuals. Finally, the type localities for these three junior synonyms of P. natalensis are in the southern part of the country, an area that is dry and at low elevation, in contrast with the habitat occupied by P. bibita sp. n.

Among Eastern African Phrynobatrachus , P. bibita sp. n. can be distinguished from P. acridoides (Cope, 1867), P. auritus Boulenger, 1900, P. dendrobates (Boulenger, 1919), P. graueri (Nieden, 1911), P. keniensis Barbour & Loveridge, 1928, P. krefftii Boulenger, 1909, P. perpalmatus Boulenger, 1898, P. petropedetoides Ahl, 1924, and P. versicolor Ahl, 1924, by a completely hidden tympanum ( Cope 1867, Boulenger 1882, 1909, Razzetti and Msuya 2002, Pickersgill 2007). This species can further be distinguished from P. auritus Boulenger, 1900, P. dendrobates , P. graueri , P. kinagopensis Angel, 1924, P. krefftii , P. mababiensis FritzSimons, 1932, P. parvulus (Boulenger, 1905), P. rouxi (Nieden,1912), P. scheffleri (Nieden, 1911), and P. ukigensis Grandison & Howell, 1983, by the white and light grey coloration of adult males’ throat ( Boulenger 1909, Angel 1924, FritzSimons 1932). The new species differs from P. bullans , P. kakamikro Schick, Zimkus, Channing, Köhler, & Lötters, 2010, and P. perpalmatus by the presence of femoral glands in adult males ( Crutsinger et al. 2004, Pickersgill 2007, Schick et al. 2010). The new species is also larger than P. pallidus , P. parvulus , P. scheffleri , and P. ungujae Pickersgill, 2007 ( Boulenger 1905, Pickersgill 2007, Schick et al. 2010), and possesses longer legs than P. kakamikro and P. pallidus Pickersgill, 2007, as well as longer feet than P. pallidus and P. ungujae ( Pickersgill 2007, Schick et al. 2010). Finally, P. bibita sp. n. can be distinguished from P. acridoides , P. bullans , P. mababiensis , P. natalensis , P. pallidus , P. ungujae , and P. versicolor Ahl, 1924, by the absence of dark bars on the lower jaw ( Cope 1867, Boulenger 1882, Fritzsimons 1932, Crutsinger et al. 2004, Pickersgill 2007).

Description of the holotype.

Adult male (Figure 2; Table 1), body slender, head longer than wide (HW/HL = 0.96), the snout is pointed and the canthus rostralis is marked and concave from nostril to eye. Tympanum not visible. Pupil horizontal. Eyelid spine absent. Maxillary teeth present, vomerine teeth absent. Tongue elongate, free for two thirds of its length, tip divided in two short lobes. Legs and feet long for the genus (TL/SVL = 0.6; FL/SVL = 0.6). Tarsal tubercle and outer metatarsal tubercle present, but inner metatarsal tubercle not distinguishable. Subarticular tubercles absent on the hands and small and barely distinguishable on the feet. Palmar tubercle absent. Fingers and toes elongated and their tips slightly swollen but not forming discs. First fingers shorter and swollen but nuptial pads absent. Fingers free of webbing. No webbing between toe I and II, and minimal webbing between the other toes. Finger formula: I<II<IV<III. Toe formula: I<II<V<III<IV. Skin of dorsum and dorsal side of limbs covered by small pointy asperities. Femoral glands hardly visible but present (Suppl. material 2: Figure S1B). Chevron-shaped gland in scapular region extending from above the shoulder girdle to approx. mid-body. Two rows of small warts form faint ridges between the eye and the scapular region but do not reach the chevron-shaped gland. The skin of the throat is not thin and loose as in many other Phrynobatrachus species and gular folds and spinulae are absent.

Coloration of holotype in life.

The body is golden (Figure 2A); a dark chevron is present in the scapular region underlining two short back ridges. A faint dark triangle is present between the eyes and a faint dark bar is present between the eye and the shoulder. The upper eyelids are darker than the rest of the head. Loreal region faintly darker than the rest of the head. Upper and lower jaws lack any blotching. The flanks and the sides of the head present numerous small white spots, extending to the upper lip below the eye. Iris dark gold with a golden ring around the iris interrupted by a black dot at the bottom of the iris. The thighs and legs are faintly and irregularly banded. The back of the thighs is yellowish with irregular coverage of melanophores. A dark triangle is present in the vent region. Ventral region is white molted with light grey. The throat is white, with the anterior third light grey with white freckles. Ventral side of the thighs and legs shows irregular melanophores and white spots.

Coloration of holotype in preservative.

After euthanasia, the individual’s coloration darkened notably (Figure 2B). In preservative, the dorsum presents a grey coloration.

Variation.

Morphometric variations of the type series are summarized in Table 1. Inner metatarsal tubercles are not visible in all individuals. Subarticular tubercles are more pronounced in some individuals. Webbing of the feet presents small variation, ranging from minimal to no webbing for toes I and II, and webbed to the first phalange for toes III to V. The dorsal and leg skin is mostly smooth with small warts more pronounced in some individuals. The two ridges in the scapular region and two oblique ridges behind the eyes were visible in all examined specimens. In four specimens, they formed a chevron in the scapular region, in three specimens the ridges jointed laterally, forming an hourglass, and in two specimens all four ridges were disjointed.

Males and females differ in size and in proportions (Figure 2). Finger and toe tips are enlarged in females and fingertips are slightly enlarged in males. Males’ first finger is short and swollen but no distinct nuptial pad is visible. Adult males present femoral glands, which are lacking in females. Bicolor eggs are visible through the skin of gravid females.

Coloration of the body varies from golden to light brown, with few large light green blotches in some individuals. Most specimens present a dark chevron in the scapular region underlining two short back ridges that are either jointed or disjointed with two oblique ridges between the back of the eye and the shoulders. A more or less pronounced dark stripe is present between the nostril and the eye, and between the eye and the arm. A more or less distinctive dark bar is present between the eyes and some individuals present a lighter or green snout. The flanks and the sides of the head present numerous small white spots, extending to the upper lip in some individuals. The thighs and legs of some individuals are very faintly and irregularly banded. The back of the thighs is cream with small light grey spots. Vocal sac in adult males is white with the frontal third of the throat light grey with white freckles, while in females the throat and the ventral skin is light grey or yellowish molted with white. Some individuals have a thin light line on the backside of the thigh from the vent to the tarsal tubercle. A dark triangle is generally present in the back of the thigh, around the vent.

Etymology.

The specific name refers to Bibita Mountain, the type and only known locality for the species. It is an invariable noun used in apposition.

Habitat, distribution, and natural history.

All individuals were collected in a single large overgrown forest pond (Figure 3A), at night. The surrounding forest consisted of large trees with overhanging epiphytes and dense undergrowth. All females and the amplected pair were found on vegetation ca. 30 cm above water (Figure 3B). A single male was found in the water, presumably while calling. All collected females were gravid, and bicolor eggs were visible through the skin. Females seemed to aggregate in specific areas of the pond, were numerous egg clutches were found on leaves overhanging the water (Figure 3B). Laying eggs on vegetation overhanging the water is unusual in Phrynobatrachus , most species laying their eggs directly in the water ( Zimkus et al. 2012). We thus confirmed that these eggs belonged to Phrynobatrachus bibita sp. n. by sequencing their mitochondrial rRNA 16s. Various forms of terrestrial egg deposition have been described in the genus Phrynobatrachus ( Zimkus et al. 2012): most similarly to P. bibita , P. sandersoni (Parker, 1935) lays its eggs on vegetation up to 2 m above small puddles, small streams or water-saturated soil ( Amiet 1981) and P. krefftii lays its eggs above the water, on rocks or vegetation ( Harper and Vonesh 2010). Phrynobatrachus guineensis Guibé & Lamotte, 1961 lays its eggs on the bark of trees above water-filled tree holes ( Rödel 1998) and P. dendrobates lays its eggs in tree holes or above streams ( Zimkus et al. 2012). Finally, P. phyllophilus Rödel & Ernst, 2002, P. tokba (Chabanaud, 1921), and P. villiersi Guibé, 1969 lay their eggs on the leaf litter or the forest floor ( Rödel and Ernst 2002a, 2002b; Zimkus et al. 2012). Phrynobatrachus bibita sp. n. thus adds to the diversity of reproductive modes in the genus.

Five egg clutches were photographed and contained approximately 30 eggs each (range 22 to 33). All observed clutches were found between 20 - 40 cm above water on vegetation, and up to two clutches were found on a single leaf. When laid, the eggs are bicolor, heavily pigmented, and encased in a thin gelatinous layer; as the egg develops, the pigmentation is more evenly distributed at its surface, with dark brown freckle, and the gelatinous layer becomes much thicker. It is possible that female P. bibita sp. n. are guarding their egg clutches in a similar manner as P. sandersoni females, which attend their eggs at night by standing over them or staying in the close proximity ( Amiet 1981). Additional behavioral observations are necessary to determine whether females P. bibita sp. n. truly display such parental care. Other frog species were found in the same pond or nearby, including Afrixalus enseticola Largen, 1974, Leptopelis vannutellii (Boulenger, 1898), Paracassina obscura (Boulenger, 1895), and Ptychadena cf. erlangeri (Ahl, 1924).

The forest in Bibita Mountain appears to be well preserved, as there are no settlements inside the forest, and a footpath is the only way to cross the forest. People from the surrounded villages harvest wild honey from the forest, but this is the only noticeable human activity there. Based on satellite imagery, no human disturbance is apparent at the higher elevations of the mountain; however, most areas below 1900 m have been transformed into agricultural land.

Call.

The advertisement call is composed of a series of pulsed notes with a slight upward frequency modulation within each note (Figure 4A). It is similar to that of Phrynobatrachus minutus (Figure 4B) in spectral structure, note length, and dominant frequency. The vocalization of P. bibita sp. n. can be distinguished from that of P. natalensis (Figure 4C) by its higher dominant frequency, indicative of its smaller body size. Dominant frequency: 3318 ± 94 Hz. Note duration = 630 ± 80 ms. Interval inter-notes = 1085 ± 630 ms. The mediocre quality of our recording prevents us from analyzing the advertisement call of P. bibita sp. n. in further details, and a more complete description of their vocalization will be needed in the future, in particular to distinguish it from other species of dwarf Phrynobatrachus .

Estimates of evolutionary relationships.

The goal of our phylogenetic analysis was to test the phylogenetic position of the new species, not to obtain a phylogeny of the genus, which has been previously done elsewhere ( Zimkus et al. 2010). Our phylogenetic estimates recovered three main lineages in the genus Phrynobatrachus (Figure 5); however, several of the nodes have low support. We briefly describe the resulting topology below.

We recovered a basal group of Phrynobatrachus , which included the species P. acutirostris Nieden, 1912, P. krefftii , P. dendrobates , P. petropedetoides Ahl, 1924, and P. sandersoni . This clade is consistent with the "Clade A" of Zimkus et al. (2010), however this clade received low support (posterior probability (pp) <0.5). All remaining species in the genus were grouped into two major clades, which received strong support (pp = 1). The first one of these two groups is similar to the "Clade B" of Zimkus et al. (2010), which includes the Ethiopian species P. minutus , P. inexpectatus , and P. sp. n. “Oromia” of Zimkus and Schick (2010), while the second group is consistent with the "Clade C" of Zimkus et al. (2010), and includes the Ethiopian species P. natalensis . Phrynobatrachus bibita sp. n. was not closely related to any of the other Ethiopian species, and instead basal to the last group ("Clade C" of Zimkus et al. (2010)), however with high support (pp = 1). Our analysis grouped the egg clutch collected (SB422) with the adults P. bibita sp. n. (holotype SB440 and paratopotypes SB419, SB420, SB425), ensuring that it belonged to the new species. We did not include samples of three junior synonyms of P. natalensis from Ethiopia (see Comparisons ), however, samples referable to these junior synonyms have previously been included on a phylogeny of members of the P. natalensis group ( Lara 2016), and they are well nested within P. natalensis , we can thus conclude that they do not represent P. bibita .