Leptopelis diffidens, Tiutenko & Zinenko, 2021

Tiutenko, Arthur & Zinenko, Oleksandr, 2021, A new species of Leptopelis (Anura, Arthroleptidae) from the south-eastern slope of the Ethiopian Highlands, with notes on the Leptopelis gramineus species complex and the revalidation of a previously synonymised species, ZooKeys 1023, pp. 119-150 : 119

Leptopelis diffidens sp. nov. Figures 2 , 3 , 4

Type locality.

Between rivers Welmel and Yadot , Harenna Forest, Bale Zone, Oromia Region, Ethiopia (6°35'N, 39°45'E) .

Material.

Holotype: Ethiopia • ♀; Segoba glade, Harenna Forest, Bale Zone , Oromia Region; 6°35'10.5"N, 39°44'30.7"E, 1770 m a.s.l.; 1 June 2019; A. Tiutenko leg.; "Found in a tussock near calling male on a flooded forest glade during rainy season"; ZSM 81 /2019 . Paratypes: Ethiopia • 1♂; Woraba glade, Harenna Forest, Bale Zone , Oromia Region; 6°35'39.9"N, 39°45'15.2"E, 1800 m a.s.l.; 5 June 2019, A. Tiutenko leg.; "Found in wet grass near slow flowing temporary stream in rainy season"; GenBank: MN 909553 ; ZSM 83 /2019 • 1 juvenile; Woraba glade, Harenna Forest, Bale Zone , Oromia Region; 6°35'39.3"N, 39°45'14.5"E, 1800 m a.s.l.; 5 June 2019; A. Tiutenko leg.; "Found in marshy grass about 10 m from a slowly flowing stream on a flooded glade"; ZSM 82 /2019 • 1♀; Katcha, Bale Mts ; 06°42'N, 39°44'E. 2400 m a.s.l.; 5 August 1986; Harenna Forest Expedition, M. J. Largen leg.; "Small stream near border between grassy clearing and dense Schefflera/Hagenia forest. Males calling from grass bordering stream"; LIV 1986.212.198 • 24 juveniles; Swamp near Shawe R ., Bale Mts ; 06°40'N, 39°44'E. 1980 m a.s.l.; 7 August 1986; Harenna Forest Expedition, M. J. Largen leg.; "Crawling up tall grasses surrounding swamp with open water and tall bushes, in Aningeria forest. Just metamorphosed juveniles, green and brown phases, latter with 3 longitudinal dark stripes, median confluent with interorbital bar"; LIV 1986.212.199-222.

Additional material.

Ethiopia • 1 larva; Woraba glade, Harenna Forest, Bale Zone , Oromia Region; 6°35'39.0"N, 39°45'14.6"E, 1800 m a.s.l.; 5 June 2019; A. Tiutenko leg.; "temporary puddle"; ZSM 172 /2019.

Diagnosis.

Medium-sized ( SVL of males ca. 24-29 mm, females ca. 35-40 mm) ground-dwelling and burrowing frog with robust body, relatively wide and short head, and short limbs. Terminal phalanges of toes and fingers expanded to small, but distinct discs. Only base phalanges of toes II-IV with broad web; on toe V the web extends along phalanges 1 and 2. The rest of toe phalanges with feeble fringe. Fingers are free of web or fringe. Light-brown or light olive-green from above; either no dorsal pattern, or three indistinct broad longitudinal bands (one vertebral and two dorsolateral) present that are slightly darker than the ground colour. Males with pectoral glands. Advertisement call: quiet, high-pitched singleton ‘quack’, repeated at intervals of ca. 20 seconds.

Description of holotype.

ZSM 81/2019, adult female. SVL 35.9 mm. The head slightly wider than its length ( HW /HL 0.77), ca. 1/3 of the SVL (HL/ SVL 0.32). Body oval. Eyes rather large ( ED /HL 0.39, ED/SL 1.15, UEW / HW 0.20), positioned laterally ( IOD 4.2 mm) and directed slightly forwards. Pupil vertical. Snout strongly curved downwards, especially in front of the nares. Canthus rostralis rounded. Naris approximately at half the distance between eye and snout tip. Tympanum visible, very close to posterior border of the orbit ( ETD ca. 1.0 mm), comparatively small: TD 1.4 mm, TD/ED 0.31. Hind limbs rather short TL+THL/SVL 0.76. Tibio-tarsal articulation reaches to the anterior of the tympanum. Outer metatarsal tubercle absent. Inner metatarsal tubercle very large ( IMT / TSL 0.24), compressed, ca. half as broad as long ( IMTW / IMT 0.57). Terminal phalanges of toes and fingers with small, but rather conspicuous discs, e.g., Fin2W/Fin2DW 0.35, Fin4W/Fin2DW 0.67, Toe4W/Toe4DW 0.68. Fingers free of webbing. Toe I free of web. On toe II the web does not reach beyond phalanx 1. On toes III and IV it reaches to ca. 1/2 of the phalanx 2, and on toe V it even extends up to the joint of the phalanges 2 and 3. The web continues as a feeble fringe along the rest of the phalanges on all toes except toe I that is free of it. Dorsal skin finely granular, almost smooth, with scattered singleton tubercles. Feeble vomerine teeth form two small groups.

Colouration in life pale green-brown or grey-green from above. Feeble one vertebral and two dorsolateral bands, a little darker than the ground colour, bordered with small irregular tubercles. The dorsal colour transits to light green and blue at thighs and shoulders. Venter off white, feebly mottled with grey. Gular area without pattern. Black band along canthus rostralis on each side of the head extending over the nostril to the eye. Behind the eye it continues over the tympanum and above the shoulder to approximately the middle of the flank. This band is not outlined. Iris dark bronze.

Colouration in preservative: Dorsum grey. Venter pale, grey mottled. Dorsolateral pattern as in life.

Variation of paratypes.

The paratype series comprises 27 specimens of various ages and sexes.

Paratype 1 (Fig. 2B ): ZSM 83/2019, adult male. It has a much slenderer body than the holotype, whose body is almost round, and is much smaller than the female: SVL 21.8 mm versus 35.9 mm. The head is similarly large (HL/ SVL 0.34), but even wider than in the female: HL/HW 0.88. Tympanum is of similar size: TD/ED 0.30. Pectoral glands visible. Digital discs are slightly larger: Fin2W/Fin2DW 0.51, Fin4W/Fin4TW 0.72, Toe4W/Toe4DW 0.77. Skin on all parts of the body and head smooth. Colouration in life: Dorsal colouration is very similar to that of the holotype: Pale olive-green from above, with three indistinct longitudinal bands. Ventral colouration is different and pale, without pattern. Throat with some scattered dark spots. Sides of thighs with dark grey blotches and bands. Similar pattern also between flanks and venter; this pattern is absent in the holotype. Similar to holotype, a dark band extends from snout tip, over the eye and above tympanum; however, it does not continue at both sides of the body beyond the shoulder. Unlike in holotype, there are no dark blotches on flanks. Colouration in preservative like in life but dorsal colour is grey.

Paratype 2 (Fig. 2C ): ZSM 82/2019, metamorph at Gosner stage 45. Very small individual, SVL 12.9 mm, with tail stub - 5.9 mm. Head is similarly large in comparison to body as in adult individuals: HL/SVL 0.34. Other proportions are similar: HL/HW 0.89, ED/HL 0.46, ED/SL 0.95, TD/ED 0.30, TL+THL/SVL 0.63, IMT /TSL 0.29. The inner metatarsal tubercle is flatter than in adults: IMTW / IMT 0.25. Digital discs are similarly conspicuous in this stage of the life cycle: Fin2W/Fin2DW 0.83, Toe4W/Toe4DW 0.75. In life, the colouration was very similar to that of the male Paratype 1.

Paratype 3 (Fig. 2D ): LIV 1986.212.198, adult female with even larger SVL than in the holotype - 39.6 mm, but with almost identical morphology and very similar appearance: HL/SVL 0.32, HL/HW 0.79, ED/HL 0.36, ED/SL 0.91, TD/ED 0.39, TL+THL/SVL 0.69, IMT /TSL 0.27, IMTW / IMT 0.99, Fin2W/Fin2DW 0.51, Fin4W/Fin4TW 0.72, Toe4W/Toe4DW 0.72. It is an old specimen that had been probably fixated or even kept for some time in formalin prior to current storage in alcohol; therefore it is hardly possible to compare its colouration with more recently collected and immediately ethanol preserved specimens. In its current state the dorsum of this specimen appears bluish-grey; the venter is yellowish. No pattern is recognisable, except dark lateral band which is similar to that in the holotype. Probably the live colour of dorsum of the Paratype 3 has been more uniformly green.

Paratypes 4-27 (Fig. 4 ): LIV 1986.212.199-222, recently metamorphosed individuals with SVL 11.1-14.5 mm (see Suppl. material 5: Table S1 for morphometrics of each specimen). They closely resemble the male Paratype 1 in body shape but are smaller and lack pectoral glands. In comparison with Paratype 2, they lack tails, i.e., have been at a more advanced development stage (Gosner stage 46) at time of collecting, though they are of similar size or just slightly larger. Like all other specimens of the type series, they have more or less continuous dark lateral bands extending from the snout tip to the middle of the flanks or further. Eight specimens appear rather bluish, compared to the rest of the series whose dorsal colour is brown. This may indicate that they were green in life. Among brown specimens some have also bluish areas which may have been green in life as well. The ventral side is uniformly pale, without any spots or pattern.

Differential diagnosis.

Leptopelis diffidens sp. nov. shares the following traits with the rest of the members of the genus Leptopelis : The terminal phalanx of finger kinked (out of alignment with the rest of phalanges); no fingers opposing each other; digital discs on fingers present; outer metatarsal tubercle absent; singleton subarticular tubercles; tympanum visible; pupil vertically elliptic; vomerine teeth arranged in two groups between choanae.

The digital discs both, on fingers and toes, are in the new species significantly more conspicuous than L. sp. ‘Shewa’ and in the parapartic high-altitude population in Bale and Arsi Mountains - L. montanus nom. nov. (Figs 5 , 6 ). In the examined specimens of both high-altitude species the digital discs were virtually absent: On their fingers there are just feeble pads (Fin2W/Fin2DW 0.92, Fin4W/Fin4 DW 0.86), and on toes the tips are even narrower than the phalanges (Toe4W/Toe4 DW 1.8). The digital discs in L. diffidens sp. nov. are not significantly larger than in L. gramineus sensu stricto (Fig. 6 ). In two paralectotypes and five topotypic specimens that we measured the average ratio 'disc width to phalanx width’ was 0.85 on finger II (Toe2W/Toe2DW) and 0.70 on toe IV (Toe4W/Toe4DW).

The examined males of both, L. sp. ‘Shewa’ and L. montanus nom. nov., are much larger than in the new species: SVL 27.2-40.6 mm (mean 35.6 mm). Females in these two species are even larger, with SVL 47.5-56.6 mm (mean 51.5 mm). Generally, females of L. diffidens sp. nov. are sized like males of the montane species in this species complex (see comparison photographs in Suppl. material 12).

All examined males of L. gramineus sensu stricto (three specimens, including paralectotype BMNH 1947.2.10.20) were significantly larger than of L. diffidens sp. nov.: mean SVL 36.1 mm; the size difference between male and female in L. gramineus appears to be small. Two of the examined three females (including paralectotype BMNH 1947.2.10.19) had SVL 39.72 mm and 39.37 mm. The third female (BMNH 1975.1633) with SVL 24.39 mm may be immature.

A principal component analysis showed that the low-altitude geographic group from the Harenna Forest that we describe here as L. diffidens sp. nov. is separated from the rest of the groups along the first axis accumulating 44.99% of morphological variation. The other groups are partially separated along the second axis accumulating 10.85% of variation while the most divergent groups are L. gramineus sensu stricto and the highland populations from the east of the GRV ( L. montanus nom. nov.). The specimens from ‘Borana/Sidamo’ population are placed between them and partially overlap with both. See this plot in Suppl. material 10.

A canonical discriminant analysis separated the three groups L. diffidens sp. nov., L. montanus nom. nov., and L. gramineus sensu stricto with high confidence (see Fig. 7 ): Wilks’s Lambda approx. 0.00048, F (58, 26) = 20.111, p <.0000. Four measurements (HL, TL, Toe3L, Toe4L) had the largest contribution to the discriminant function. A discriminant analysis summary and standardised coefficients for canonical variables are provided in Suppl. material 1 and 3. In spite of moderate sample size and combining both sexes in a single multivariate analysis, we revealed strong differences in the body proportion between three lineages of the L. gramineus complex, manifested in head proportions, tibia, and longest toe lengths. As Lada et al. (2005) showed for European Pelobates , the fossorial way of life is a strong constraint for morphological variation and sexual dimorphism. Thus, the morphological divergence that we observed in the studied species of L. gramineus complex may be related to different extent of fossoriality.

Compared to all other frogs currently assigned to L. gramineus , the new species appears to have smoother skin on dorsum, with just some small singleton warts. L. diffidens sp. nov. differs from other members of the complex also in overall dorsal colouration: It is rather grey-green or pale beige while L. gramineus and other high-altitude species of this complex have more bright green or yellow-brown colouration. Furthermore, the new species has no such dorsal pattern of scattered blotches and ocelli as in the parapatric L. montanus nom. nov. The latter, in turn, usually lacks continuous black lateral bands that seems to be typical for L. diffidens sp. nov. (Fig. 3 ). Canthus rostralis in L. diffidens sp. nov. is not outlined light yellow or light green as in other members of this complex.

From non-sympatric, but also ground-dwelling, L. bocagii the new species differs by less robust body form, smaller head, rounded canthus rostralis (versus strongly angled in L. bocagii ). The digital discs in L. bocagii are similarly expanded except on toes IV and V where they seem to be larger. The inner metatarsal tubercle is similarly large and compressed. Unlike in L. bocagii that has strong vomerine teeth, they are feeble in L. diffidens sp. nov. and additionally, the colouration of L. bocagii is very different, see Largen (1977) for a detailed description and illustration.

Leptopelis diffidens sp. nov. is generally ground-dwelling and does not occur on trees or scrubs, although males and juveniles may occasionally climb grass. From all scansorial members of the genus distributed at the Horn of Africa, including sympatric L. ragazzii (Boulenger, 1896), it differs by much more robust body (particularly in females), comparatively small head, shorter limbs, reduced digital discs. Moreover, L. ragazzii has a flatter head with less curved snout and comparatively larger eyes. This applies also to non-sympatric L. vannutellii (Boulenger, 1898), which does not differ morphologically from L. ragazzii . Both, L. ragazzii and L. vannutellii , have small inner metatarsal tubercle and well-developed webbing on feet, extending on toes III and IV well beyond the joint of the phalanges 1 and 2. Similarly to L. ragazzii that has two clearly different colour phases, bright green and brown, occurring together. Leptopelis diffidens sp. nov. exhibits such dichromatism as well, although not that distinct: unlike in L. ragazzii , the brown phase in the new species is not purely brown, but rather olive-green or grey-brown. No plain green phase was ever recorded, but grey-green or pale green individuals occur in populations of L. vannutellii . Overall, the colour of L. vannutellii is not like that of L. diffidens sp. nov.

The non-sympatric species L. susanae and L. yaldeni Largen, 1977 have much broader digital discs, small inner metatarsal tubercles, and more extensive webbing on toes III and IV than the new species. Both are scansorial forms with more slender bodies. Leptopelis yaldeni has two colour phases which are clearly distinct, green and brown, like in L. ragazzii. Similar to L. diffidens sp. nov., L. susanae does not show such clear dichromatism, but its general colouration clearly differs very much from the new species (compare photographs and description in Largen 1977).

Another species, L. concolor Ahl, 1929, recorded from the Juba river basin, south-east of the distribution area of L. diffidens sp. nov., is a scansorial form with a similar body size that occupies coastal savanna habitats. Similar to the new species, it has reduced pedal webbing but larger discs, particularly on toes III, IV, and V. Like other arboreal Leptopelis , it has a slender body and a large head. The colouration of L. concolor is rather constant: light brown from above, with dark interorbital bar and an indistinct reversed ‘Y’ on dorsum.

Advertisement call.

Males usually call from tussocks surrounded by water, but also from tall and dense grass at margins of pools and puddles, and occasionally from shrubs surrounding the glade. The animal sits near the roots of the grass, but not on grass and not in water. When approached, it stops calling and becomes silent for many minutes. The calls are rather weak and may be difficult to perceive in places with background chorus of other frogs, particularly of Ptychadena , or with loudly shouting birds. During a rainy season, in May-June, we did not notice any significant difference in the intensity of vocalisations between day and night. However, in the same season we observed more calling individuals and more frequent calls in the parapatric population of L. montanus nom. nov. in the Bale Mountains.

Like other Ethiopian members of this species complex ( Largen 1977; Schiøtz 1999), the advertisement call of L. diffidens sp. nov. is a ‘quack’. It consists of a single note that contains eight pulses with interval of ca. 20 ms. The tone frequency is 990-2700 Hz (Fig. 8A ). It is higher than in the call of the high-altitude member of the L. gramineus complex from the Bale Mountains that also consists of a single note, however, of five pulses and with a frequency of 344-3270 Hz (Fig. 8B ). According to our observations, L. diffidens sp. nov. repeated their ‘quack’ with equal intervals of ca. 20 seconds. Another male may use the pause for his call. We did not observe two or more males calling synchronously. Largen (1977) mentioned a ‘scream’ that sometimes precedes the ‘quack’ in L. gramineus from high-altitude population ( L. montanus nom. nov.). We did not observe such vocalisations in the new species at any localities we visited.

The vocalisation of L. ragazzii partially resembles that of L. diffidens sp. nov. in containing a ‘quack’ sound of similar pitch and frequency. However, it is normally preceded by a screech, lasting for ten or more seconds, that consists of 4 or 5 notes with increasing pitch and frequency (Fig. 8C ). L. diffidens sp. nov. does not produce such sounds.

Genetics.

The 16S mitochondrial gene sequence (GenBank no. MN 909554 ) of the specimen ZSM 83 /2019 (Paratype 1) matches to 99.8% the corresponding sequences obtained by other workers from tissue samples of frogs that had been collected in the Harenna Forest at Katcha and in vicinity of Rira and identified as L. gramineus . Freilich et al. (2016) applied phylogenetic reconstructions based on the COI gene and found four highly divergent mitochondrial lineages in L. gramineus with average distance of 6.4-8.0% between them. They restricted three of these clades to the east of the GRV and called ‘Arsi’, 'Kibre Mengist’ and ‘Kasha’. These correspond to what we call here L. montanus nom. nov., L. sp. ’Borana/Sidamo’, and L. diffidens sp. nov. (see Suppl. material 8: Table S 4) .

As Reyes-Velasco et al. (2018) showed with their genetic study using multiple RAD loci, the population from the Harenna Forest and the related population from Bore and Kibre Mengist area ( L. sp. ‘Borana/Sidamo’) represent together a sister clade of L. susanae . This lineage diverged from the high-altitude species L. montanus nom. nov. around 4 my ago, during the Pliocene. The split between eastern and western lineages of the L. gramineus complex took place ca. 6 my ago. Therefore, the new species from the Harenna Forest is more distantly related to species from the west of the GRV, hence also to L. gramineus sensu stricto.

In order to align our material to the already defined groups we have constructed a statistical parsimony network of 16S haplotypes (showed in Fig. 9 ). Both, the paratype of L. diffidens sp. nov. (GenBank no. MN 909554 ) and the sequenced specimen from the Gaysay Grasslands (GenBank no. MN 909551 ) clustered respectively, with the representatives of the geographic groups of the low-altitude Harenna Forest (i.e., clades ‘Harenna’ sensu Reyes-Velasco et al. 2018 and ‘Kasha’ sensu Freilich et al. 2016) and of the high-altitude population that Freilich et al. (2016) called ‘Arsi’ and Reyes-Velasco et al. (2018) called ' Bale Mountains’ .

Etymology.

The Latin adjective Leptopelis diffidens (used here in nominative singular) means diffident, anxious, shy, mistrustful. This name refers to the behaviour of this frog that appears even more cryptic and defensive than L. gramineus from other populations. The vocalisation of a male is so quiet that the animal is very difficult to locate and to spot. When approached, at a distance of only 5 m, it stops calling and remains silent until the disturber has gone away or has not moved for at least 10 minutes. As a vernacular name for this species, we propose 'Harenna Burrowing Treefrog’.

Distribution and habitat.

According to our current knowledge, the new species is endemic to the Harenna Forest where it occurs at elevations of 1400-2300 m a.s.l. We observed these frogs and recorded vocalisations at the following localities, which are glades in a tall forest: Gola, Hacho, Harawa, Haro Alati, Hordoba, Kaffa Guasaa, Manyate, Megano, Segoba, Sire, Woraba, and Yagana (Fig. 9 ). This species was also collected many times from Katcha and nearby localities.

Leptopelis diffidens sp. nov. seems to be confined to glades in the forest that are temporarily flooded with rain water during rainy season. These frogs do not live in areas of dense high trees, where they are replaced by L. ragazzii , typically found along streams. These two species never occur syntopically. We also did not encounter L. diffidens sp. nov. in man-made open areas in the forest, such as clearings, plantations, or settlements. However, we found two males of L. ragazzii (brown and green phase) on Aboye, a large clearing regularly used for barley plantation. Moreover, unlike L. ragazzii and the highland members of the L. gramineus complex whose larvae live in fast-flowing water, the new species uses lentic or slow-flowing waterbodies for breeding and does not occur at fast flowing streams (Fig. 9 ). We assume therefore that this frog requires a certain flora composition and environmental, particularly hydrological, conditions that are available only in natural glades.

Leptopelis diffidens sp. nov. shares breeding sites with Phrynobatrachus inexpectatus , Ptychadena sp., and Xenopus clivii . We observed this even in the middle of Manyate, a large village in the south of the Harenna Forest, because it has been built around a natural glade that retained its flora and hydrological characteristics.

An altitudinal and ecological barrier exists between L. diffidens sp. nov. and the not-yet-named montane grassland-dwelling L. montanus nom. nov.: L. diffidens sp. nov. does not ascend the Harenna Escarpment and is absent already in the sub-alpine heathlands, i.e., above 2500 m a.s.l. On the other hand, L. montanus nom. nov. that is distributed in the high-altitude areas, including the Sanetti Plateau, was not recorded in the areas directly adjacent to the Harenna Escarpment that have elevations around 4000 m a.s.l.; hence, no contact zone seems to exist here (Fig. 9 ). Leptopelis diffidens sp. nov. may not range west beyond the Ladamo mountain chain because suitable glades are absent there. This ridge also seems to isolate L. diffidens sp. nov. from populations of L. sp. ‘Borana/Sidamo’, a not-yet-named sister clade which was reported from localities further west, in Borana and in northern Sidamo ( Reyes-Velasco et al. 2018). In the east, the range of L. diffidens sp. nov. reaches into Berbere area of the Harenna Forest where the forest gradually transits to bushland and savanna. The southern range boundary seems to be at Manyate and Haro Alati glade. South of this line glades disappear and the forest merges with Acacia bushland.

Sexual characters.

Males can be distinguished from females by a much smaller body size, less robust body, larger head, and the presence of pectoral glands.

Natural history.

Leptopelis diffidens sp. nov. is a ground-dwelling and semi-fossorial frog that spends most of its life hidden. In the dry season it is nocturnal, but in the rainy season we did not notice a significant difference between diurnal and nocturnal activity patterns at least in males.

Since we found a gravid female of L. diffidens sp. nov. and individuals at the last stages of metamorphosis on the same day and at the same locality, we assume that this species breeds more than once a year. Its breeding seasons appear to be out of sync with that of the parapatric high-altitude population of L. montanus nom. nov. in the Bale Mountains. We found metamorphs of L. diffidens sp. nov. ( ZSM 82/2019 and ZSM 172/2019) in June, at the end of the short rainy season. A few days earlier we had visited a population of L. montanus nom. nov. in the Gaysay Grasslands and found neither metamorphs nor larvae. The rainy season there had just begun. In the Web Valley, Bale Mountains, we observed large tadpoles of L. montanus nom. nov. in the middle of February, at the end of a dry season. However, just a week later at Katcha in the Harenna Forest we found all puddles dried, and no frogs were around. During another trip, we found tadpoles in advanced development stages and metamorphs of L. montanus nom. nov. in vicinity of Dinsho village, Bale Mountains, in early November but saw neither adults nor larvae of L. diffidens sp. nov. after that in the Harenna Forest where the rainy season was approaching its end.

Larva.

The tadpole of L. gramineus has been described by Largen (1977) based on specimens from high-altitude populations and compared with the tadpole of sympatric L. ragazzii by Tiutenko and Zinenko (2019). Considering the morphological similarity and close phylogenetic relationship of L. diffidens sp. nov. we expect the tadpole of the new species to be similar to the tadpoles in the rest of this species complex. Except for one specimen ZSM 172/2019 at Gosner stage 43, no larvae of L. diffidens sp. nov. have been collected so far. This specimen was at time of collection shortly before leaving water and resembles an adult L. diffidens sp. nov. closely enough to permit positive identification. It has completely developed forelimbs. Instead of larval mouthparts it has an almost completely developed oral cavity and even a tongue. However, the development of the mouth opening is incomplete: it is only 2.4 mm wide, oval, and directed forward. Additionally, the tympanum is not yet present. All measurements are a little larger than those of another specimen ( ZSM 82/2019) that we found at the same locality in a more advanced metamorphotic stage (see Suppl. material 5: Table S1).

Status of other cryptic species in Leptopelis gramineus complex.

Largen (1977) pointed out in his review of the Ethiopian Leptopelis that there is a " great variation of size which has been demonstrated between, but not within, populations " of L. gramineus . Since his so-called ‘populations’ are now treated as cryptic species, the pattern of these size differences has become clearer. The high-altitude species are generally considerably larger than the low-altitude ones. The 12 small male specimens from Wando ( BMNH 1973.2164-2176) that Largen mentions are probably of the species that we call here L. sp. ‘Borana/Sidamo’ which is closely related to L. diffidens sp. nov. that is characterised by the same ‘dwarfism’. The largest ( BMNH 1975.1638) of the 27 males examined of this species measured 29.6 mm. The largest of four females examined measured 40.0 mm. The mean SVL values are 25.4 (males) and 38.6 mm (females).

The population at the high-altitude lake Wonchi that Largen also mentions as an example of small-sized L. gramineus requires an additional study. Unfortunately, we did not find the specimens that Largen refers to ( AAU H.548/1-2 and AAU H.718/1-4) during our visit in the ZNHM. Geographically this population should be within the range of L. sp. ‘Shewa’, and the locality is situated at high elevation of almost 3500 m a.s.l. This was also confirmed by DNA barcoding performed by Mengistu et al. (2012). Whether small body size is typical for this population, is yet to be determined.

For resolving the taxonomic status of the western populations of L. gramineus , new material and DNA samples from the type locality or from nearby highlands in Ethiopian administrative zones Gamo Gofa, Keffa, Wolayita, Kenta, and Dawuro are required together with further studies of morphological differences, bioacoustics, ecology, and contact zones. Mengistu et al. (2012) mentioned tissue samples of specimens from Gamo Gofa, but did not report any further results of their analysis. Other workers ( Freilich et al. 2016; Reyes-Velasco et al. 2018) did not include topotypic material in their published results. Reyes-Velasco et al. (2018) had obtained nuclear DNA sequences of vouchers from Dorse and Chencha (Gamo Gofa), and from Bonga (Keffa), places at and close to the type locality, but apparently did not use them in their analysis for some reason. Jimma is the geographically closest locality from which DNA sequences were analysed so far. Reyes-Velasco et al. (2018) found that this population is rather different from L. gramineus further north, around Addis Ababa, Wonchi, and at other localities in Shewa and Gojam ( L. sp. ‘Shewa’). We compared specimens of L. gramineus sensu stricto (paralectotypes and topotypic specimens housed in BMNH) with specimens from the north and found considerable difference between these populations in digital discs size, skin surface structure, body size and colouration. Therefore, it appears likely that the ‘northern’ population ( L. sp. ‘Shewa’) represents a different species. Ahl (1924: 9) described a new species Pseudocassina rugosa , with type locality in Meta. In his later work he assigned it to the genus Leptopelis ( Ahl 1931: 222) without any explanation. Largen synonymised L. rugosus with L. gramineus in the course of a revision of Ethiopian Leptopelis ( Largen 1977). If comparison of genetic, acoustic, and morphological data with topotypic material would confirm a sufficient level of divergence, reinstatement of Leptopelis rugosus (Ahl, 1924) may be considered. Also, it is not clear how far this putative species ranges southwards, so there is no confidence that it does reach the type territory of L. gramineus .

In the same paper Ahl (1924) described Pseudocassina ocellata , another species that Largen (1977) synonymised with L. gramineus . Its type locality is Dida Plateau ('Hochebene Didda’) which he first erroneously placed in Somalia, as he did with some other specimens collected by Neumann and Erlanger (see discussion in Largen 1975). This plateau is in fact situated in the Arsi Mountains, and the Leptopelis population there is certainly conspecific with frogs from other localities in Arsi and Bale. According to the published genetic data, this clade is reproductively isolated from other lineages of L. gramineus complex to the west from the GRV and could have limited exchange of genetic material with Kofele lineages sensu Reyes-Velasco et al. (2018). Therefore, we resurrect this species from synonymy and raise L. ocellatus (Ahl, 1924) to the status of a valid species in genus Leptopelis . As the vernacular English name we propose 'Ocellated Burrowing Treefrog’ because it describes the typical colouration of this animal and matches the original description by Ahl (1924). However, the Latin name Leptopelis ocellatus is not available in this genus because a valid species from West Africa was described earlier and now bears the same name: L. ocellatus (Mocquard, 1902). In accordance with Article 50 and Paragraph 3 of Article 60 of ICZN, we establish a substitute name Leptopelis montanus nom. nov. for Leptopelis ocellatus (Ahl, 1924) to resolve its homonymy with Leptopelis ocellatus (Mocquard, 1902).

Etymology.

The new specific name Leptopelis montanus (living in mountains, montane) refers to the fact that this species ranges into afromontane areas up to elevation of almost 4000 m a.s.l., thus being a Leptopelis with probably the highest altitudinal distribution. The name is an adjective in nominative singular.

Figures

Figure 2. Types of Leptopelis diffidens sp. nov. (in dorsal and ventral view) A holotype, female, ZSM 81 / 2019 B paratype 1, male, ZSM 83 / 2019 C paratype 2, metamorph, ZSM 82 / 2019 D paratype 3, female, LIV 1986.212.198.

Figure 3. Adult females of three species of the Leptopelis gramineus complex in life, shown to the same scale A L. diffidens sp. nov., Harenna Forest (holotype, ZSM 81 / 2019) B Leptopelis montanus nom. nov., Gaysay Grasslands, Bale Mountains (not collected) C L. sp. ' Shewa', Menz-Guassa (not collected).

Figure 4. Paratypes 4 - 28, immature individuals, LIV 1986.212.199 - 222 (in dorsal and ventral views).

Figure 5. Comparison of digital discs (on right hand and foot) and of pedal webbing in A Leptopelis diffidens sp. nov., and B L. montanus nom. nov.

Figure 6. Pedal webbing of five members of the Leptopelis gramineus complex A L. diffidens sp. nov., LIV 1986.212.198 B L. gramineus, BMNH 1947.2.10.19 (paralectotype) C L. gramineus, BMNH 1975.1619 D L. sp. ' Borana / Sidamo', PEM 3821 E L. montanus nom. nov., BMNH 1975.1621 F L. sp. ' Shewa', BMNH 1969.969.

Figure 7. Grouping of three species of Leptopelis gramineus complex on a plane of two canonical roots. Discriminant analysis summary and standardised coefficients for canonical variables are provided in Suppl. material 1 and 3.

Figure 8. A comparison of singleton advertisement calls of Leptopelis diffidens sp. nov. and L. montanus nom. nov. B oscillograms (top) and spectrograms (bottom) of advertisement calls of L. diffidens sp. nov., L. montanus nom. nov.), and L. ragazzii within 40 seconds.

Figure 9. Network of haplotypes and a map showing the distribution of the Leptopelis gramineus species complex members: L. gramineus (filled circle), L. sp. ' Shewa' (empty circle), L. montanus nom. nov. (empty triangle), L. diffidens sp. nov. (filled square), L. sp. ' Borana / Sidamo' (empty diamond). Type localities of the described species are marked with a cross. The colours on the haplotypes chart correspond to various groups recognised by Mengistu et al. (2012), Freilich et al. (2016) and Reyes-Velasco et al. (2018). The symbols in the legend correspond to populations shown on the map.