Dryophytes leopardus Shimada et Matsui, 2025

Dryophytes leopardus Shimada et Matsui sp. nov.

(English name: East Japan tree frog)

(Japanese name: Higashi-Nihon-Ama-gaeru)

( Fig. 8 View FIGURE 8 )

urn:lsid:zoobank.org:act:107E307B-3F40-4C3D-8F5A-4E4E890C9AC0

Hyla viridis View in CoL ?: Hallowell 1861, p. 500. “Simoda, Japan ” (=Shimoda, Shizuoka, Japan)

Hyla arborea View in CoL (part): Hilgendorf 1880, p. 120. “Yeso” (= Hokkaido, Japan)

Hyla arborea var. japonica View in CoL : Bedriaga 1889, p. 475, 487. “ Tokyo ” and “Yezo” (= Hokkaido, Japan)

Hyla japonica View in CoL : Camerano 1879, p. 895. “ Tokio ( Giappone)” (= Tokyo, Japan)

Hyla arborea japonica View in CoL : Sclater 1892, p. 32. “Yezo, Japan ” (= Hokkaido, Japan); “Kiga, Japan ” (=Hamamatsu, Shizuoka, Japan)

Hyla stepheni View in CoL (part): Nikolski 1905, p. 401. Jesso (= Hokkaido, Japan)

Hyla View in CoL new taxon A: Dufresnes et al. 2016

Dryophytes japonicus View in CoL (part): Duellman et al. 2016, p. 23

Holotype. KUHE 66109 (former AUEZ 3999), an adult male from Yoshiwara , Toyota City, Aichi Prefecture, Japan (35°00’56’’ N, 137°05’38’’ E, 10 m asl), collected on 25 October 2023 by Tomohiko Shimada. GoogleMaps

Paratypes. KUHE 66111 (former AUEZ 4001) one adult male and GoogleMaps KUHE 66110, 66112–66113 (former AUEZ 4000, 4002–4003), GoogleMaps three adult females, data same as the holotype GoogleMaps .

Referred specimens. KUHE 8812, 29190–29191, 37629, Aomori Prefecture; KUHE 29240, 29244, 29246, 32922, 32925, Iwate Prefecture; KUHE 39830, Miyagi Prefecture; KUHE 40056, T3124, Akita Prefecture; KUHE 13131–13132, 13383, 18480, 18485, Yamagata Prefecture; KUHE 43347, Fukushima Prefecture; KUHE 36528, Tochigi Prefecture; KUHE 28075–28076, Ibaraki Prefecture; KUHE 36146, 38829–38830, 38832, 39551–39552, 39554–30555, AUEZ 4026, 4036, 4039–4040, 4048, 4068, 4076, 4089–4091, 4099, Niigata Prefecture; KUHE 7950, 33017, Ishikawa Prefecture; KUHE 16099, 30510–30511, 30513, 42971–42973, 43010, AUEZ 3760–3761, Gifu Prefecture; KUHE 37382, 39976, Shizuoka Prefecture; AUEZ 2616, Aichi Prefecture; AUEZ 13455–13456, 25559, 42928–42929, 42934, 42936, 42938, 42941, Mie Prefecture, KUHE 24152, 25433, 33109, 34913, 39940, 41350–41351, Shiga Prefecture; KUHE 43235–43236, Kyoto Prefecture.

Etymology. The specific epithet is a Greek (and Latin) noun denoting a leopard, alluding to the color pattern of the rear of thigh observed in this species.

Diagnosis. A moderate-sized species of the genus Dryophytes , with adult SVL 25–35 mm in males and 27–42 mm in females. It shares most morphological characters with its sister species, D. japonicus , but usually differs in having the black and/or white patterns on the rear of thigh, and they definitely differ in mitochondrial and nuclear genome characters. It differs from other congeneric species in East Asia, D. suweonensis and D. immaculatus , in the presence of finger webbing and more developed toe webbing, as well as in the pattern of the rear of thigh.

Description of holotype (in millimeters). Snout-vent length (SVL) 30.9; body robust; head short, wider (HW 12.2, 39.4%SVL) than long (HL 10.3, 33.4%SVL); snout truncate, tip rounded in dorsal outline; projecting beyond lower jaw, slightly rounded in lateral profile; canthus sharp; lore vertical, concave; nostril below canthus, midway between tip of snout (S-NL 1.5, 4.9%SVL) and anterior margin of upper eyelid; internarial distance (IND 2.5, 8.0%SVL) subequal to nostril to eye (N-EL 2.3, 7.3%SVL); eye large, length (EL 4.2, 13.5%SVL) 1.8 times eyenostril distance, equaling to snout length (SL 4.2, 13.5%SVL); interorbital (IOD 3.0, 9.7%SVL) wider than width of upper eyelid (UEW 2.7, 8.7%SVL) and internarial distance; pineal spot invisible; tympanum large and distinct, nearly circular (TDv=TDh 1.9, 6.0%SVL), about half eye diameter; vomerine teeth in indistinctly oval, small, and slightly oblique raised series (each of 3 teeth), the center posterior to line connecting posterior margins of choanae, connected with each other, but widely separated from choanae; tongue narrow anteriorly, moderately notched, without papilla; a pair of internal vocal sacs and vocal openings on corners of mouth. Forelimb stout (forelimb length 18.6, 60.1%SVL; LAL 15.0, 48.6%SVL); webbing poorly but distinctly developed; finger length formula: I<II<IV<III ( Fig. 8C View FIGURE 8 ), second finger as long as third; finger tips with round adhesive discs having circummarginal grooves; disk of third finger largest (3FDW 1.23, 4.0%), two thirds of tympanum diameter; indistinct palmar tubercles and supernumerary tubercles present; subarticular tubercles prominent, circular; indistinct nuptial pads on dorsal, medial, and ventral surfaces of first finger extending from its base to subarticular tubercle, covered with minute yellow asperities; inner palmer tubercle distinct, oval (IPTL 1.7, 5.5%SVL), subequal to tympanum diameter; indistinct skin fold present in outer side of forelimb, starting from disk of fourth finger to elbow. Hindlimb long (HLL 45.8, 148.6%SVL), about 2.5 times the length of forelimb; tibia (TL 13.8, 44.7%SVL) shorter than foot (FL 13.8, 44.8%SVL); heels do not touch with each other when limbs are held at right angles to body; tibiotarsal articulation of adpressed limb reaching posterior corner of eye; toe tips with round adhesive discs having circummarginal grooves; toe length formula I<II<III<V<IV; third toe subequal to fifth; toes moderately webbed, formula I 1–2 II 0–2 1/2 III 0–2 IV 2–1 V ( Fig. 8D View FIGURE 8 ); excision of membrane between two outer toes reaching middle subarticular tubercle of fourth when toes in contact; webs thick, not crenulate; subarticular tubercles prominent, rounded; inner metatarsal tubercle distinct, oblong (IMTL 1.2, 4.0%SVL), equaling to 3FDW, less than half length of first toe (1TL 3.5, 11.2%SVL); outer metatarsal tubercle small but distinct; indistinct tarsal fold present, connecting tibiotarsal articulation to inner metatarsal tubercle. Two distinct skin folds connecting both arms present on pectoral region, starting from anterior and posterior edges of arms, respectively. Head and dorsum smooth; no dorsolateral fold; a supratympanic fold from eye, curving to axilla; ventral and ventrolateral side of trunk and ventral side of thigh coarsely granular.

Color in life. Dorsum green fringed with a narrow yellowish white line on canthus, eyelid, supratympanic fold, and lateral side of trunk; a dark brown band running below canthus and supratympanic fold, vaguely expanding to lateral side of trunk; lores green below canthus; upper lip white with dark dots; upper half of tympanum surrounded by the band below supratympanic fold; dorsal side of thighs marked with dark crossbars; rear of thighs marked with several white blotches with reticulated pattern formed by cluster of pigmentation; anterior half of ventrum white; vocal sac and posterior half of ventrum yellowish white; ventral surfaces of thigh and tibia yellowish pink.

Variation. Females are significantly larger in SVL (mean ± SD = 33.0 ± 3.2 mm, n = 40) than males (30.4 ± 2.0 mm, n = 40; t-test, P <0.01). Size of N-LL, IND, and some characters of distal part of forelimbs (1FL, HAL, 3FDW, 1TL, and FL), all relative to SVL, tended to be greater in females than males (Mann-Whitney U-test, two-tailed, P <0.05). Similar sexual differences tended to be seen in 4TDW and LAL, but p-values were slightly larger than 0.05 (p=0.052 and 0.058, respectively). Males tended to have more developed toe webbing than females. In the second and the third toes, outer edge of web reach at toe disk in males, but reach at the first articulation in females. In the fourth toes, two phalanges are free from inner webs in males, while three phalanges are free in females.

Although the center of vomerine teeth lays posterior to the line connecting the posterior borders of the choanae in the holotype, it lays anterior the line in a part of paratypic specimens (i.e. KUHE 66110), and the latter status seems to be common in this species ( Matsui & Matsui, 1982).

Eggs and larvae. In the topotypic population, Aichi Prefecture, eggs laid at a time ranged from 109–161 (mean ± SD = 124 ± 25, n = 4) (Motohiro Nakamura, personal communication). Eggs each 1.2 mm in diameter and light brown in animal hemisphere. Eggs are usually laid in small clumps. A total of eight tadpoles in stages 31–35 (total length [TOTL] = 25.8–38.3 [mean ± SD = 33.5 ± 3.8] mm, head body length [HBL] = 11.1–14.0 [mean ± SD = 12.3 ± 1.0] mm), and three in stages 36–41 (TOTL 33.8–40.9 [mean = 37.4] mm, HBL=14.5–15.3 [mean = 14.8] mm), from the type locality were closely examined. Head and body slightly flattened above, spheroidal below; head body width (HBW) maximum slightly anterior to level of spiracle 54–68% (median = 62%) of HBL; head body depth (HBD) 89–108% (median = 97%) of HBW; snout rounded; eyes dorsolateral, visible from below; nostril open, dorsal, rim raised, midway between tip of snout and eye; internarial 29–58% (median = 49%) of interorbital. Oral disk anteroventral, emarginate, width 31–43% (median = 39%) of HBW; marginal papillae on upper labium with wide gap; lower labium with a continuous row of papillae, submarginal papillae present near corners; denticles 2(2)/3 ( Fig. 9D View FIGURE 9 ) or 2(1–2)/3; beaks with black outer margins; outer surface smooth; margin finely serrate; upper beak weekly convex medially; neither beak divided. Spiracle sinistral, tube pointing upward and backward, almost completely attached to body wall. Anal tube dextral, attached to ventral fin; loops of gut visible ventrally in young larvae. Tail moderately long and lanceolate, both margins weakly convex, tapering gradually to slightly rounded tip; tail length 178–219% (median = 197%; only specimens with non-damaged tail) of HBL, maximum depth 30–36% (median = 36%) of length; dorsal fin origin at midpoint of body, deeper than ventral fin except near tail tip and body; ventral fin origin continuous to vent; caudal muscle moderately strong, maximum tail width 36–51% (median = 44%) of HBW; muscle depth at anterior one-third of tail 34–45% (median = 41%) of tail depth, steadily narrowed posteriorly, shallower than either fin in distal half of tail. Neuromasts on body surface are not discernible. In life dorsal and lateral body brown, spotted with clusters of dark pigmentations; venter white, scattered with dark gray on throat; tail scattered with black spots ( Fig. 9A–C View FIGURE 9 ).

Karyotype. Diploid chromosome 2n = 24, with six large and six small pairs, that are homomorphic and lacking sexual difference ( Seto 1964). Chromosomes Nos. 1 and 2 in the larger group and Nos. 8 and 9 in the smaller group are metacentric, while Nos. 3, 4, 5 in the larger group and Nos. 7, 10, 11, and 12 in the smaller group are submetacentric. The large chromosome No. 6 is subtelocentric, having satellites at the tip of the short arms. Although nucleolus organizer region (NOR) is supposed to exist on No. 6 as is usual in Dryophytes frogs (Anderson 1991), an additional examination is needed for this character because there are no reports which could be exactly assigned to this species.

Call. We analyzed mating calls of ten males, recorded at the topotypic population, Aichi Prefecture, Japan at an air temperature of 16.8°C on 1 June 2023 by T. Shimada. Calls (N=10; acoustic parameters of each individual were estimated from average of five continuous notes) consisted of a series of notes each emitted at an interval (between the beginnings of two successive notes) of 0.29 ± 0.01 (0.25– 0.32) s ( Fig. 10 View FIGURE 10 ). Each note was composed of 17.3 ± 0.57 (13–20) short pulses and lasted for 0.10 ± 0.00 (0.07–0.12) s. The fundamental frequency was 1.68 ± 0.08 (1.55–1.88) kHz and the dominant frequency was 3.48 ± 0.12 (3.30–3.80) kHz. There are weak frequency and intensity modulations and some clear harmonics. Although we collected those data from males calling alone, it is known that a pair of males often call alternately, and the intervals of notes become longer in such cases compared with those emitted alone ( Aihara et al., 2006). The pulse duration is known to correlate positively with male body size, while the dominant frequency negatively correlate with it ( Takahashi et al. 2016).

Comparisons. This species shares most morphological characters with its sister species, D. japonicus . However, the typical specimens of the new species is conspicuous in having a pigmented pattern of the rear of thigh ( Fig. 1 View FIGURE 1 ), which is not popular in D. japonicus and other congeneric species of East Asia ( Fig. 6 View FIGURE 6 ; Fei & Ye 2016). Yet, this diagnosis is not perfect because there is a pattern (category C) which is sometimes appear both in D. japonicus and the new species. Further, this diagnosis cannot totally be applied for the populations close to the specific border (Kinki region). Although several morphometric differences were detected among local groups of those species, no significant differences were found between D. leopardus sp. nov. and its adjacent populations of D. japonicus (Chugoku-Shikoku group) (See “Ratios of each character to SVL” in Morphometric comparison of Results section). Dryophytes immaculatus and D. suweonensis differ from the new species in lacking finger webbing (vs. rudimentary finger webbing present) and less developed toe webbing with its excision reaching the line connecting dorsal ends of the middle subarticular tubercle of fourth toe and distal subarticular tubercle of fifth toe (vs. excision of webbing not reaching the line; Kuramoto, 1980; Borzée et al. 2020).

From the New World congeners, the new species, D. leopardus sp. nov., differs morphologically in the following manner (data from Beheld & King,1979 and Duellman, 2001): The new species has nearly smooth dorsal skin [vs. skin granulated in D. gratiosus (Le Conte) ] and has finger webbing [vs. no webbing between fingers in D. plicatus (Brocchi) and D. walkeri (Stuart) ]. In the new species, lores green below canthus and upper lip white with dark dots [vs. dark-edged light spot beneath eye in D. arenicolor (Cope) , D. avivoca (Viosca) , D. versicolor (Le Conte, 1825) , and D. chrysoscelis (Cope) ], and a narrow yellowish white line fringing on canthus and eyelid, and a narrow light line and a dark brown band running below canthus and supratympanic fold, vaguely expanding to lateral side of trunk [vs. white-edged lavender stripe on side in D. andersonii (Baird) , a light-edged purple to black stripe from snout through eye and along side in D. eximius (Baird) , D. arboricola (Taylor) , and D. wrightorum (Taylor) , white stripe along upper jaw and side of body in D. squirellus (Daudin) , and sharply defined light stripe along upper jaw and side of body in D. cinereus (Schneider) ]. In the new species, color pattern of rear of thigh is variable, and large or small white dots present, and if absent, black patterns with clusters of melanophores present [vs. uniformly brown in D. plicatus (Brocchi) , yellow spots on dark brown in D. euphorbiaceus (Günther) and D. bocourti (Mocquard) , gray to reddish-brown; yellow to white spots on dark in D. femoralis (Daudin) , and uniformly tan in D. walkeri (Stuart) ].

Range. Northeastern Japan (Hokkaido, Honshu, and some adjacent islands) and Sakhalin. Hokkaido region: Hokkaido Prefecture (including Rishiri Is., Yagishiri Is., Teuri Is., Kunashiri Is. and Shikotan Is.). Tohoku region: Aomori, Iwate, Miyagi, Akita, Yamagata (including Tobi Is.), and Fukushima Prefectures. Kanto region: Tochigi, Gunma, Ibaraki, Chiba, Saitama, and Kanagawa Prefectures (including Eno Is.) and Tokyo Metropolis. Chubu region: Niigata, (including Sado Is. and Awa Is.), Toyama, Ishikawa, Fukui (except for western tip, where mtDNA of D. japonicus was found), Yamanashi, Nagano, Gifu, Shizuoka, Aichi (including Saku Is.), and Mie Prefectures (including Toshi Is., Sakate Is., and Suga Is.) (except for southern tip, where mtDNA of D. japonicus was found). Kinki region: Shiga, Osaka, Kyoto, and Nara Prefectures. Although most part of Osaka Prefecture is supposed to be occupied by this species, the hybrid zone with D. japonicus exists at the northern area (Minoo City, Toyono Town, Nose Town) (See Fig. 2 View FIGURE 2 ). In Kyoto Prefecture, D. leopardus sp. nov. is restricted to southeastern half, and the middle to northwestern half are supposed to be D. japonicus or (if any) hybrid populations of those two species (See Fig. 2 View FIGURE 2 ). In Nara Prefecture, we genetically examined only a sample from the southern tip, which had mtDNA of D. japonicus , but judging from genetic identification of surrounding prefectures, it is natural to consider that at least northern Nara population is D. leopardus sp. nov. In Wakayama Prefecture, most populations seem to belong to D. japonicus , but we found mtDNA of D. leopardus sp. nov. at the northern and eastern tips. At least, one of them (Koya Town) is supposed to be a hybrid population, and we need further studies to examine if pure populations of D. leopardus sp. nov. exist in this prefecture. Outside of Japan, it is only distributed in southernmost part of Sakhalin ( Kuzmin & Maslova 2003).

Natural History. Dryophytes leopardus sp. nov. inhabits widely in plains and low mountains, but is sometimes seen in highland. Its altitudinal range occupies from shoreline (Oga city, Akita Pref.; Kimura et al. 2017) up to at least alt. 1,150 m (Minamimaki village, Nagano Pref.; Shimada’s personal observation). It uses various types of habitats, such as paddy fields, forests, wetlands, and gardens. In Honshu Is., it emits calls during most part of spring and summer [late April to middle August; Yamamoto 2012; Shimada et al. 2013], but some of them might include rain calls, and ovipositional activities itself are supposed to finish before high summer ( Shimada et al. 2013). The timing of reproductive start is similar in Hokkaido Is. (late April; Tokuda 2011), but is reported to be later in Sakhalin and Kunashiri Is. (late May to early June; Basarukin 1982, 1984). It breeds in various types of still water such as rice fields, ponds, ditches, wetlands, and sometimes small artificial tanks. Although Basarukin (1984) reported that females lay each single egg separately in Sakhalin, this remark should be re-checked because eggs laid in small batches are popular in other populations. Eggs are often attached to vegetation such as weeds, roots, and aquatic plants, but those laid just on muddy bottoms are seen as well.

Conservation status. Dryophytes japonicus including D. leopardus sp. nov. is listed as Least Concern (LC) in IUCN category ( Kuzmin et al. 2017). It is not listed in the Japanese Red List by the Ministry of Environment, but in Tokyo Metropolis, populations assigned to D. leopardus sp. nov. are treated as Endangered (urban area), Vulnerable (northern and southern Tama), and Near Threatened (western Tama) by the local government. Populations in Rishiri and Yagishiri Is. in Hokkaido are listed as the threatened local populations by the local government, although the former might have already been extinct (K. Kazama and M. Kazama, personal communications).