Bioacoustics of Scinax rupestris

Diagnosis. Scinax rupestris sp. nov. is a member of the S. ruber clade for having the single morphological synapomorphy known for the group, the tadpole vent tube that does not reach the free margin of the lower fin. The new species can be diagnosed by the following set of characters: (1) moderate size (SVL in males 21.9–27.7 mm , females 26.7–31.7 mm ); (2) snout acuminate in dorsal view, rounded in profile; (3) tympanum medium-sized (TD 61.1–71.0% of ED); (4) vocal sac single, median, subgular, that does not reach the pectoral region, and externally evident by the loose skin on the sides of jaw; (5) iris iridescent yellow, with some thin, darker reticulations; (6) tadpoles with ventral oral disc; (7) regular P-3, unmodified as a labial arm; (8) absence of keratinized and colored plates on the sides of the lower jaw-sheath; (9) presence of a keratinized and colored spur on each side behind the lower jaw-sheath; (10) dorsolateral eyes, invisible ventrally; and (11) advertisement call composed of 8–14 notes each with 4–18 pulses, and duration of 290–420 ms. FIGURE. 1 . Scinax rupestris sp. nov. , holotype (MZUSP 112877 ; SVL 25.2 mm ). A: Dorsal view. B: Ventral view. Photos: M.R.C. Comparison with other species. The Scinax ruber clade includes 66 species, of which 55 are not included in the two monophyletic groups currently recognized: Scinax rostratus and Scinax uruguayus groups ( Faivovich et al. 2005 ; Frost 2014 ). For this reason we present comparisons with all species. The structure for the comparison is based first on obvious size differences in adults (that is, no overlapping nor a minimal gap between size ranges), followed by more detailed comparisons with species that cannot be differentiated on the basis of size, or in case that there are conspicuous external morphological characters.

Description of holotype . Body moderately robust, head rounded in profile, acuminated in dorsal view; head as large as wide, 37.7% of SVL. Nostrils dorsolateral, elliptical, slightly protruded; distance between nostrils 64.3% of IOD. Canthus rostralis evident and convex. Loreal region slightly concave. Eyes protuberant, ED 10.7% larger than IOD, almost equal to END . Tympanum rounded, separated from eye by a distance almost half TD. TD 61.3% of ED. Supratympanic fold barely evident, from the corner of the eye to the insertion of the arm. Vocal sac single, median, subgular, externally evident by the loose skin on the sides of jaw. Vocal slits present, located diagonally to the longitudinal body axis, originating laterally to the tongue and running towards the corner of the mouth. Tongue elliptical, free laterally and posteriorly notched. Vomerine teeth in two slightly convex series between and only just posterior the choanae, each bearing six teeth. Choanae oval. Axillary membrane absent. Upper arm slender, forearm moderately robust. Fingers slender, subarticular tubercles single, conical in fingers I and II; rounded on fingers III and IV. Subarticular tubercle in the third finger smaller than the others. Supernumerary tubercles absent. Relative finger length I<II<IV<III. Discs elliptical, wider than long; disc on Finger I slightly smaller than the others. Inner metacarpal tubercle single, medium-sized, elliptical; outer metacarpal tubercle flat, medially divided. Webbing absent between fingers I and II; basal between fingers II, III, and IV. Basal fringe between fingers I and II. Thick, wide, light colored nuptial pad covering almost the complete dorsal surface of Metacarpal I and medially obscuring nearly half of the inner metacarpal tubercle (Fig. 3A). Hind limbs robust; TL 52.8% of SVL. Toes slender; subarticular tubercles single, conical, and rounded. Supernumerary tubercles poorly developed, single, and rounded. Relative toe length I<II<V<III<IV. Discs elliptical, wider than long. Inner metatarsal tubercle single, oval; outer metatarsal tubercle simple, rounded. A poorly developed fringe on the lateral margin of foot, originating approximately at middle of metatarsus and joining the lateral fringe of Toe V. Webbing formula I 2 -–2- II 1 1/2 –2 1/ 2 III 1– 2 - IV 2– 1 V. Skin on dorsum smooth, with some scattered granules located mainly on the interorbital portion and between the canthus rostralis. Ventrally the body, ventral surfaces of thighs, and subcloacal region densely covered with rounded, low granules. Ventral surfaces of arms and tibiae smooth. Pectoral fold present, with pre- and postaxillar elements present. No glandular patches apparent in the pectoral region. FIGURE. 3 . A: Nuptial pad of Scinax rupestris sp. nov. , holotype (MZUSP 112877 ). Scale bar = 1 mm . Color in preservative of the holotype . Dorsum brown with some scattered small round and irregular dark blotches; fore- and hindlimbs with irregular, transverse dark markings. Flanks and inguinal region lighter, with diffuse dark blotches. Hidden area of thigh and shank brown with some diffuse light blotches. Venter creamy white. Measurements of the holotype (mm). SVL 25.2; HL 9.5; HW 9.4; IND 1.8; IOD 2.8; ED 3.1; END 2.8; TD 1.9; TL 13.3; FL 10.8. Variation in the type series. Some measurements are presented in Table 1 . The number of vomerine teeth in males varies between three and seven on both the right and left processes. In one male paratype (MZUSP 112859 ) the right process (three teeth) is smaller (about half of the left process size) than the left process (four teeth). Dorsally, small, scattered, slightly conical and rounded tubercles are evident mainly on head (68.2%) and body (68.2%), sometimes on limbs (36.4%), and less commonly on eyelid (22.7%). Ulnar tubercles, small and low are in line on the ventrolateral portion of the forearm, starting immediately after the elbow reaching approximately the wrist (68.2%). In some individuals these ulnar tubercles are white (13.6%). In addition, a fringe on the lateral margin of the metacarpal is present; it starts at level of the anterior margin of the palmar tubercle for after to join to the external fold of the Finger V, and can be complete (9.1%) or interrupted (50.0%). Measurements Females ( n = 2) Mean ± SD Range Males ( n = 27) Mean ± SD Range SVL 29.18 ± 3.49 26.7–31.6 24.94 ± 1.33 21.9 – 27.7 HL 10.57 ± 0.95 9.9–11.2 9.42 ± 0.52 8.5 – 10.7 HW 10.05 ± 1.33 9.1 – 11.0 9.24 ± 0.48 8.3 – 10.3 ED 3.03 ± 0.49 2.7 – 3.4 2.92 ± 0.18 2.6 – 3.3 IND 1.85 ± 1.78 1.7 – 2.0 1.87 ± 0.08 1.7 – 2.0 IOD 3.24 ± 3.17 3.1 – 3.4 2.64 ± 0.26 2.3 – 3.3 TD 2.16 ± 0.11 2.1 – 2.2 1.91 ± 0.16 1.6 – 2.3 END 3.02 ± 0.47 2.7 – 3.3 2.70 ± 0.24 2.0 – 3.1 TL 14.37 ± 2.10 12.9 – 15.8 12.79 ± 0.75 11.4 – 14.6 FL 10.81 ± 1.85 9.5 – 12.1 10.05 ± 0.73 8.6 – 11.5 Tarsal tubercles, small and low could be present on the ventrolateral portion of the tarsus, starting immediately after the tibia-tarsus articulation and almost reaching the outer metatarsal tubercle (18.2%). These tubercles can also be white pigmented (22.7%). An internal fold, elliptical is present on the internal margin of the tarsus, starting immediately after of the tibia-tarsus articulation followed by three or four small tubercles towards to the inner tubercle metatarsal, reaching the anterior third of the tarsus (31.8%); the fold and tubercles are white pigmented in two specimens. Otherwise, the small tubercles are absent in some individuals (36.4%). Additionally, calcar tubercles small and low can be present in some individuals (22.7%). Webbing formulae among paratypes varies as follows: I (2–2-) – (2–2-) II (1–1-) – (2-–2½) III (1–1-) – (2- 2 ½) IV (2–1½) – 1 V. The two available mature females are larger than males. Their forearms are not robust as in males. There is a single male (MZUSP 112875 ) that shows on the internal margin of the left arm a light colored, glandular pad, that we consider an aberrant occurrence considering that it is absent on the right arm and in all other male specimens available to us. Most specimens have a thickening of the ventral portion of the webbing between toes IV and V that is also evident on the dorsal aspect as a number of wrinkles or an ampulla-like dilation. This is common in several species of Scinax , and seems related to a parasite infection, as unidentified eggs are sometimes visible under high magnification in the affected webbing. In life, the species show notable color differences by night and day. At night, the dorsal color pattern of head, body and limbs could be dark brown-olivaceous, with small round and irregular dark-brown blotches (Fig. 4D–E); dark-brown with dark round and irregular blotches and some white spots on head and body, but with limbs with dark transversal bars (Fig. 4D); or head and body overall dark-brown, with limbs yellowish with some irregular brown blotches (Fig. 4C). Flanks vary from yellowish to olivaceous. Venter immaculate white. Iris iridescent yellow, with some thin, darker reticulations. During the day, the dorsal color pattern of head, body and limbs are marbled creamy white, with small, round and irregular dark gray blotches. In some individuals, these blotches form transversal bars on the limbs and a discrete interocular blotch (Fig. 4A–B). Flanks are pale-yellowish. FIGURE. 4. Scinax rupestris sp. nov. A and B: Diurnal pattern color. C, D and E: Nocturnal pattern color. Photos: R.A.B. In preservative, the dorsal color pattern of head, body and limbs varies from light and dark gray to dark brown, with some round and irregular dark blotches; in some specimens, small white spots are present on head, body and limbs. Additionally, two individuals show an inverted interocular triangular blotch. Hidden surfaces of thigh and shank light or dark brown, with light, diffuse blotches. Flanks and inguinal region lighter, with diffuse dark blotches. A dark brown canthal line is present in some individuals (45.5%). Transversal dark bars on limbs. FIGURE. 5. Waveform (above) and spectrogram (below) of the advertisement call of Scinax rupestris sp. nov. (CHUNB 73649) recorded at Chapada dos Veadeiros, Goiás, Brazil , depicting 10 notes, with 5–14 pulses. Notice isolated single pulses between notes, and at the beginning and end of the call. The final pulse of a given note is more intense than the surrounding ones and the uprising sound intensity towards the end of the call. Calls. Air temperature and humidity during recordings were 22°C and 80%, respectively. The advertisement call (Fig. 5; n = 1 male ; CHUNB 73649; n = 16 calls) consists of 8–14 (11.06 ± 1.57) multipulsed notes. Each note is composed of 4–18 (7.06 ± 3.06) pulses that are increasingly modulated within each note, with the amplitude suddenly falling at the beginning of the contiguous note. Single pulses (2.95 ± 0.98 ms) are sometimes isolated between notes, and also at the beginning and end of the call. Note duration is 26.70 ± 12.40 ms (14.0–76.0 ms) and they are separated by an interval of 1.0–12.0 ms (5.40 ± 2.74). Pulse duration is 1.96 ± 0.57 ms (1.0–3.0 ms), interval between pulses is 1.0–2.0 ms. Calls have a mean duration of 350 ± 30 ms (290–420 ms), with a call rate of 4–6 calls/min At the end of the call, the single pulses can be organized in groups of up to eight pulses (3.50 ± 2.39). The first and last notes are longer (first: 21.0–76.0 ms; last: 21.0–48.0 ms) than the intervening notes (14–25 ms), and are composed of 4–18 (11.33 ± 3.37) pulses (first note) and 6–14 (9.71 ± 2.15) pulses (last note), as opposed to the 4–8 pulses (5.52 ± 0.92) in the intervening notes. The call has two emphasized harmonics, with the fundamental frequency being also the dominant frequency, between 2067–2239 kHz (2085.66 ± 54.27 kHz), and the second between 3962.1–4478.9 kHz (3986.70 ± 246.83 kHz). Tadpole description. Tadpole description is based on five specimens (CFBH 38063) in Gosner stage 38, for which the morphological measurements are given separately below. Body robust, elongated and slightly ovoid in lateral view and elliptical in dorsal view (Fig. 6A −B), wider than high (BW/BH = 1.13–1.14); BH about 53.5% of BL (BH/BL = 0.53–0.54); BL about 38% of TL. Maximum body height in the middle of body length and maximum body width just behind the eyes. Snout rounded in dorsal, ventral and lateral views. Eyes relatively large (ED/BL = 0.16–0.17), dorsolateral, and not visible in ventral view (Fig. 6C). IOD about 81% of BW (IO/BW = 0.79–0.82). Nostrils rounded, small (diameter about 22% of ED), dorsolateral, located nearer to the eyes than the tip of snout (ESD/NSD = 0.76–0.97). NSD about 11% of body length (NSD/BL = 0.10–0.12). Narial opening with a pigmented marginal rim (Fig. 6A–B). Oral disc ventral, not emarginated, width about 56% of BW (ODW/BW = 0.55–0.57). Marginal papillae small, homogeneous in size, conical, with pointed tip; arranged as a simple row around the oral disc; dorsal gap about 9.0–9.8% of BW. Many submarginal papillae conical, scattered, located in the lateral portion of the angular and posterior regions, reaching the posterior portion of P3 (Fig. 6D–E). Large conical papilla on each side of the internal portion of the angular region. Labial tooth row formula (LTRF): 2(2)/3 or 2(2)/3(1); A1>A2; A2 with a very narrow gap in the medial portion; P1, P2, and P3 similar in size; P3 regular, not modified as a labial arm. In one individual, the P1 is interrupted medially. Jaw sheaths densely pigmented, serrated; upper jaw-sheath archshaped, with medial projection and long lateral process; lower jaw-sheath V-shaped (Fig. 6E). Behind the lower jaw-sheath, there is a colored keratinized spur on each side. Spiracle single, sinistral, short, located at the middle of the body, below the body midline, oriented posterodorsally, with inner wall fused to body wall, except for its posterior margin, and external wall shorter than inner wall; spiracle opening elliptical. Vent tube dextral, as large as wide, positioned above the margin of the ventral fin; vent tube opening elliptical and directed downward. The intestinal mass is positioned at a right angle to the longitudinal body axis. Tail elongated (TL 51.9–68.1% longer than BL), with pointed tip. Tail musculature moderately developed (TMH/MTH = 0.43–0.46). Dorsal fin continuous, originating at posterior third of body; ventral fin origin at posteroventral-most edge of the body; both fins tapering to the tip of the tail in the last half; dorsal fin slightly higher that the ventral fin (DFH/VFH = 1.20–1.26). The lateral line is visible as a series of elliptical, vertical whitish stitches. The V-shaped supraorbital line includes 17−21 stitches that diverge medially on the head between the nares and continues anteroventrally onto the snout. The postorbital series consist of a short dorsolateral row that includes 6−8 stitches. Tadpole color. In life, body dorsally yellowish brown, marbled with irregular black blotches; two golden interrupted dorsolateral stripes, from the tip of snout to the body-tail junction, both delimited above by a narrow black stripe, which extends from the nostrils to the tail tip, along the basis of the dorsal tail. A golden elliptical blotch on snout, marbled laterally with irregular black blotches on the dorsal half of the body (Fig. 7C). Iris black with a wide golden ring around the pupil. Venter white and translucent (intestinal mass is visible). Tail musculature with some scattered brown blotches and a vermiculated dark brown stripe between the epaxial and hypaxial musculature extending from the tail-body junction to tail tip. Dorsal and ventral fins with dark blotches and some thin, dark reticulations near the free margins of both fins. FIGURE. 6. Tadpole of Scinax rupestris sp. nov. (CFBH 38063, stage 38. TL = 29.63 mm ). A: Dorsal view. B: Lateral view. C: Ventral view. D and E: Oral disc. Photos A, B and C: M.R.C. Scale bar = 1 mm . In preservative, the color pattern is similar to that of living tadpoles, but fades and loses its golden and yellowish tones. The venter is totally translucent. The iris becomes black and the black tones become dark brown. Body dorsally translucent with small brown rounded spots, densely distributed and brown blotches; the black and golden stripes disappear. Measurements ( n = 5, mean ± standard error, range into parenthesis). TL 27.10 ± 2.24 (24.3–29.6); BL 10.21 ± 0.62 (9.5–11.1); TAL 16.89 ± 1.91 (14.5–18.6); MTH 6.37 ± 0.71 (5.6–7.2); TMH 2.85 ± 0.19 (2.6–3.1); BH 5.46 ± 0.32 (5.1–5.9); BW 6.19 ± 0.34 (5.8–6.6); ED 1.65 ± 0.10 (1.6–1.8); ODW 3.44 ± 0.23 (3.1–3.8); END 0.99 ± 0.02 (1.0–1.02); NSD 1.15 ±1.37 (1.0–1.4); ND 0.37 ± 0.02 (0.3–0.4); TMW 2.78 ± 0.11 (2.6–2.9); IND 2.97 ± 0.18 (2.7–3.1); IOD 4.92 ± 0.4 (4.5–5.4); DFH 2.29 ± 0.35 (1.9–2.8); VFH 1.90 ± 0.29 (1.6–2.2). Variation among tadpoles. There are no evident morphological variations in the oral disc, marginal and submarginal papillae, and vent tube in the different development stages examined. LTRF is either 2(2)/3 ( 20 specimens ) or 2(2)/3(1) ( 16 specimens ). In addition to the submarginal papillae on the lateral portion of the angular and infraangular regions, some individuals also present submarginal papillae anteriorly, immediately below the marginal papillae, near A1 (two specimens). Some individuals show loss of labial teeth (four specimens), and damage in the papillae (one specimen). One individual (stage 30) shows P1 (half of its length) and P3 (one third of its length) reduced, and P2 taking place on one side of P3. Most color variation among larvae involves the density of dorsal pigmentation, but some individuals show the vermiculated stripe replaced by brown blotches in the first two thirds of the tail musculature, or a straight stripe (less common), instead of the vermiculated one. In other individuals, the vermiculated stripe can be either interrupted, incomplete or absent; in this latter case, it is replaced by some blotches and many small spots until the tail tip. The oral disc can be pigmented near the marginal papillae (eight specimens), and in the submarginal papillae (five specimens). The margin of the vent tube can also be pigmented (two specimens). Furthermore, we detected some injuries in the tails of tadpoles of Scinax rupestris sp. nov. (six specimens; in the developmental stages 25 to 28, 34 and 36) at several stages of regeneration. Natural history and Geographic distribution. Scinax rupestris sp. nov. is an inhabitant of high altitude temporary rock-bed rivulets and creeks in the quartzitic rock mountains in Chapada dos Veadeiros region, northern state of Goiás, Central Brazil . This species has not been observed calling from vegetation (the individual of Fig. 4C was jumping away from observer and stopped on a clump of grass, where it was photographed), but males typically call from rocks along these streams. When disturbed, the individuals stop calling and shelter under rocks or rock crevices, exhibiting high ability for climbing on rock surfaces. The dorsal coloration confers camouflage to the individuals against rock surfaces. Although the males call several meters from each other, some small aggregations can be observed close to larger rivulets pools. Tadpoles are apparently diurnal. Its coloration is similar to that of the quartzitic rocks on rivulets beds that they inhabit (Fig. 7C). These rivulets and creeks are located on open habitats, and the sunlight can reach the rivulet bed. The golden blotches and stripes of the tadpole reflect the light in similar way that quartzitic sand, pebbles and rocks. The new species shares the same habitats in Chapada dos Veadeiros with Ameerega flavopicta (Dendrobatidae) , Bokermannohyla pseudopseudis (Hylidae) , Leptodactylus tapiti (Leptodactylidae) , Proceratophrys goyana and P. salvatori (Odontophrynidae) . One individual of Scinax rupestris sp. nov. was found in the stomach of B. pseudopseudis .