Bioacoustics of Ranitomeya benedicta

Ranitomeya benedicta can be distinguished from other species of Ranitomeya by the combination of a relatively large adult SVL and a characteristic red head. Ranitomeya reticulata also possesses red coloration on the head, although this coloration extends well onto the dorsum whereas in R. benedicta the red coloration is confined to the head only. Furthermore, R. reticulata is smaller than R. benedicta (SVL 13.7–16.6 mm vs. 15.0– 20.2 mm ). Ranitomeya benedicta is most similar in external morphology to R. fantastica and the second new species, R. summersi , described below. Ranitomeya benedicta has comparatively shorter hind limbs, with a significantly shorter tibia than R. fantastica (mean tibia length/SVL = 0.444 in R. benedicta , 0.486 in R. fantastica , P = 0.01, Fig. 2 ). Certain color morphs of R. fantastica have head coloration approaching red (i.e. bright orange), but never a true red as in R. benedicta . Furthermore, the call of R. benedicta is much louder (audible from ~ 8 m vs. ~ 3 m in R. fantastica ), decreases in tone in consecutive notes (vs. constant tone across notes in R. fantastica ) and has shorter pulses than the call of R. fantastica (150 ± 20 ms vs. 290 ± 30 ms in R. fantastica , Fig. 3 ). For differences between R. benedicta and R. summersi , see below. Ranitomeya benedicta is distinguished from its closest relatives on the basis of 8 unique unambiguous mitochondrial gene nucleotide site substitutions. Measurements (in mm) of holotype . The undissected holotype ( Fig. 4 ) is an adult female as shown by its relatively large size and lack of vocal slits. SVL 17.2; FL 8.1; TL 8.3; KK 15.1; FoL 7.3; HaL 5.0; HL 5.0; HW 5.5; BW 5.5; UEW 2.57; IOD 2.50; IND 2.21; TD 0.75; ED 2.14; DET 0.50; L1F 1.86; L2F 2.86; W3D 0.96; W3F 0.43. Measurements of additional specimens are given in the appendix ( Table 1 ). Description of holotype . Widest part of head is at jaw articulations. Head width as wide as body. Tongue ovoid; teeth absent. In life, head bright red with comma-shaped black spots over the eyes and tympanum. Red coloration extends posteriorly to level of the axillae; black ground color with blue reticulation on dorsal body. Limbs and digits black, limbs reticulated with blue. Underside of head red, lacking paired gular spots. Venter black with discrete blue reticulation. Nares black surrounded by black spots. Iris black. In life, skin texture nearly smooth on the dorsal surfaces of the body and head; limbs and rump weakly granular. Venter weakly granular on limbs and body, ventral surface of head nearly smooth. Snout sloping and rounded in lateral profile, round or slightly blunted in dorsal profile. Nares situated at tip of snout and directed laterally; both nares visible from ventral and anterior view but not from dorsal view. Canthus rostralis rounded, loreal region flat and nearly vertical. Upper eyelid approximately equal in width to interorbital distance; internarial distance roughly equal to eye width. Tympanum round, partially concealed posterodorsally. Hands relatively large, length 29 % of SVL. Relative length of appressed fingers III> IV ≈ II> I; first finger 65 % length of second; finger discs moderately expanded, width of disc on finger III 2.2 times width of adjacent phalanx. An unpigmented median metacarpal tubercle is present on base of palm; inner metacarpal tubercle present near base of finger I but angled posteriorly; unpigmented proximal subarticular tubercles present on base of each digit, except on finger I, where tubercle is part-way up the digit; distal subarticular tubercle visible only on fingers III and IV. All tubercles raised above level of hands; scutes present on dorsal surface of fingers. Hind limbs moderate length, with heel of appressed hind limbs reaching level of eye. Femur and tibia roughly equal in length, tibia 102 % length of femur; knee-knee distance 88 % of SVL. Relative lengths of appressed toes IV> V ≈ III> II> I; first toe short with unexpanded disc; second toe with slightly expanded disc, discs on toes III–V moderately expanded. Two unpigmented metatarsal tubercles present on base of foot, one situated medially near base of toe I, the other situated laterally at the base of the fifth metatarsal. Proximal subarticular tubercles present at base of each toe but most notable on toes I and II due to their lack of pigmentation. Toes III and V with two subarticular tubercles, toe IV with three subarticular tubercles. A tarsal keel is present starting below the knee and turning into the medial metatarsal tubercle at the foot. Tarsal tubercle absent; feet and hands lacking webbing and lateral fringing. Variation. Adults 15.0– 20.2 mm SVL, females slightly larger than males: three adult males 15.0– 17.5 mm (mean 16.5 mm ); four adult females 16.8–20.2 mm (mean 18.4 mm ). Widest part of head is at jaw articulations. Head about as wide as body except in two apparently gravid females whose bodies are wider than the head. Head width 92 % of body width in three males (range 80–106 %), 94 % percent of body width in four females (range 74–121 %). Head width 27–34 % of SVL in adults. No apparent sexual dimorphism in external morphology except that males are smaller, possess faint vocal slits on the floor of the mouth, and have a slightly expanded subgular pouch. Live animals bear a diagnostic scarlet-red head with black spots over the eyes. These spots are medially fused in some individuals and form a W-shaped head crest which descends over the eyes creating a face-mask; in most individuals this mask also covers the tympanum. The red coloration on the head extends posterior to level of the axillae; from here the dorsal body coloration changes to a black ground color with blue reticulation. This reticulation is diffuse in some individuals, sometimes to the point of the body appearing uniform blue on the rump. Limbs and digits are colored as the dorsum: individuals with defined dorsal reticulation have limbs reticulated with blue; individuals with diffuse reticulation on the body have either diffuse limb reticulation or limbs colored uniformly blue. Underside of head red, most individuals possessing some amount of black marbling on chin. Venter black with diffuse or discrete blue reticulation. Nares black and surrounded by black spots, these spots are sometimes fused to form a bar across the loreal region. Hands relatively large, length 24–29 % of SVL. First finger 56–81 % length of second; finger discs moderately expanded in both males and females, width of disc on finger III 2 –2.3 times width of adjacent phalanx. Tibia 85–115 % length of femur (mean 99 %); knee-knee distance 81–93 % of SVL (mean 87 %). In preservative, red coloration turning pale pink and blue reticulation turning purple-grey. Tadpole. One tadpole (captive bred from topotypic parents) was used for the mouthpart description. The body of the tadpole was destroyed after preservation making a full description impossible, but the mouthparts remained intact. Oral disc emarginate, anterior and posterior labia forming flaps free from body wall. Marginal papillae absent on anterior labium except for lateral-most portion (3–4 papillae present), present in one complete row on posterior labium. Papillae white, rounded; submarginal papillae absent. Jaw sheaths deep in longitudinal width, serrate, lacking indentations. Lateral processes short, extending barely past lower jaw. Labial tooth row formula is 2(2)/3. A-1 complete, A-2 with medial gap, same width as A-1. P-1, P-2, and P-3 complete; P-1 and P-2 equal width, P-3 slightly shorter. We examined two R. fantastica tadpoles from Tarapoto and the upper Cainarachi Valley (stages 27 and 29) and the mouthparts were indistinguishable from the single R. benedicta tadpole. Vocalizations. The advertisement call is a series of buzz-like notes ( Fig. 3 ). The dominant frequency of the call starts at 4240 Hz and gradually decreases in subsequent notes to 3190 Hz (air temperature 24° C). The notes in the calls were very short with a duration of 150 ± 20 ms (vs. 290 ± 30 ms in R. fantastica and 440 ± 30 ms in R. summersi ); brief pauses between each note lasting 140 ± 20 ms (vs. 200 ± 20 ms in R. fantastica and 150 ± 20 ms in R. summersi ). The ratio of note length to pause length is approximately 1:1 (vs 1.4: 1 in R. fantastica and 2.8: 1 in R. summersi ). The call can be heard from as far as ~8 meters which is comparatively loud for a member of the fantastica group (vs. 3 m in R. fantastica and 1 m in R. summersi ).

Distribution and natural history. This frog appears to be widely distributed throughout the lowland forest of the Pampas del Sacramento in southern Loreto and eastern San Martin ( Fig. 5 ). The Pampas del Sacramento are enclosed by Rio Ucayali to the east, Cordillera Azul and Rio Huallaga to the west and northwest, and the flooded forests of Pacaya-Samiria to the north. Ranitomeya benedicta appears to occur throughout this region in forests over ca. 150 m elevation. The southern margin of its range likely extends at least to Rio Cushabatay, but not reaching Pucallpa, as frequent surveys along the Aguaytia-Pucallpa road since 2006 have failed to encounter any members of the fantastica group. Ranitomeya benedicta has never been observed west of Rio Huallaga or east of Rio Ucayali, and both rivers appear to be boundaries restricting its westward and eastward distribution, respectively. The Ucayali and the lower Huallaga are generally surrounded by 10–50 km-wide flood plains and wetlands; areas which are inhospitable to most poison frogs and may be more effective barriers than the rivers themselves. These frogs are most frequently encountered amidst the tangled branches in or near tree-falls. They are extremely alert and agile, and when encountered, immediately flee to the ground and disappear amidst the leaf litter, jumping in erratic spurts. These frogs are rarely encountered and often after days of searching we were only able to find one or two individuals. Their distribution appears to be patchy but in some areas locally abundant. This species appears to favor slightly elevated areas within the forest and have not been found areas susceptible seasonal flooding. Although R. benedicta is primarily a lowland species, individuals have been found in higher elevation areas in the Huallaga Canyon at 315 m and in the upper Cushabatay at 405 m (M. Ramírez Zárate pers. comm.). These records may represent the upper altitudinal limits of this species, although further exploration of the Cordillera Azul may uncover additional montane localities. In the northwestern extent of its distribution, R. benedicta is sympatric with R. imitator and R. ventrimaculata . Reproduction occurs terrestrially, with clutches of 4–6 grey eggs being laid amongst humid leaves accumulated on the forest floor. Egg development lasts 12–16 days ( MSR , pers. obsv.). Tadpoles are transported to larger phytotelmata, such as Aechemea bromeliads, and (in captivity) metamorphose in about 100 days ( MSR , pers. obsv.). Due to the low phytotelmata abundance in the forests of the Pampas del Sacramento and the observation that R. ventrimaculata are the tadpoles most commonly encountered, there is likely strong competition for phytotelmata access, and phytotelmata abundance may be a major limiting factor on population sizes in R. benedicta ( Donnelly, 1989 ) . Aside from terrestrial reproduction and foraging, these frogs appear to be arboreal, which could partially explain why they are so difficult to find. Near the village of Shucushuyacu, farmers felling large trees frequently encounter this frog fleeing from the large canopy bromeliads (i.e. Aechemea ). Timber harvesters along the Rio Ucayali have also reported the same observation. These frogs also are extremely tolerant of heat. We observed one frog carrying tadpoles across a large, deforested area in full, midday sun (36° C) and was seemingly unaffected. Conservation status. Under the IUCN Red List criteria (IUCN, 2001), we suggest R. benedicta be listed as Vulnerable (VU) under the following criteria (1) we estimate its extent of occurrence at 19,000 km 2 (although the area of occupancy may be much lower), and ongoing deforestation along the lower Huallaga will reduce the amount of suitable habitat substantially over the coming years, (2) this species is found primarily in undisturbed rainforest and does not occur in areas disturbed by human activity, (3) populations appear to be patchily distributed, and (4) there will be a high demand for this species in the pet trade. This species does occur in a protected area (Cordillera Azul National Park), although the extent to which it occurs there is unknown. We fear that by describing this species (and others), we are also putting them at risk of illegal smuggling, and expect that there will be a large influx of illegally acquired R. benedicta to the North American and European markets over the next few years. This is particularly frustrating for the authors who try to minimize these negative impacts through sustainable harvest and legal exportation (i.e. Understory Enterprises). This organization is concerned with preserving species in their native habitats, while sustainably harvesting individuals for the pet trade. We encourage anyone interested in owning any poison frog to exercise restraint and purchase only sustainably-harvested or captive-bred frogs of legal origin. Further, consumers must understand that when these projects are done correctly; they take a considerable amount of time and money. When a species is illegally smuggled, it reduces the potential market for the legal business, and as a result, reduces the amount of money these organizations can generate for land protection. Ranitomeya summersi , sp. nov. Figures 1 , 5 , 6 , 10

Ranitomeya summersi can be distinguished from other species of Ranitomeya by the combination of relatively large adult SVL, distinctly bicolored pattern consisting of continuous bands (as opposed to spots or dashes), and a buzz-call. This species is most similar in external morphology to R. fantastica and some populations of R. imitator . Ranitomeya summersi can be distinguished from R. fantastica using the following characteristics: shorter hind limbs, with a significantly smaller tibia/SVL ratio (mean tibia length/SVL = 0.458 in R. summersi , 0.486 in R. fantastica , P = 0.044; Fig. 2 ); the call of R. summersi has longer pulses than the call of R. fantastica (440 ± 30 ms vs. 290 ± 30 ms in R. fantastica , Fig. 3 ); and the ratio of note length to pause length is 2.8:1 (vs 1.4: 1 in R. fantastica ). All known R. summersi individuals possess a bicolored pattern (orange and black), whereas R. fantastica typically are tricolored (orange, black, and blue or white) and have reticulated leg netting (see Fig. 8–10 ). Ranitomeya summersi can be distinguished from R. benedicta by adult coloration; R. benedicta has a red head with blue dorsal and ventral reticulation (compared to the bicolored pattern in R. summersi as described above). Ranitomeya summersi is considered a Müllerian mimic of some sympatric populations of R. imitator ( Symula et al. , 2001 applying the name Dendrobates fantasticus ) and can be distinguished from these populations by its soft ‘buzz’ call, audible from less than 1 m (vs. loud ‘trill’ call, audible from greater than 5 m in R. imitator ). In our experience, the black eye-mask of R. imitator usually does not cover the tympanum, and the black head spot of R. imitator is ovoid, whereas the head spot in R. summersi is pentagonal in shape. Ranitomeya summersi usually has paired black gular spots, rarely present in R. imitator . This species can be distinguished from its sister taxon, R. fantastica , on the basis of 16 unique unambiguous mitochondrial gene nucleotide site substitutions, and can be distinguished from R. benedicta on the basis of 20 unique unambiguous mitochondrial gene site substitutions, and is remarkably distant, genetically, to R. imitator (see Symula et al., 2001 ). Measurements (in mm) of holotype . The undissected holotype ( Fig. 6 ) is an adult male as determined by the presence of vocal slits. SVL 19.5; FL 8.1; TL 8.5; KK 16.7; FoL 8.0; HaL 5.5; HL 5.3; HW 6.3; BW 7.3; UEW 2.86; IOD 1.93; IND 2.29; TD 1.00; ED 2.07; DET 0.50; L1F I 2.07; L2F 3.14; W3D 0.96; W3F 0.36. Measurements of additional specimens are given in the appendix ( Table 1 ). Description of holotype . Widest part of head is at jaw articulations. Head narrower than body. Head width 86 % of body width. Subgular pouch not visibly expanded. Tongue grey, ovoid, attaching anteriorly; teeth absent. Pattern bicolored; black on the head, body, and limbs, highlighted by several golden orange bands. The head is black and orange, the black confined to the top of the head and around the eyes and tympanum. The black eye mask covers the tympanum and does not connect with the black on top of the head. Dorsum jet black with one incomplete orange cross-band near the sacral region; broken medially left of vertebrae. The hind limbs are black and lack reticulation, but possess a solid orange band extending from the rump over the femur, down across the anterior surface of the knee, back across the posterior surface of the tibia, and across the posterior surface of the tarsus to the foot. The dorsal surface of the foot is half orange and half black, toes orange. The front limbs are orange dorsally; this orange is fused with the orange on the face. Dorsal surfaces of the hands are both orange and black, fingers orange, finger tips black. Ventrally, the belly is black with irregular and coarse orange marbling. From a ventral aspect, the hind limbs have irregular orange longitudinal bands. Ventral surfaces of the forelimbs are uniformly black. The underside of the head is orange; two well-defined black gular spots laterally. Black bar present on the anterior surface of the snout between the nares. Nares black; iris black. In life, skin texture granular on the dorsal surfaces of the body and head; dorsal surfaces of limbs weakly granular. Venter weakly granular on belly; limbs and ventral surface of head nearly smooth. Snout sloping and rounded in lateral profile, rounded in dorsal profile. Nares situated at tip of snout and directed laterally; both nares visible from ventral and anterior view but hidden in dorsal view. Canthus rostralis rounded, loreal region slightly concave and vertical. Upper eyelid approximately equal in width to interorbital distance; internarial distance roughly equal to eye width. Tympanum round, partially concealed posterodorsally. Hands relatively large, length 28 % of SVL. Relative length of appressed fingers III> IV> II> I; first finger 67 % length of second; finger discs moderately expanded, width of disc on finger III 2.6 times width of adjacent phalanx. An unpigmented median metacarpal tubercle is present on base of palm; inner metacarpal tubercle present near base of finger I but angled posteriorly; unpigmented proximal subarticular tubercles present on base of each digit, except on finger I, where tubercle is part-way up the digit; distal subarticular tubercle visible only on fingers III and IV. All tubercles raised above level of hands; scutes present on dorsal surface of fingers. Hind limbs moderate length, with heel of appressed hind limbs reaching reaching to eye. Femur and tibia roughly equal in length, tibia 105 % length of femur; knee-knee distance 86 % of SVL. Relative lengths of appressed toes IV> III ≈ V> II> I; first toe short with unexpanded disc; second toe with slightly expanded disc, discs on toes III–V moderately expanded. Two metatarsal tubercles present on base of foot, one situated medially near base of toe I, the other situated laterally at the base of the fifth metatarsal. Proximal subarticular tubercles present at base of each toe but most notable on toes I and II due to their lack of pigmentation. Toes III and V with two pigmented subarticular tubercles, toe IV with three subarticular tubercles. A faint tarsal keel is present starting below the knee and turning into the medial metatarsal tubercle at the foot. Tarsal tubercle absent; feet and hands lacking webbing and lateral fringing. Variation. Adults range from 15.5–20.4 mm SVL, females and males roughly equal in SVL. Head width 86 % of body width in type series (range 66–127 %). Head width 30–33 % of SVL in adults. No apparent sexual dimorphism in external morphology except that males tend to be less rotund and possess vocal slits on the floor of the mouth. Subgular pouch not visibly expanded in males. The black eye mask invariably covers the tympanum and does not connect with the black on top of the head, but the head-black and eye-black can fuse with the black on the dorsum. Dorsum jet black with one orange cross-band near the sacral region. The cross-band is usually complete but in some individuals is broken medially. Some individuals have longitudinal extensions of the cross-band in line with the vertebrae, which can extend up the back, or posteriorly connecting with the orange bands of the hind limbs ( Fig. 10 , A– G). The dorsal surface of the foot is usually half orange and half black; toes can be either orange or black. Dorsal surfaces of the hands are both orange and black, fingers are usually either all orange or all black. The underside of the head is orange; most individuals have two well-defined black gular spots laterally, in some individuals these spots are medially fused. Most individuals in the type series possess a black bar on the anterior surface of the snout between the nares. Hands relatively large, length 26–31 % of SVL. First finger 66–76 % length of second; finger discs moderately expanded in both males and females, width of disc on finger III 2 –2.7 times width of adjacent phalanx. Femur and tibia roughly equal in length, tibia 100–107 % length of femur (mean 105 %); knee-knee distance 86–93 % of SVL (mean 90 %). In preservative little color change occurs, except that the orange coloration turns to pale gold or white. Tadpole . A stage 29 tadpole was used for the description ( Fig. 2 ). Total length 19.0 mm, body length 7.6 mm , body width 5.5 mm . Snout rounded when viewed from above; body ovoid in dorsal view. Eyes black, dorsal, angled laterally, pupils white in preservative. Nares not forming tube, situated half-way between eye and tip of snout, directed dorsolaterally. Spiracle sinistral; vent dextral. Tail 3.4 mm in depth measured halfway along length; ventral tail fin begins at tail base, dorsal tail fin begins just posterior to plane of vent opening, ventral and dorsal fins relatively uniform in thickness throughout tail, tapering towards tip. Musculature depth uniform throughout, tapering towards tip, musculature depth 2.0 mm. The mouth is directed anteroventrally. Oral disc emarginate, anterior and posterior labia forming flaps free from body wall, 2.1 mm in width. Marginal papillae absent on anterior labium except for lateral-most portion (3–4 papillae present), present in one complete row on posterior labium. Papillae white, rounded; submarginal papillae absent. Jaw sheaths deep in longitudinal width, serrate, lacking indentations. Lateral processes very short, extending barely past lower jaw. Labial tooth row formula is 2(2)/3. A-1 complete, A-2 with medial gap, same width as A-1. P-1, P-2, and P-3 complete; P-1 and P-2 equal width, P-3 slightly shorter. In preservative, the head appears light brown. Pigmentation on dorsum is mottled brown with a white ground color. Ventral coloration is white with irregular brown flecking which is most dense near the mouth. Tail musculature brown, fins transparent with brown mottling. Life colors grey. One additional stage 27 tadpole was examined and agrees with this description. Additionally, the above description agrees with two R. fantastica tadpoles (see above). Mouthparts are identical between these two species. It appears that tadpole morphology is consistent within the R. fantastica species group. Vocalizations. The advertisement call is a series of soft buzz-like notes, similar to R. fantastica ( Fig. 3 ). The notes in the calls are 440 ± 30 ms (vs. 290 ± 30 ms in R. fantastica ) in duration with brief pauses of 150 ± 20 ms between each note (vs. 200 ± 20 ms in R. fantastica ). The ratio of note length to pause length is 2.8:1 (vs 1.4: 1 in R. fantastica ). The call is audible from 1 meter (vs. 3 m in R. fantastica ). The dominant frequency of the call is 2890 Hz at an air temperature of 24° C (vs. 3290 Hz in R. fantastica at 24° C). The call of R. summersi can be distinguished from R. benedicta on the basis of the following parameters: audible from < 2 m (vs. up to 8 m in R. benedicta ); note length> 400 ms (vs. 150 ± 20 ms in R. benedicta ), and ratio of note length to pause length is> 2:1 (vs. 1: 1 in R. benedicta ). This call can be distinguished from R. imitator in that R. summersi has soft buzz-like notes audible from < 2m (vs. loud, trill-like call easily audible from> 5 m in R. imitator ).