Bioacoustics of Tachiramantis lassoalcalai

Description of the holotype : an adult female, of 26.2 mm of SVL. Body slender; head slightly longer than wide, HW 38.16% of SVL. Snout subacuminate in profile ( Fig. 3 a), subacuminate in dorsal view ( Fig. 3 b); EN slightly shorter than ED; nostrils non- protuberant, directed dorsolaterally; canthus rostralis distinct, rounded; loreal region slightly concave. Upper eyelid without tubercles or warts. Cranial crests absent. Tympanum distinct, 33.3% of ED, surrounded by a tympanic annulus, with a supratympanic fold covering a small portion of its posterodorsal section. Choanae small, round, not concealed by palatal shelf of maxillary arch; vomerine dentigerous processes small, slightly oblique, bearing 4 teeth each, posterior and medial to choanae. Tongue round, slightly notched posteriorly, posterior one third free. Dorsal skin shagreen ( Fig. 3 c); one large tubercle posteroventral to tympanum present; occipital ridges poorly-defined; middorsal raphe only present, but difficultly detectable, on the head; dorsolateral folds present on the anterior half of the body, low and indistinct. Throat and chest smooth, belly and ventral surfaces of thighs areolate ( Fig. 3 d); posterior surfaces smooth. Ulnar, tarsal, and heel tubercles absent. Relative lengths of adpressed fingers III>IV>II>I; when adpressed, Finger I not reaching disk on Finger II. Finger disks much broader than long, disk on Finger III twice width of adjacent phalanx; oval except on Finger II, which is rounded; disk on thumb round, not distinctly expanded. Finger II disk on right hand 1.5 times wider than adjacent phalanx. Lateral fringes on fingers absent; webbing absent. Bifid palmar tubercle, and ovoid thenar tubercles low, indistinct; subarticular tubercles protuberant, single, round; supernumerary tubercles protuberant, in rows under each finger ( Fig 3 e). Hind limbs moderately short; shank 49.6% of SVL. Relative lengths of appressed toes IV>V>III>II>I. Disk on Toe IV slightly smaller than disk on Finger III. Toes without lateral fringes, unwebbed. Disks slightly wider than long, wider than phalanges, horizontally oval, except on Toe I, which is round. Inner metatarsal tubercle oval, indistinct; outer small, protuberant, round; subarticular tubercles protuberant, single, round; four supernumerary tubercles distinguishable under left plant ( Fig 3 f). Colour in life ( Fig 2a ). Dorsum golden brown with small scattered irregular dark brown spots; head slightly lighter, especially over the external borders of the upper eyelids and snout; with two dark brown stripes, one interorbital, another transversal irregular black bar anterior to the upper eyelids; dark brown canthal and supratympanic stripes; a narrow labial stripe interrupted by lighter transverse intersections. Ill defined dark brown transverse bars on anterior limbs; on hind limbs little more defined over a light grey background; hidden surfaces of hind limbs grey with small round white spots surrounded by dark brown. Throat light grey, chest belly and inferior part of limbs light grey spotted with white. Iris metallic golden with greenish tonalities and finely black reticulated. Colour in preservative . Dorsum grey, with small black spots irregularly spread along dorsum; on the head, an irregular black, broken interorbital bar, and another transversal irregular black bar anterior to the upper eyelids ( Fig. 3 b). Canthal stripe black, wide, covering nare; supratympanic stripe black, covering upper half of tympanum and turning behind tympanum towards the upper arm; a narrow longitudinal labial grey stripe on each side. Transverse dark bars on hind limbs only on shanks and tarsi. Anterior surface of thighs with pale spots surrounded by dark grey. Throat grey, chest, belly and ventral surfaces of arms dark grey with white marbling ( Fig 3 d); downsides of shanks and tarsi dark grey with white round little spots. Measurements of holotype (in mm): SVL: 26.2; ShL 13; HeL: 10.3; HW: 10; InD: 2.4; EN: 3; ED: 3.6; TD: 1.2; ETS: 4.7; FD: 1.2; T4D: 1; 1FiL: 3; 2FiL: 3.8. Variation: Females are consistent in the majority of characters with the female holotype . Some variation can be seen in the shape of the middorsal raphe (distinct on MHNLS 18460, 18894, 18895, present but indistinct on MHNLS 18877, 18893, 18897). The two dorsolateral folds are present on all females although they are low and ill-defined; they are completely indistinct on 18460 due to its bad preservation state. There also is consistently present a tubercle posteroventral to the tympanum. Small calcars are present at least in two specimens photographed in life (a male MHNLS 18878, Fig. 2a ; a female MHNLS 18877, Fig. 2 b) but not appreciable in preservative; small ulnar and tarsal tubercles are also appreciable in the male MHNLS 18878 ( Fig 2a ), but are indistinct in preservative. Appendix II shows the dorsal pattern variation of the type series. Males dorsally can be dark grey (MHNLS 18878), pale grey (MHNLS 18879, 18873–74), to bronze (MHNLS 18876), with transverse bars on the limbs, a W postoccipital mark, irregular spotting on dorsum, and a well defined black interorbital bar. The paler specimens (MHNLS 18879, 18873–74) have a uniform dorsum, with no spotting at all, except a few small irregular marks on the head of MHNLS 18876. The most appreciable variation is on ventral pattern and disposition of spots on the groin and on the hind limbs ( Fig. 4 a–d). On all females but MHNLS 18894 the belly is patterned with dark grey marbled with white large mostly round spots. On MHNLS 18894 the pattern is discernible but paler. On females MHNLS 18460 and 18894 there are many pale spots surrounded by black on the groin ( Fig 4 a), anterior and posterior surfaces of the thighs, as is characteristic for the species in the lentiginosus group of Rivero (1988). Variation on the belly pattern is as follows: MHNLS 18876 is the paler, with an almost unpatterned belly ( Fig 4 b), MHNLS 18873 and 18874 have a pale reticulation; MHNLS 18879 has a spotted belly, without marbled or reticulation; MHNLS 18900 has a mostly white belly with irregularly scattered small round spots; and MHNLS 18878 has a dark grey belly with a few whitish small spots; the most patterned belly is on MHNLS 18877 ( Fig. 4 c). Males accord with all characters with females, and there is not a clear sexual dimorphism in pattern. The male MHNLS 18878 has also a profusion of white round spots on the groin and posterior side of the thighs ( Fig.4 d). Sexual dimorphism is only evident by the smaller size, presence of vocal slits and single white nuptial pads on males. Natural history: One of the localities at Cerro Las Antenas corresponds to a section of a narrow rapid creek at 1780 m , surrounded by a dense primary cloud forest. At this site 18 specimens were collected and two more observed during the expedition of March 2008 (end of the dry season); all were found on different bush leaves in the creekside foliage, rocks in the creek’s bed, and in sympatry with Cryptobatrachus remotus Infante-Rivero, Rojas-Runjaic & Barrio-Amorós, 2008 , Cochranella sp., and Hyalinobatrachium tatayoi Castroviejo-Fischer, Ayarzagüena & Vila, 2007 . The other locality is a secondary low forest, at the summit of Cerro Las Antenas, at 1933 m , with a high density of arboreal ferns, vines and Cecropia . At this site Pristimantis lassoalcalai was found in sympatry with P. rivasi sp. nov. During the expedition of March 2008 many males were heard vocalizing profusely between 1600 and 1900 h, from leaves, usually exposed. All specimens seen or collected were at less than 1 m over the ground; several called from dry folds of Cecropia leaves. After 1900h the call activity declined abruptly and only a few males called sporadically until 2100h. During July 2008 many vocalizing males were heard calling profusely in the early morning hours (around 0800h) in a clear day near the summit of Cerro Las Antenas. The vocalization of P. lassoalcalai consists of single notes emitted every few seconds to every few minutes. The call remains a lament and could be described onomatopoeically as “nheek, nheek, nheek”.

evolutionary relationships of Tachiramantis within Craugastoridae are yet unclear as its position varied in all previous phylogenetic hypotheses in which samples of this genus were included ( Heinicke et al . 2015 ; Guayasamin et al . 2017 ; Heinicke et al . 2018 ). However, all of these hypotheses were congruent in recovering Tachiramantis as monophyletic, not closely related to Pristimantis , and deeply divergent to its sister linage. Our phylogeny ( Fig. 2 ) also differs from previous hypotheses with regard to the intergeneric relationships of Tachiramantis with other craugastorids but it is concordant with the results of Heinicke et al . (2015) and Guayasamin et al . (2017) in recovering the three species of Tachiramantis as closely related, with T. lentiginosus and T. douglasi as sister species, and T. prolixodiscus as the most external of the genus. The only difference with these two previous hypotheses corresponds to the position of the newly included P. lassoalcalai , which is well-nested within Tachiramantis as sister to T. lentiginosus + T. douglasi (rendering Tachiramantis paraphyletic). This newly discovered phylogenetic position confirms previous suspicions of a close evolutionary relationship between this species and Tachiramantis based on its overall morphological resemblance with T. lentiginosus and T. douglasi . Based on this result and in order to promote a taxonomy based on monophyletic groups, we transfer Pristimantis lassoalcalai Barrio-Amorós, Rojas-Runjaic & Barros, 2010 to Tachiramantis as Tachiramantis lassoalcalai comb. nov. With the new addition, Tachiramantis is now composed by four species distributed on the northern part of the Cordillera Oriental of Colombia , Tamá massif, Sierra de Perijá and Cordillera de Mérida. Tachiramantis remains a poorly known genus, distributed in a region undersampled by systematic studies using molecular data ( Heinicke et al . 2015 ) and with vast areas poorly inventoried for amphibians, as the Sierra de Perijá ( Rojas-Runjaic et al . 2011 ; Meza-Joya 2016 ; Rojas-Runjaic et al . 2018 ) and the Tamá massif ( Acevedo et al . 2014 ; Meza-Joya et al . 2019 ). Thus, we expect that the inclusion of both new and already described terraranas currently allocated to Pristimantis , in future molecular phylogenies, will result in the discovery of additional members of Tachiramantis . On the other hand, although no external morphology synapomorphies are known for Tachiramantis , the occurrence of pale spots on the posterior surface of thighs ( Rivero 1984 ) is shared by T. douglasi , T. lassoalcalai , and T. lentiginosus (absent in T. prolixodiscus ). Thus, this character state can be useful to diagnose these three species from other sympatric terraranas by external morphology. The phylogenetic relevance of this phenotypic character should be objectively assessed. In addition, osteological characterization of T. lassoalcalai will be required in order to corroborate the occurrence in this species of the putative cranial synapomophies defined by Heinicke et al . (2015) for Tachiramantis . Taxa 1 2 3 4 5 6 7 8 9 10 1. T. prolixodiscus SBH 268371 - 2. T. douglasi JDL 26116 6.4 - 3. T. douglasi AJC 3486 6.4 0.4 - 4. T. douglasi AJC 3472 6.4 0.4 0.0 - 5. T. douglasi AJC 3492 6.4 0.4 0.0 0.0 - 6. T. lentiginosus CVULA 9100 7.2 5.3 5.5 5.5 5.5 - 7. T. lassoalcalai MHNLS 18873 5.1 4.5 4.1 4.1 4.1 5.6 - 8. T. lassoalcalai MHNLS 18874 5.1 4.5 4.1 4.1 4.1 5.6 0.0 - 9. T. lassoalcalai MHNLS 18892 5.1 4.5 4.1 4.1 4.1 5.6 0.0 0.0 - 10. T. lassoalcalai MHNLS 18893 5.1 4.5 4.1 4.1 4.1 5.6 0.0 0.0 0.0 - As it is known, acoustic signals play a critical role in mate recognition and mate choice, and in most cases are species-specific ( Wells 2007 ), which make them a useful source of evidence for anuran systematics and taxonomy (Khöler et al . 2017). Until very recently there was no bioacoustic information available for any Tachiramantis , but the almost simultaneous publication of the call description of T. douglasi ( Mendoza-Roldán et al . 2020 ) with our description of the advertisement call of T. lassoalcalai allow us to confirm that bioacoustics also constitutes a valuable tool to diagnose species in this group. These two species exhibit strikingly different advertisement calls (values for T. lassoalcalai in parentheses): T. douglasi calls last ca. 1.7 s and consist of trills of 7–9 notes, with the first being the longest (calls represented by single notes of ca. 0.7 s ), with dominant frequency of ca. 5.1 kHz (dominant frequency between 2.9–3.0 kHz). Considering that the known geographic distributions of T. lassoalcalai and T. douglasi are relatively close ( Mendoza-Roldán et al . 2020 ; Barrio-Amorós et al . 2010 ) and that both species are morphologically similar (see Fig. 1 ), their bioacoustical characterizations will be valuable to diagnose them during fieldwork and in the absence of molecular data.