Glassfrogs of Ecuador: Diversity, Evolution, and Conservation

Guayasamin, Juan M.; Cisneros-Heredia, Diego F.; McDiarmid, Roy W.; Peña, Paula; Hutter, Carl R.

doi:10.3390/d12060222

Open AccessArticle

Glassfrogs of Ecuador: Diversity, Evolution, and Conservation

by

Juan M. Guayasamin

^1,2,*,

Diego F. Cisneros-Heredia

^3,4,5

,

Roy W. McDiarmid

⁶,

Paula Peña

^7,8 and

Carl R. Hutter

^9,10

¹

Instituto BIÓSFERA-USFQ, Colegio de Ciencias Biológicas y Ambientales COCIBA, Laboratorio de Biología Evolutiva, Campus Cumbayá, Universidad San Francisco de Quito USFQ, Quito 170901, Ecuador

²

Department of Biology, University of North Carolina, Chapel Hill, NC 27599, USA

³

Museo de Zoología & Laboratorio de Zoología Terrestre, Instituto de Diversidad Biológica Tropical iBIOTROP, Colegio de Ciencias Biológicas y Ambientales COCIBA, Universidad San Francisco de Quito USFQ, Quito 170901, Ecuador

⁴

Department of Geography, King’s College, London WC2R 2LS, UK

⁵

Instituto Nacional de Biodiversidad INABIO, Quito 170135, Ecuador

⁶

USGS Patuxent Wildlife Research Center, National Museum of Natural History, Smithsonian Institution, P.O. Box 37012, Washington, DC 20013, USA

⁷

Ingeniería en Biodiversidad y Recursos Genéticos, Centro de Investigación de la Biodiversidad y Cambio Climático–BioCamb, Av. Machala y Sabanilla, Universidad Tecnológica Indoamérica, Quito 170103, Ecuador

⁸

Universidad Internacional Menéndez Pelayo, Isaac Peral, 23-28040 Madrid, Spain

⁹

Biodiversity Institute and Department of Ecology and Evolutionary Biology, University of Kansas, Lawrence, KS 66045–7561, USA

¹⁰

Museum of Natural Sciences and Department of Biological Sciences, Louisiana State University, Baton Rouge, LA 70803, USA

^*

Author to whom correspondence should be addressed.

Diversity 2020, 12(6), 222; https://doi.org/10.3390/d12060222

Received: 30 December 2019 / Revised: 20 April 2020 / Accepted: 6 May 2020 / Published: 2 June 2020

(This article belongs to the Special Issue Systematics and Conservation of Neotropical Amphibians and Reptiles)

Abstract

:

Glassfrogs (family: Centrolenidae) represent a fantastic radiation (~150 described species) of Neotropical anurans that originated in South America and dispersed into Central America. In this study, we review the systematics of Ecuadorian glassfrogs, providing species accounts of all 60 species, including three new species described herein. For all Ecuadorian species, we provide new information on the evolution, morphology, biology, conservation, and distribution. We present a new molecular phylogeny for Centrolenidae and address cryptic diversity within the family. We employ a candidate species system and designate 24 putative new species that require further study to determine their species status. We find that, in some cases, currently recognized species lack justification; specifically, we place Centrolene gemmata and Centrolene scirtetes under the synonymy of Centrolene lynchi; C. guanacarum and C. bacata under the synonymy of Centrolene sanchezi; Cochranella phryxa under the synonymy of Cochranella resplendens; and Hyalinobatrachium ruedai under the synonymy of Hyalinobatrachium munozorum. We also find that diversification patterns are mostly congruent with allopatric speciation, facilitated by barriers to gene flow (e.g., valleys, mountains, linearity of the Andes), and that niche conservatism is a dominant feature in the family. Conservation threats are diverse, but habitat destruction and climate change are of particular concern. The most imperiled glassfrogs in Ecuador are Centrolene buckleyi, C. charapita, C. geckoidea, C. medemi, C. pipilata, Cochranella mache, Nymphargus balionotus, N. manduriacu, N. megacheirus, and N. sucre, all of which are considered Critically Endangered. Lastly, we identify priority areas for glassfrog conservation in Ecuador.

Keywords:

anura; biogeography; centrolenidae; systematics; taxonomy

1. Introduction

The Neotropical family Centrolenidae Taylor 1951 is a monophyletic taxon that contains about 150 species classified into 12 genera [1] (Figure 1). Historical biogeographic evidence in a phylogenetic context suggests that the group originated in South America and subsequently dispersed to Central America [2,3]. Centrolenids share a unique morphology and behavior that makes them readily identifiable; some of the most evident traits include a green dorsum in most species (Figure 2), completely or partially translucent venter (Figure 3), out-of-water deposition of eggs along streams (Figure 4), and forward-directed eyes (Figure 2A). Other interesting features that have evolved within Centrolenidae include parental care [4] (Figure 5), fighting behavior [5] (Figure 6), and humeral spines in males of some species [6] (Figure 7).

The monophyly of Centrolenidae is well supported by morphological [7,8,9] and molecular [2,8,10,11,12,13,14] studies. Osteological characters shared by all glassfrogs (Figure 8) include a dilated medial process on Metacarpal III [7], T-shaped terminal phalanges [15], intercalary element between distal and penultimate phalanges [15], and complete or partial fusion of tibiale and fibulare [15,16]. The diversity of glassfrogs is growing and in constant revision (e.g., [1,6,17,18,19,20,21]). Centrolenid species richness is concentrated in mountain chains, where humidity is high, and streams provide suitable reproductive habitats. Therefore, it is no surprise that the Andes, by far, is the center of diversity and endemism for the group and that Colombia and Ecuador maintain the highest species richness.

In 1973, John D. Lynch and William E. Duellman [22] provided the first review of the glassfrogs of Ecuador, and they reported the presence of 20 species and suggested the occurrence of an additional one. Only 47 years later, the species richness of this family has tripled, reaching the incredible number of 60 species in Ecuador. Since Lynch and Duellman’s pioneering work, our understanding of centrolenid biology [23,24,25], morphology [6,9,17], systematics [1,6,17,26,27], biogeography [3,28], and evolution [2,19,25] has improved substantially. Still, there are novel challenges and conspicuous gaps that need to be filled.

Herein, we provide a new review of Ecuadorian glassfrogs, bringing an update of what is known about the group, highlighting issues pending resolution, and providing a framework that we hope will facilitate further research, particularly with species identification, discovery, and conservation. We employ a candidate species system (e.g., [29]) to identify putative undescribed species to be investigated and to streamline the species discovery process. These tasks are particularly urgent in the current global amphibian extinction phenomenon [30,31,32,33].

Figure 3. Ventral transparency in glassfrogs. (A) Complete transparency of parietal peritoneum and pericardium; H. aureoguttatum. (B) Partial transparency: parietal peritoneum is transparent only posteriorly; N. posadae. (C) Venter opaque (no ventral transparency): ventral parietal peritoneum and the urinary bladder peritoneum are opaque (white); N. gradisonae. Photos by Martín Bustamante.

Figure 4. Egg deposition sites in glassfrogs. (A) On upper side of leaves (C. sanchezi). (B) On tip of leave (N. wileyi). (C) On underside of leaves (H. cappellei, photo by C. Barrio-Amorós). (D) On the margin of underside of leaves (Teratohyla spinosa, photo by R. Puschendorf). (E) On rocks (e.g., Sachatamia albomaculata, photo by R. Puschendorf). Figure modified from Guayasamin et al. [1].

Figure 5. Parental care in glassfrogs. Adult male of Hyalinobatrachium aureoguttatum, with egg clutch. Photo by Luis A. Coloma.

Figure 6. Combat behavior in glassfrogs. Note diversity of positions. Illustrated species: Nymphargus grandisonae. Photos by Carl R. Hutter.

Figure 7. Humeral spine in Cochranella litoralis. Note that the humeral spine is formed by the prolongation of the crista ventralis. Photo by Luis A. Coloma; drawing by Linda Trueb.

Figure 8. Synapomorphies of Centrolenidae. (A) Partial (Nymphargus posadae, QCAZ 25090) and complete ventral transparency (Hyalinobatrachium aureoguttatum, QCAZ 32070). (B) Partial (N. wileyi, QCAZ 26029) and complete fusion between tibiale and fibulare (H. munozorum, KU 155497). (C) Presence of medial process on Metacarpal III and intercalary element (Teratohyla spinosa, KU 32935). The presence of T or Y-shaped terminal phalanges is a synapomorphy of Allocentroleniae (Allophrynidae + Centrolenidae). Photos in (A) by M. Bustamante. Modified from Guayasamin et al. [1].

2. Methods

2.1. Taxonomy and Species Concept

Throughout this work, we use the name Centrolenidae as originally intended by Taylor [15]. The taxonomic arrangement below the family level (i.e., subfamily and genus) follows the proposal by Guayasamin et al. [1]. We use quotations to denote species with uncertain phylogenetic position to differentiate them from monophyletic clades (e.g., “Centrolene”, “Cochranella”), as proposed by Guayasamin et al. [1]. An updated taxonomy for all the species in Centrolenidae is provided in Table S1.

For recognizing species, we adhered to the evolutionary species concept first proposed by Simpson [34,35], modified by Wiley [36] and de Queiroz [37,38]. This concept incorporates important theoretical factors such as lineage independence, identity, and evolutionary tendencies, and provides a flexible framework when reproductive isolation is difficult to test (e.g., allopatric populations). Evidence supporting the validity of a species can come from different sources (e.g., morphology, DNA, behavior, ecology), and no trait alone can be considered a biological property that a species must have in order to be recognized [37]. In other words, under the evolutionary species concept, the only necessary property for an entity to be a recognized as a species is that it corresponds to a temporal segment of a metapopulation lineage evolving separately from other lineages [37,38]. Evidence for independent evolution is gathered from different data sources, where integrative taxonomy plays a fundamental role when assessing what represents (or not) a distinctive species (e.g., [39,40]).

2.2. Characters and Terminology

For general terminology and descriptions of morphological characters we follow the proposals by Lynch and Duellman [22] and Cisneros-Heredia and McDiarmid [17]. We illustrate some of the most relevant traits for glassfrog identification, including dorsal color pattern (Figure 2), skin texture (Figure 9), and snout shape (Figure 10). To facilitate comparison with previous literature dealing with anurans, fingers are numbered preaxially to postaxially from I–IV. However, we stress that from an evolutionary perspective, anuran fingers should be numbered from II–V, to reflect the loss of Digit I in anurans [41,42,43]. Webbing formulae follow the method of Savage and Heyer [44], as modified by Guayasamin et al. [20] (Figure 11). Larval characters follow the terminology recommended by McDiarmid and Altig [45]. The morphology of nuptial excrescences and prepollical spines (Figure 12) follows the types proposed by Flores [46], with the additions and modifications detailed in Cisneros-Heredia and McDiarmid [17] and Guayasamin et al. [1]. Other key traits in centrolenid taxonomy are humeral spines (Figure 7, Figure 13, and Figure 14), ventral transparency (Figure 8), peritoneum and pericardium (with or without iridophores; Figure 13), enlarged subcloacal warts on thighs below vent (Figure 15), and tubercles on the external edge of arm, hand, and foot (Figure 16).

When discussing parental care, we adopt the terminology by Delia et al. [25]. Egg brooding refers to a specific form of ventral contact where the parent positions its body over the egg clutch; this behavior reduces embryonic mortality by protecting embryos from dehydration and, possibly, by preventing fungal development and predation [25]. Parental care is divided into the following behaviors [25]: (i) Short-term maternal care, where brooding is provided for a few hours just after oviposition; (ii) prolonged male care, where parental care is provided for several weeks; and (iii) prolonged female care, where parental care is provided for several weeks (only observed in Ikakogi tayrona [47]).

Figure 10. Snout shape in Glassfrogs. (A) Round, Hyalinobatrachium munozorum, KU 118054. (B) Bluntly round, Centrolene ballux, KU 164725. (C) Truncate, Nymphargus megacheirus, KU 143269. (D) Slightly protruding, “Centrolene” medemi, KU 164493. (E) Protruding, “Cochranella” balionota, KU 164708. (F) Sloping, Nymphargus grandisonae, KU 164688. Drawings by Juan M. Guayasamin.

Morphometrics—morphological variables were measured with digital callipers to the nearest 0.1 mm, as follows: (1) Snout–vent length (SVL) = distance from tip of snout to posterior margin of vent; (2) tibia length (TL) = length of flexed leg from knee to heel; (3) foot length (FL) = distance from proximal margin of outer metatarsal tubercle to tip of Toe IV; (4) head length (HL) = distance from tip of snout to posterior angle of jaw articulation; (5) head width (HW) = width of head measured at level of jaw articulations; (6) interorbital distance (IOD) = distance between upper eyelids, representing the width of the underlying frontoparietals; (7) upper eyelid width (UE) = greatest transverse width of upper eyelid; (8) internarial distance (IN) = distance between nostrils; (9) eye–nostril distance (EN) = distance from posterior margin of nostril to anterior margin of eye; (10) snout–eye distance (SE) = distance from tip of snout to anterior margin of eye; (11) horizontal eye diameter (ED) = distance between anterior and posterior borders of eye; (12) tympanum diameter (TD) = distance between anterior and posterior margins of tympanic annulus; (13) eye–tympanum distance (ET) = distance from posterior border of eye to anterior margin of tympanic annulus; (14) radio–ulna length (RUL) = length of flexed forearm from elbow to proximal border of palmar tubercle; (15) hand length (HDL) = distance from the proximal margin of palmar tubercle to tip of Finger III; (16) Finger-I length (F1L) = distance from outer margin of palmar tubercle to tip of Finger I; (17) Finger-II length (F2L) = distance from outer margin of palmar tubercle to tip of Finger II; (18) disc of Finger III (3DW) = greatest width of disc of Finger III; and (19) Finger-III width (F3W) = width of Finger III measured at the level of distal subarticular tubercle, including lateral fringes and excluding webbing. For comparing different body sizes (SVL) among glassfrogs, we considered the average size of males and categorized them according to the following criteria: Minute (SVL < 22 mm), small (SVL 22–25 mm), medium (SVL 25–30 mm), large (SVL 30–50 mm), and giant (SVL > 50). Eye diameter was divided into small (eye diameter < 10% SVL), moderate size (eye diameter 10%–15% SVL), and large (eye diameter > 15% SVL). Tympanum was considered to be very small (tympanum < 20% of eye diameter), small (tympanum 20%–30% of eye diameter), moderate (tympanum 31%–40% of eye diameter), large (tympanum diameter 41%–50% of eye diameter), and very large (tympanum diameter > 50% of eye diameter.

Figure 11. Webbing in glassfrogs. (A) Terminology used for webbing formula in hands and feet (see Guayasamin et al. [20]; modified from Savage and Heyer [44]). Roman numerals (I, II, III, IV, V) represent fingers or toes. Arabic numerals represent the number of phalanges completely or partially free of webbing. We use 0⁻ to indicate that webbing reaches the distal margin of the disc; 0 indicates that webbing reaches the middle of the disc; 0⁺ indicates that webbing reaches the proximal margin of the disc; 1⁻ indicates that webbing reaches the distal margin of the intercalary cartilage; 1 indicates that the webbing the middle of the intercalary cartilage; 1⁺ indicates that the webbing the proximal margin of the intercalary cartilage; 2⁻ indicates that webbing reaches the distal margin of the distal subarticular tubercle; 2 indicates that webbing reaches the middle of the distal subarticular tubercle; 2⁻ indicates that webbing reaches the proximal margin of the distal subarticular tubercle. For example, webbing formula in the illustrated foot is absent between Toes I and II (lateral fringes are not considered as webbing); II 1^2/3—3⁺ III 1^2/3—3⁻ IV 3⁻—2 V. Figure modified from Guayasamin et al. (2006). (B) Simplified type of hand webbing. Absent: Nymphargus cochranae, QCAZ 31113. Moderate: Chimerella mariaelenae, QCAZ 22363. Extensive: Centrolene geckoidea, KU 164490. Drawings by Juan M. Guayasamin.

Figure 12. Prepollex and nuptial pad morphology in glassfrogs. (A) Condition of the prepollex. Concealed, Nymphargus cochranae, QCAZ 31113. Distinct, Teratohyla spinosa, KU 164668. (B) Nuptial pad morphology. Type I: Large to medium-size nuptial excrescence present on the dorsal, lateral, and/or ventral sides of the thumb; Cochranella posadae, QCAZ 26023. Type II: Small circular or squarish nuptial excrescence present on the dorsal or dorsolateral face of the thumb; Centrolene lynchi, MCZ 97846 (figure modified from Flores 1985). Type III: Medium-size spinous nuptial excrescence extending from the lateral side of the thumb to its dorsomedial surface; Nymphargus armatus, UVC 9400 (modified from Lynch and Ruiz-Carranza 1996). Type IV: Large nuptial excrescence formed by a granular pad that extends from the side of the thumb to its dorsomedial surface, and on the proximal dorsolateral surface of finger II; Cochranella litoralis, ICN 13821. Type V: Medium-size diffuse nuptial excrescence formed by glandular clusters and individual glands; a pad as such is absent; Hyalinobatrachium aureoguttatum, QCAZ 27429. Type VI (not illustrated): Nuptial excrescences formed by a combination of clustered and individual glands that sparse along the flanks of the body (see text).

Figure 13. Peritonea and humeral spines in Centrolenidae. (A,Left): Pericardium lacking iridophores, hepatic, and visceral peritonea with iridophores (Hyalinobatrachium aureoguttatum). (A,Right): Pericardium with iridophores, hepatic, and visceral peritonea lacking iridophores (Centrolene buckleyi). (B,Top): Absence of humeral spine (H. fleischmanni). (B,Bottom): Presence of humeral spine in males (Espadarana callistomma). Figure modified from Guayasamin et al. [1].

Figure 14. Variation of humeral spines, crista medialis, and crista ventralis, in adult males of Centrolenidae (modified from Guayasamin et al. [1]). Illustrated specimens are: Centrolene sanchezi, KU 170116; C. geckoidea, ICN 5598; C. pipilata, KU 143286; Cochranella euknemos, KU 77534; Cochranella litoralis, QCAZ 27693; Teratohyla spinosa, KU 32935; Nymphargus griffithsi, KU 288992, 188148; N. cochranae, KU 123218; N. megacheirus, KU 143271; Sachatamia albomaculata, KU 65185; S. ilex, LACM 72910; Chimerella mariaelenae, QCAZ 21252; H. valerioi, KU 178091.

Figure 15. Ventral surfaces of tights in glassfrogs. (A) With enlarged subcloacal warts. (B) Lacking enlarged warts. Illustrations by Juan M. Guayasamin.

Figure 16. Tubercles on limbs. (A) Tubercles on ventrolateral edge of Finger IV and arm, Cochranella resplendens, KU 118053. (B) Tubercles on ventrolateral edge of Toe V and tarsus, C. mache, QCAZ 22412. Photos by Martín Bustamante.

Vocalizations—Calls were recorded in the field by different researchers (see Acknowledgments) using an Olympus Linear PCM Recorder LS-10 tape recorder and a Sennheiser K6-ME67 directional microphone. Recordings obtained by us are stored at the Laboratorio de Biología Evolutiva of the Universidad San Francisco de Quito (LBE). Sounds were recorded in PCM format at a sampling frequency of 44.1 kHz. Audiospectograms and oscillograms were generated in the R package SeeWave [48]. Frequency information was obtained through fast Fourier transformation, with a 50% window overlap (Hanning window size of 1024 and a frequency window of 43.1 Hz). The following call variables were measured: Call duration, duration between calls, number of notes per call, note duration, duration between notes, dominant frequency, first harmonic, second harmonic [49,50,51] (Table 1). A call is defined as the sound produced in a single exhalation of air. Calls and notes were divided into two categories, tonal and pulsed, based on distinct morphology. Tonal calls/notes have non-peaked amplitude sustained throughout the duration of the note. Pulsed calls/notes are characterized by having at least one clear amplitude peak. Pulses were defined as a visible increase and decrease of amplitude peaks on the oscillogram within a single note, and notes were defined by a single and complete amplitude rise from and return to the base frequency of the background noise. For a review on the use of bioacoustics in taxonomy and specific variable definitions, see Köhler et al. [52].

Specimens examined—We examined ethanol-preserved specimens from the following herpetological collections: Centro de Biodiversidad y Genética, Cochabamba, Bolivia (CBG); Círculo Herpetológico de Panamá (CHP); Centro Jambatu de Investigación y Conservación de Anfibios, San Rafael, Ecuador (CJ); División de Herpetología, Museo Ecuatoriano de Ciencias Naturales, Quito, Ecuador (DHMECN); Field Museum, Division of Amphibians and Reptiles, Chicago, USA (FMNH); Instituto de Ciencias Naturales, Universidad Nacional de Colombia, Bogotá, Colombia (ICN); University of Kansas, Museum of Natural History, Division of Herpetology, Lawrence, Kansas, USA (KU); Natural History Museum of Los Angeles County, Section of Herpetology, Los Angeles, California, USA (LACM); Museum of Comparative Zoology, Harvard University, Cambridge, Massachusetts, USA (MCZ); Museo de Historia Natural La Salle, Caracas, Venezuela (MHNLS); Museo de Zoología, Universidad Tecnológica Indoamérica, Quito, Ecuador (MZUTI); Museo de Zoología, Pontificia Universidad Católica del Ecuador, Quito, Ecuador (QCAZ); Colección de Herpetología, Escuela de Biología, Universidad de Costa Rica, San José, Costa Rica (UCR); Museo de Vertebrados, Universidad del Valle, Cali, Colombia (UVC); and Museo de Zoología, Universidad San Francisco de Quito, Ecuador (ZSFQ). When specimens were not available for direct comparison, we relied on descriptions in the literature. Sexual maturity of specimens was determined by the presence of vocal slits and nuptial pads in males and by the presence of eggs or convoluted oviducts in females.

2.3. Evolutionary Relationships

Taxon and gene sampling—we combined genetic sequences available from previous work (mostly [2,3,19]) and new sequences generated during this study (Table S2). Genomic extraction, amplification, and sequencing are as described in Guayasamin et al. [2] and Castroviejo-Fisher et al. [3]. The final dataset contains complete or partial sequences of 10 genes representing 6513 bp of data (mitochondrial: 12S rRNA, 16S rRNA, ND1; nuclear: BDNF, C-MYC exon 2, CXCR4, NCX1, POMC, RAG1, SLC8A3). We sampled 49 outgroup taxa from a large range of families within Hyloidea (Table S2), and include all three species from Allophrynidae, the sister group to Centrolenidae [2,12]. Finally, ingroup taxon sampling includes 251 terminals, representing 113 named species, and 24 putative new species. The percentage of named species sampled (i.e., matrix completeness) for each marker is as follows: 12S = 90%; 16S = 94%; BDNF = 55%; C-MYC exon 2 61%; CXCR4 = 61%; NCX1 = 59%; ND1 = 81%; POMC = 61%; RAG1 = 62%; and SLC8A3 = 60% (complete marker statistics can be found in Table S3). For most species of Ecuadorian glassfrogs, we obtained a total of ~2733 bp from the following mitochondrial markers: 12S rRNA (~907 bp), 16S rRNA (~864 bp), and ND1 (~960 bp). See Table S2 for genes sequenced for each species and GenBank Accession Numbers.

Candidate species—to assist in identifying putative new species for future study, we employed the candidate species designation system of Vieites et al. [29]. Prior to this study, divergent centrolenid lineages have been designated in inconsistent ways (using sp. or aff. or museum numbers), which can result in confusion in identifying and studying tentative new species. Given the increasing number of putative new species in Centrolenidae, establishing candidate species aims to better organize and maintain the growing discovery of undescribed lineages. Importantly, designating a divergent lineage as a putative new species does not necessarily mean it is a new species; rather, the system is meant to identify these lineages for future study, incorporating multiple lines of evidence (i.e., morphology, bioacoustics, biogeography, or nuclear genes) in order to determine their species status.

Candidate species are typically identified through evidence from genetic distances or phylogenetic lineage divergence through widely used genetic markers that provide a basis for comparison (i.e., 12S, 16S, CO1; [29,53]). In this study, we used the mitochondrial genetic markers 12S, 16S, and ND1 and integrated evidence from genetic distances and phylogenetic relationships to identify candidate species. We first used a threshold of 3% in identifying divergent lineages, which is a threshold that most named centrolenid species exceed (this study). Putative new species were then numbered using a scheme of “sp_CaXX”, with numbering beginning at 01 for each genus. Finally, we note that Vieites et al. [29] also uses “unconfirmed” and “confirmed” designations, whereas “confirmed” candidate species have additional evidence (morphology or calls) for their status as species and are simply awaiting detailed analyses and subsequent description. We did not use these designations, as most putative new centrolenid species are supported only through mitochondrial genetic divergence.

Phylogenetics—the analyzed dataset contains complete or partial sequences of 10 genes representing 6513 bp of data (mitochondrial: 12S rRNA, 16S rRNA, ND1; nuclear: BDNF, C-MYC exon 2, CXCR4, NCX1, POMC, RAG1, SLC8A3). See Tables S2 and S3 for details. Analyses were conducted using maximum likelihood (ML) and Bayesian (BA) criteria. We did not perform searches under the parsimony criterion because of its limitations under certain conditions (long branch attraction; [54,55]) and lack of theoretical support for transition/transversion bias and variation in substitution rates among different nucleotide sites [56]. The 12S and 16S rRNA sequence data were aligned using MAFFT 7.4 [57], using the Q-INS-i algorithm that takes RNA secondary structure into consideration. Protein coding genes were also aligned in MAFFT using the AUTO function and manually inspected for accuracy and open reading frames.

Maximum likelihood was run in the IQ-TREE v1.5.5 software [58]. The data were automatically partitioned, and the best model was implemented using ModelFinder within IQ-TREE [59], which groups partitions with the same model and similar rates and simultaneously searches model and tree space. Node support was assessed via 100 ultra-fast bootstrap replicates [60].

For Bayesian phylogenetic analyses and divergence dating, we used BEAST v2.4.5 [61]. We used a single secondary calibration point using ages estimated from Hutter et al. [62], which estimates divergence dates for Hyloidea using 18 genetic markers and 8 fossil/geographic calibrations. We used median age of Centrolenidae with a normal distribution to capture the 95% confidence interval from this study (Mean = 33.4 Myr; Sigma = 6). We used a single uncorrelated lognormal relaxed clock prior linked across all partitions. We partitioned data by codon position for protein-coding genes and used the Bayesian bModelTest package within BEAST to select the best model for each partition. We estimated the relaxed clock rate using an initial value of 1e-9 and a broad prior (in our case, a gamma distribution with a shape parameter of 0.001 and scale parameter of 1000). We used a Yule speciation process for the tree prior. We ran Markov chain Monte Carlo (MCMC) searches for a total of 100 million generations, sampling every 10,000 generations. Stationarity was assessed by examining the standard deviation of the split frequencies and by plotting the -lnL per generation, using Tracer v1.5 [63]; trees generated before stationarity were discarded as “burn-in”, which was 20% of trees.

Species conservation status—global and local (Ecuador) conservation status of glassfrogs follow the categorization and criteria established by the International Union for Conservation of Nature (IUCN) [64] (Figure 17), including rate of decline, population size, area of geographic distribution, and degree of distribution fragmentation. The following categories were used: (i) Not Evaluated: The species has not yet been evaluated against the criteria, (ii) Data Deficient: There is inadequate information to make a direct or indirect assessment of its risk of extinction based on its distribution and/or population status, (iii) Least Concern: The species is widespread and abundant and not under immediate risk of extinction, (iv) Near Threatened: For species that are not currently threatened, but are close to qualifying for or are likely to qualify for a threatened category in the near future, (v) Vulnerable: A taxon is Vulnerable when it is considered to be facing a high risk of extinction in the wild (Criteria A to E for Vulnerable), (vi) Endangered: When the species is considered to be facing a very high risk of extinction in the wild (Criteria A to E for Endangered), (vii) Critically Endangered: When the species is considered to be facing an extremely high risk of extinction in the wild (Criteria A to E for Critically Endangered), (viii) Extinct in the Wild: When the species is known only to survive in cultivation, in captivity, or as a naturalized population, outside its historical range. A taxon is presumed Extinct in the Wild when exhaustive surveys in known and/or expected habitats, at appropriate times, throughout its historic range have failed to record an individual, (ix) Extinct: When there is no reasonable doubt that the last individual of the species has died. A taxon is presumed Extinct when exhaustive surveys in known and/or expected habitats, at appropriate times (diurnal, seasonal, annual), throughout its historic range have failed to record an individual.

2.4. Biogeographic Regions of Ecuador

In this study, we applied biogeographic regions of continental Ecuador as a simplification of vegetation types [65,66] (Figure 18). This system has the virtue of combining the characteristics of the vegetation and the historic isolation between the western and eastern slopes of the Andes and the eastern and western lowlands. Mean annual precipitation and mean annual temperature for each of the recognized regions are shown in Table 2. We also summarized the land cover of Ecuador, illustrating which ecosystems have suffered intensive deforestation and which are still preserved (Table 3, Figure 18).

Dry Shrub—characterized by a combination of warm and extremely dry conditions. Annual precipitation can be as low as 60 mm in the westernmost locality (Salinas, Santa Elena Province). This region covers an area of 8033 km² and is restricted to the coastal margin of central Ecuador (Figure 18). In some areas, grasses introduced for raising livestock have replaced native plants. In the drier habitats, xerophytic plants are dominant, especially cacti and other thorny plants [67].

Western Deciduous Forest—this forest occurs at an elevation of 50–300 m in central and northern Ecuador (100–400 m in southern Ecuador) and covers 25,673 km² (10.3% of Ecuadorian territory, Figure 18). Conditions are drier and the terrain has lower tree densities than in evergreen forests. The trees are generally shorter than 20 m with an understory that can be dense with abundant herbaceous plants. Some tree species lose their leaves during the dry season [67]. More than half of the land cover of this habitat type has been converted for agriculture and grazing cattle.

Chocoan Tropical Rainforest—this rainforest is the second largest biogeographic region in Ecuador, with 31,732 km² at elevations ranging from sea level to 300 m (Figure 18). It has a closed canopy with trees that can reach 30 m in height and with an understory dominated by ferns and Araceae [67]. Tree diversity is high, with more than 100 species/ha with diameter at breast height >10 cm, but lower than in the Amazonian Tropical Rainforest [68]. Habitat destruction rate in this region is the highest in Ecuador and only 18.3% of its natural vegetation remains.

Western Foothill Forest—this forest covers 15,305 km² on the western Andean slopes with an elevational range of 300–1300 m (400–1000 m in southern Ecuador). Plant endemism is high, especially between latitudes 0° and 3° S [67]. This forest is structurally similar to its counterpart from the eastern Andean slope, although the amphibian communities are highly differentiated.

Figure 18. Land use, biogeographic regions, and provinces of Ecuador. Land cover: Modified from Ron et al. [66]; “Mosaics” are mixtures of natural vegetation and either agricultural land or pastures. “Other” includes urban areas, shrimp farms, lakes, rivers, glaciers, and sand banks. Biogeographic regions: Shown as a simplification of vegetation types (Sierra et al. [65], as modified by Ron et al. [66]). Provinces: Provinces are divided into three broad geographic regions: Coast, Andes, and Amazon.

Western Montane Forest—this evergreen forest covers 21,576 km² at an elevational range between 1300–3400 m (1000–3000 m in southern Ecuador; Figure 18). The canopy is generally below 25 m with a high abundance of epiphytic plants (especially mosses, ferns, orchids, and bromeliads). At intermediate elevations, especially during the afternoon, the forests become misty and receive horizontal precipitation from low, overhanging clouds. Western Montane Forest is restricted to narrow stretches between the basin of the Mira River (close to the Colombian border) and the basins of the Chanchán and Chimbo rivers (2° S). It is replaced by drier habitats (principally Andean Shrub) south of 4° S, close to the border with Peru. Only 35% of its natural vegetation remains unaltered (Table 3).

Páramo—this is the vegetation type that reaches the highest elevation and covers 15,976 ha (6.1% of the territory; Figure 18). Depending on the region, its lower limit lies between 3000 and 3600 m. Short herbaceous plants generally forming tight clumps dominate the vegetation. The plants are adapted to cold temperatures and to low availability of water. Open grassy areas predominate but are mixed with small patches of forest or shrubs [69]. At higher elevations, the vegetation is restricted to sparse clumps on otherwise bare land. Because of the occurrence of frequent freezes, agriculture is limited, and this has ameliorated anthropogenic habitat degradation. In this region, only 21.1% of the natural vegetation has been cleared or severely fragmented, the lowest proportion for any region (Table 3). However, the Páramo is the region with the highest proportion of endangered amphibians.

Andean Shrub—this biogeographic region lies between 1400 and 3000 m and has an area of 11,266 km²; it is found in the inter-Andean basins between the Cordillera Occidental and Cordillera Oriental (Figure 18). As a result of rain shadow effects from both mountain chains, the Andean Shrub has a relatively low precipitation (Table 2). Although originally dominated by shrubs, most of the vegetation has been replaced by crops, pastures, or forests of exotic trees (Eucalyptus and Pinus; [69]). In dry valleys (e.g., Chota, Guayllabamba, and Patate) the native vegetation is spiny. Andean Shrub is almost unrepresented in the Ecuadorian National System of Protected Areas. Habitat degradation is severe; more than half the land cover is devoted to agriculture or to raising cattle (Table 3).

Eastern Montane Forest—this forest covers 31,555 km² between 1300 m and 3600 m on the eastern Andean slopes (Figure 18). The vegetation is structurally similar to that from the Western Montane Forest. Above 2900 m, the soil of the forest is covered by moss and the trees are irregularly shaped [69].

Eastern Foothill Forest—this evergreen forest covers 13,133 km² between elevations of 600 m and 1300 m (Figure 18), and is a mixture of tree species from the Andes and the lowlands of the Amazon Basin [68]. The canopy reaches up to 30 m in height and encloses a dense sub-canopy and understory. Tree diversity is lower (130 species/ha, >10 cm DBH) [68] than in the Amazonian Tropical Forest. Average annual precipitation is the second highest of all regions (2833 mm).

Amazonian Tropical Rainforest—the Amazonian Tropical Rainforest is the most extensive biogeographic region in Ecuador with a total area of 73,909 km² (29.8% of the Ecuadorian continental territory; Figure 18). It is restricted to elevations below 600 m and has the highest average annual precipitation of any region (3349 mm). The dominant forest type, Terra Firme, is characterized by well-drained soils. The canopy is 10–30 m high, punctuated with emergent trees up to 40 m (and rarely 50 m); small gaps created by fallen trees [68,70] are common. Tree diversity is high with 200–300 species of trees/ha (>10 cm DBH) [68,70]. Other vegetation types in this region include Várzea forest (flooded with white water), Igapó forest (flooded with black water), riparian woodland forest, river island scrub, and Mauritia flexuosa palm swamps [68,71]. At the local scale (≤100 km²), amphibian diversity reaches its global peak in the Amazonian Tropical Rainforest of Ecuador [72,73].

2.5. Potential Distribution

Estimating the distribution of a species is challenging, since biotic, abiotic, and historic factors come into play. As an approximation to species distributions and being aware of the associated caveats [74,75], we modelled the ecological niche of all species for which we had at least 10 independent localities (>1 km apart). Given that our ecological models did not include variables such as biotic interactions, random extinction, vagility, influence of diseases or introduced species, we used our results only as proxy, and are fully aware that predicted areas might be, in most cases, overestimated; nevertheless, models are useful for conservations assessments. Locality data were obtained from the following museums: AMNH, BMNH, FHGO, KU, QCAZ, MECN, MZUTI, ZSFQ, and USNM. All specimens were directly examined by Juan M. Guayasamin or Diego F. Cisneros-Heredia.

We used all 19 climatic variables from the WorldClim database (www.worldclim.org). These bioclimatic variables are derived from monthly temperature and rainfall values [76]. A correlation analysis was used to determine independent variables for the distribution of each species. The potential distributions of centrolenid species were estimated using default settings in Maxent v. 3.3.3k [77,78]. We used 70% of the presence records as training data, and the other 30% was used to evaluate the model. Maxent was run 10 times to obtain the ecological model for each species. In order to evaluate models, we employed the area under the curve (AUC), a value that is an indicator of model performance [77,79]. The AUC value was calculated in Maxent and ranges between 0.5 (random classification) and 1 (perfect fit). We discarded models that had AUC values below 0.7 [80]. Maxent continuous models were reclassified in order to obtain a binary map of potential presence or absence, using the maximum training sensitivity plus specificity threshold (MTS + S), a method that has been shown to produce highly accurate predictions [81,82]. The potential distribution models were edited using ArcMap v.10 to obtain more realistic estimates of species distributions by removing areas predicted as present that are located in inaccessible biogeographic regions (e.g., species restricted to the Amazon basin predicted as present in the Chocó and vice versa; species from the Pacific slope of the Andes predicted as present in the Amazonian slope and vice versa).

Impacts of human activities to species—in order to build a layer of human impact (LHI) for Ecuador, we used shape files containing information on land use, roads, human settlements, human population density, mining, and oil exploration and concession shape files (ESRI, 2003). The original shape files were converted to raster files in ArcMap v. 10; we then calculated Euclidean distances in each raster file to compute buffer areas around specific features (i.e., roads, oil fields), giving values according to the intensity of human impact and distance from the specific human activity (Table 4). In the case of population density, the intensity value was equal to the logarithmic scale of the population density value for each grid cell. We obtained raster layers with buffer zones and intensity values for each of the impacts, which were added using the map calculator tool (ArcMap v. 10) to obtain a single map summarizing all considered threats.

2.6. Registration of New Nomenclatural Acts

According to the International Commission on Zoological Nomenclature (ICZN), which produces the International Code of Zoological Nomenclature, the electronic publication of this article in portable document format (PDF) represents a published work. Therefore, the new species name contained in the PDF is effectively published under the International Code of Zoological Nomenclature from the electronic edition alone. This publication and the nomenclatural acts contained in it have been registered in ZooBank, the online official register for the ICZN. The ZooBank Life Science Identifiers (LSIDs) can be accessed and viewed through standard web browsers by appending the LSID to the prefix http://zoobank.org/. The online version of this work is archived and available from the following digital repositories: Diversity, CLOCKSS, and e-Helvetica.

3. Results

3.1. Phylogenetic Systematics

The relationships among glassfrog genera are shown in Figure 19. We also included a tree for each genus that occurs in Ecuador. These trees are the source of evidence for generic placement and comments on the evolutionary relationships among species discussed below and are summarized in Table S1. Each genus was well supported and congruent with the trees estimated by Guayasamin et al. [1,2], Castroviejo-Fisher et al. [3], and Twomey et al. [19], with only a few differences resulting from our improved taxon sampling. There was significant support (ML: Greater than 95% bootstrap [BS]; BA: Greater than 0.95 posterior probability [PP]) for all genera.

3.2. Potential Distribution

The area of the potential distribution of each glassfrog species is summarized in Table S5. AUCs associated to each model are reported in Table S4. The species with the largest area of potential distribution was Cochranella resplendens (77,792 km²), whereas the species with the most restricted predicted distribution was Centrolene heloderma (1067 km²). The areas with the highest impacts were the interandean valleys and the Pacific lowlands; these were also areas with the highest population density. In terms of distribution and range extensions, agriculture and ranching were the activities that had the highest impact on habitats. Other activities with high impact values are related to mining and the oil industry.

4. Species Accounts

Genus Centrolene Jiménez de la Espada, 1872 [83]
Etymology: The name Centrolene is derived from the Greek words kéntron (point or spur) and ōlénē (elbow, forearm), alluding to the humeral spine found in males of this genus [84]. Although the name Centrolene has been used as neuter in gender for almost 150 years, now, because of an intricate game of words and nomenclatural regulations, it is considered to be feminine [85], creating nomenclatural instability that, in our opinion, was unnecessary. Centrolene is the type genus for the family Centrolenidae.
Centrolene ballux (Duellman and Burrowes 1989 [86]; Figure 20, Figure 21,Figure 22 and Figure 23)
Centrolenella ballux Duellman and Burrowes, 1989 [86]. Holotype: KU 164725.
Type locality: “14 km (by road) west of Chiriboga (00°18′ S, 78°49′ W), 1960 m, Provincia de Pichincha, Ecuador” (now in Provincia de Santo Domingo de los Tsáchilas).
Centrolene ballux—Ruiz-Carranza and Lynch, 1991 [6].
“Centrolene” ballux—Guayasamin, Castroviejo-Fisher, Trueb, Ayarzagüena, Rada, and Vilà, 2009 [1].

Common names: English: Gold-dust Glassfrog. Spanish: Rana de Cristal Polvo de Oro.

Etymology: The specific name ballux is Latin; it means “gold dust” and is used in reference to the minute gold flecks on the dorsum [86].

Identification: On the Pacific versant of the Ecuadorian Andes, only Nymphargus buenaventura and some populations of N. griffithsi are similar to Centrolene ballux in having a green dorsum with small light spots (Figure 20), but they differ by possessing basal webbing between outer fingers (moderate webbing in C. ballux), concealed prepollex (distinct in C. ballux), and lacking humeral spines (present in males of C. ballux). The Colombian “Centrolene” robledoi resembles C. ballux; both lack vomerine teeth, have a small series of white spots on flanks, and have a similar webbing and snout shape. However, C. robledoi has small dark spots on its dorsum (absent in C. ballux), concealed prepollex (distinct in C. ballux), and is slightly larger than C. ballux (males, SVL 19.9–24.4 mm in C. robledoi; 19.2–22.2 mm in C. ballux). Centrolene peristicta and C. lynchi are sympatric with C. ballux in several localities, but the two species can be distinguished by having dorsal dark and light minute spots (only light spots present in C. ballux; Figure 20).

Diagnosis: (1) Vomers lacking teeth; (2) snout bluntly rounded to truncated in dorsal and lateral profiles (Figure 21); (3) tympanum oriented almost vertically, ED/TD = 31%–34%; tympanic annulus visible except for dorsal border covered by supratympanic fold; tympanic membrane partially pigmented, differentiated from surrounding skin; (4) dorsal surfaces shagreen with small warts; (5) pair of enlarged subcloacal tubercles (Figure 15); (6) anterior two-thirds of ventral parietal peritoneum white, posterior third transparent (condition P3); silvery white pericardium; no iridophores in peritonea covering intestines, stomach, testes, kidneys, gall bladder, and urinary bladder (condition V1); (7) liver tetralobed, lacking iridophores (condition H0); (8) humeral spines present in adult males; (9) no webbing between Fingers I and II, webbing between Fingers II and III basal or absent; webbing formula for outer fingers: III (2^1/4–2^3/4)—(2–2⁺) IV (Figure 21C); (10) webbing formula on foot: I 1—(2–2⁺) II 1—(2–2⁺) III (1–1⁺)—(2–2^1/3) IV 2^1/2—(1–1^1/3) V; (11) ulnar fold low, white; inner tarsal fold low; outer tarsal fold absent, but small white tubercles evident along ventrolateral margin of tarsus; (12) distinct prepollex, clearly separated from Finger I; in males, nuptial pad Type I; (13) Finger II slightly longer than Finger I (Finger I 91.4%–98.0% of Finger II); (14) disc of Finger III of moderate size, about 47.4%–59.3% of eye diameter; (15) in life, dorsum green with small white warts; upper lip white; bones green; (16) in preservative, dorsal surfaces lavender with small white and/or unpigmented spots, particularly evident on limbs; (17) iris whitish cream with dark-grey thin reticulation and pale yellow hue around pupil; (18) melanophores mostly absent from fingers and toes, except for a few on Toes IV and V and on base of Finger IV; (19) males call from the upper side of leaves; call emitted sporadically and consisting of a single short note (328.5–420.4 ms) with 7–9 pulses; mean dominant frequency at peak amplitude 4833 Hz ± 14 (range 444–464); notes are frequency modulated (Márquez et al. 1996); (20) fighting behavior unknown; (21) eggs deposited on the upper sides of leaves; short-term maternal care unknown; long-term parental care absent; (22) tadpoles undescribed; (23) minute body size; SVL 19.2–22.2 mm (X = 20.6 ± 0.911, n = 25) in males; SVL 21.0–23.3 mm (n = 3) in females.

Figure 20. Centrolene ballux in life from Reserva Las Gralarias, Pichincha province, Ecuador. (A,B) Adult males. (C) Adult male in dorsal view, QCAZ 40199. (D) Egg clutch on upper side of leaf. (E) Metamorph. (F) Adult female in ventral view, QCAZ 40196. Photos by Carl R. Hutter (B,D,E), Luis A. Coloma (C,F), and Alejandro Arteaga/Tropical Herping (A).

Figure 21. Centrolene ballux. (A) Head in lateral view, KU 164725. (B) Finger I and nuptial pad in dorsal view, KU 200275, adult male. (C) Hand in ventral view, KU 164725. Illustrations by Juan M. Guayasamin.

Color in life (Figure 20): Dorsum lime green with small greenish–white to yellowish–white warts; fingers and toes yellowish green; anterior two-thirds of ventral parietal peritoneum white; visceral peritoneum lacking iridophores; pericardium silvery white; bones green; upper lip yellowish white to white; iris cream white with fine black reticulations.

Color in ethanol: Dorsal surfaces of head, body, and limbs lavender with small white and/or unpigmented warts; hands and Toes I–III mostly unpigmented; margin of upper lip white; ulnar fold white; small tubercles on outer ventrolateral margin of tarsus white; small white tubercles posterior to cloacal opening; ventral surfaces cream. Anterior two-thirds of the ventral parietal peritoneum white, posterior third transparent; silvery white pericardium; no iridophores in peritonea covering intestines, stomach, testes, kidneys, gall bladder, and urinary bladder.

Biology and ecology: Individuals are active at night on the upper surfaces of leaves of bushes and trees 50–800 cm above and along small streams, or on ferns over roadside ditches (Arteaga et al. 2013). The reproductive activity of Centrolene ballux occurs in the rainy season (December–April [87]). One male (IND-AN 1725) was on a leaf 5 cm below another leaf on which there was a clutch of 18 eggs [86]. Another clutch found at La Planada had 13 eggs. Near Chiriboga, a gravid female (KU 164729) with green eggs and five adult males were found along a stream on 8 May 1975. At Reserva Las Gralarias, on March 2009, a clutch with 21 embryos was found on the upper side of a leaf, which was hidden by another leaf; an adult male was observed nearby (ca. 100 cm from the clutch). Centrolene ballux is relatively abundant at Reserva Las Gralarias, where it maintains reproductive populations along several streams (Ballux Creek, Five-frog Creek, Heloderma Creek, Chalguayacu River, Kathy’s Creek; [88]). It is unknown if females provide short-term parental care to egg clutches; males do not exhibit parental care [25].

Call (Figure 22): The advertisement call of Centrolene ballux was described from Las Palmeras, Ecuador, by Márquez et al. [89]. The call was emitted sporadically and consisted of a single short note (328.5–420.4 ms, n = 2) with 7–9 pulses, the first 4–5 being repeated at short, regular intervals, and the last three pulses being emitted at longer intervals (mean pulses per second = 21.0 ± 0.1, range 21.3–21.4). The dominant frequency was high (mean dominant frequency at peak amplitude 4833 Hz ± 14). The notes were frequency modulated (dominant frequency at the beginning of note is lower than at its end). Recordings obtained at Reserva Las Gralarias, Ecuador, match this description.

Figure 22. Call of Centrolene ballux from Reserva Las Gralarias, Pichincha province, Ecuador.

Tadpole: Not described.

Distribution (Figure 23): Centrolene ballux is known from six localities in the cloud forests of Carchi, Imbabura, Pichincha, and Santo Domingo de los Tsáchilas provinces (Ecuador) and one in the department of Nariño (Colombia) on the Pacific slopes of the Cordillera Occidental of the Andes at elevation between 1780 and 2340 m [86,87,89] (Specimens Examined). The species inhabits the Western Montane Forest ecoregion. In Ecuador, it has a potential distribution of 4752 km².

Conservation status: Globally, Centrolene ballux is currently listed as Endangered by the IUCN [90]. In Ecuador, the most recent records of the species are from Reserva Las Gralarias (March 2018) and Reserva Río Manduriacu (2008) [21]. In Colombia, the last published record is from Reserva La Planada on April–June 1986 [86]. Recent surveys at Quebrada Zapadores and Las Palmeras (= Río Guajalito) have been unsuccessful in finding this species [91] (DFCH unpubl. data). During the rainy seasons (December–April) of 2011–2018, reproductive populations were observed in four streams at Reserva Las Gralarias. The species has a very restricted distribution, with only five known localities in Ecuador, in an area where forest fragmentation is common. At Reserva Las Gralarias, the amphibian chytrid fungus Batrachochytrium dendrobatidis infects C. ballux with a relatively high prevalence, but no recent declines have been recorded [92].

Evolutionary relationships (Figure 24): Centrolene ballux and C. buckleyi are sister species.

Specimens examined: Centrolene ballux: Ecuador: Provincia de Carchi: ca. 5 km W La Gruel (0.916667 N, 78.13333 W; 2340 m), KU 202798. Provincia de Pichincha: Reserva Las Gralarias (0.00806 S, 78.72433 W, 1852 m), QCAZ 40195–99; Quebrada Zapadores, 5 km ESE of Chiriboga on Chiriboga–Quito road (0.245278 S, 78.7261 W, 2010 m), KU 164733. Provincia de Santo Domingo de los Tsáchilas: 14 km we’st of Chiriboga on Chiriboga–Santo Domingo road (0.265278 S, 78.8478 W, 1960 m), KU 164725 (holotype), 164726–32 (paratypes).

Localities from the Literature: Provincia de Pichincha: Las Palmeras (=Bosque Protector Río Guajalito) (0.283 S, 78.75 W; 1800 m) [89].

Figure 23. Distribution of Centrolene ballux in Ecuador (yellow dots).

Figure 24. Evolutionary relationships among species in the genus Centrolene, inferred using maximum likelihood and Bayesian criteria.

Centrolene buckleyi (Boulenger 1882 [93]; Figure 25, Figure 26, Figure 27 and Figure 28)
Hylella buckleyi Boulenger, 1882 [93]. Syntypes: BM 78.1.25.16 (Intac), 80.12.5.201 (Pallatanga). See comments below on type material. Neotype: KU 202770.
Neotype locality: Isla Wolf of Laguna Cuicocha (0°18′07″ N, 78°22′00″ W; 3070 m), Provincia Imbabura, Ecuador, collected on 28 February 1984 by W. E. Duellman.
Centrolenella buckleyi—Noble, 1920 [94].
Cochranella buckleyi—Taylor, 1951 [15]. Rivero, 1961.
Centrolenella buckleyi buckleyi—Rivero, 1968 [95].
Centrolenella johnelsi Cochran and Goin, 1970 [96]. Holotype: MLS 432. Type locality: “San Pedro, N of Medellín, Antioquia, Colombia”. Synonymy by Ruiz-Carranza and Lynch, 1991 [6].
Centrolene buckleyi—Ruiz-Carranza and Lynch, 1991 [6].

Common names: English: Buckley’s Glassfrog. Spanish: Rana de Cristal de Buckley.

Etymology: The specific name buckleyi was used to recognize Mr. Buckley, who collected the type series of the species.

Identification: Centrolene buckleyi is one of the few species of glassfrogs found in the highlands of Ecuador (2050–3070 m). It is easily recognized by the presence of a large humeral spine (in adult males), white upper lip, inclined snout in lateral profile, and reduced webbing between fingers (Figure 25 and Figure 26). In life, dorsal surfaces are uniform green, but some individuals have small whitish warts. Centrolene heloderma and C. condor can be confused with C. buckleyi; however, C. heloderma has a pustular dorsal skin (shagreen in C. buckleyi) and C. condor has small light and dark spots on the dorsum (dark spots absent in C. buckleyi). Similar species from other countries include Centrolene hesperia, C. lemniscata (Peru), C. altitudinalis, and C. venezuelensis (Venezuela). We note that, as currently defined, C. buckleyi is a species complex that requires further taxonomic studies.

Figure 25. Centrolene buckleyi in life. (Left): Adult male from locality near Oyacachi, Napo province, 3012 m, MZUTI 763, photo by Eduardo Toral. (Right): Adult male from Guarumales, Zamora Chinchipe province, 2070 m, CJ-11364, photo by Diego Acosta-López.

Comments on type material: The description of Centrolene buckleyi was based on two specimens housed at The Natural History Museum, London (formerly British Museum of Natural History). As noted by Goin [97] and Lynch and Duellman [22], one syntype (BMNH 80.12.5.201; from Pallatanga, Provincia de Chimborazo, Ecuador) is now almost completely macerated in ethanol and almost no bones remain, and the other (BMNH 78.1.25.16; from Intac, Imbabura province, Ecuador) is missing and recent searches by Jeff Streicher and DFCH were unsuccessful. Both localities are on the western versant of the Andes of Ecuador, on the slopes of the Cordillera Occidental. Because Centrolene buckleyi represents a species complex [20,98] (Figure 24), we here designate a neotype for Centrolene buckleyi (KU 202770, adult female) collected from Laguna de Cuicocha (near the syntype locality of Intac = Intag), thereby allowing clear comparisons among populations that might represent independent lineages. The morphological characteristics of the neotype fully correspond to those mentioned by Boulenger [93] in the original description of the species. Additionally, mitochondrial sequences of the neotype are included in the phylogeny shown in Figure 24. The designation of the neotype is justified on the Article 75.3 of the International Code of Zoological Nomenclature [99].

Diagnosis: (1) Vomers lacking teeth; (2) snout round in dorsal aspect, slightly sloping to sloping in lateral profile (Figure 26); (3) tympanum partially or completely hidden under skin, when visible oriented almost vertically, its diameter is 29.0%–38.6% of eye diameter; supratympanic fold moderately heavy; tympanic membrane slightly thinner than skin around tympanum; (4) dorsal skin finely shagreen, males with spicules; (5) pair of round subcloacal warts (Figure 26); (6) anterior half to two-thirds of venter covered by white parietal peritoneum, posterior portion translucent (condition P2–P3); silvery white pericardium; translucent peritoneum covering intestines, stomach, testes, kidneys, gall bladder, and urinary bladder (condition V1); (7) liver with four or five lobes, lacking iridophores (condition H0); (8) in males, humeral spines present; (9) webbing absent between Fingers I, II, and III; reduced webbing between outer fingers: III (2^1/4–3⁻)—(2⁺–2^1/2) IV (Figure 26); (10) webbing formula on foot: I (1^1/2–2⁻)—(2–2^1/4) II (1⁻–1⁺)—(2⁺–2^1/2) III (1⁺–1^2/3)—(2^1/3–3) IV (2^2/3–3)—(1^2/3–2⁻) V; (11) ulnar fold low, ventrolateral margin of arm white; inner tarsal fold evident; outer tarsal fold absent, external ventrolateral margin of tarsus white; (12) concealed prepollex; in males, nuptial pad Type I; (13) Finger II slightly longer than Finger I (Finger I 86.5%–98.7% of Finger II); (14) disc of Finger III of moderate size, about 54.2%–64.5% of eye diameter; (15) in life, dorsum uniform green that may have scattered whitish warts; upper lip white, usually continuing as a white line across the flanks; bones green; (16) in preservative, pericardium and anterior half to two-thirds of ventral parietal peritoneum white, visceral peritoneum translucent, peritoneum around kidneys translucent; (17) iris grey-white with thin black reticulation and a horizontal brown stripe; (18) melanophores mostly absent from fingers and toes, except for a few on Toes IV and V and on base of outer fingers; (19) males call from the upper sides of leaves; two call descriptions available (see Call section); (20) males fight upside down, grasping one another venter to venter; (21) eggs deposited on the upper sides of leaves; females provide short-term parental care; prolonged parental care is absent; (22) tadpoles unknown; egg clutches deposited within male’s territory, but parental care has not been reported; (23) medium body size, SVL in males 26.5–30.9 mm (X = 29.0; n = 25), in females 29.3–34.4 mm (X = 31.2; n = 9).

Figure 26. Centrolene buckleyi. (A) Head in lateral view, adult female, KU 178035. (B) Venter and thighs in ventral view, adult female, KU 178040. (C) Hand in ventral view, adult female, KU 178055. Illustrations by Juan M. Guayasamin.

Description of Neotype: Adult female, SVL 32.6 mm (KU 202770; Figure 27). Head wider than long; head length 28.5% of SVL; snout rounded in dorsal profile, sloping in lateral profile; canthus indistinct; loreal region slightly concave; upper lip white, slightly flared; nostril protuberant, closer to eye than to tip of snout, directed dorsolaterally; internarinal area barely depressed. Eye small (diameter 14% of SVL), directed anterolaterally; transverse diameter of disc of Finger III 50.9% of eye diameter. Supratympanic fold conspicuous; tympanum not visible externally. Vomers lacking teeth, choanae large, round; tongue ovoid, ventral posterior half not attached to floor of mouth and posterior margin notched; vocal slits extending posterolaterally from the posterolateral base of tongue to angle of jaws.

Humeral spine absent; ulnar fold evident, white; relative length of fingers: III > IV > II > I; webbing between Fingers I, II, and III absent, basal webbing for outer fingers: III 2^2/3—2⁺ IV; discs and disc pads expanded, elliptical; subarticular tubercles large, round, simple; numerous fleshy supernumerary tubercles present; palmar tubercle elliptical, simple. Length of tibia 51.5% of SVL; inner tarsal fold evident; outer tarsal fold absent, ventrolateral margin of tarsi white; feet about three-fourths webbed; webbing formula on foot I 1^1/2—2 II 1—2⁺ III 1^1/2—3 IV 3—1^2/3 V; discs on toes elliptical; disc on Toe IV narrower that disc on Finger IV; inner metatarsal tubercle large, ovoid; outer metatarsal tubercle not evident; subarticular tubercles small, round; numerous fleshy supernumerary tubercles evident.

Skin on dorsal surfaces of head, body, and lateral surface of head and flanks shagreen, lacking spicules; throat smooth; venter and lower flanks areolate; cloacal opening directed posteriorly at upper level of thighs; small, white tubercles located immediately posterior to cloacae. In ventral view, pair of enlarged subcloacal tubercles not evident.

Figure 27. Centrolene buckleyi, adult female, neotype, KU 220770. Ecuador, Imbabura province, Isla Grande, Lago Cuicocha, 3070 m. Photos by Juan M. Guayasamin.

Color in life (Figure 25): Dorsal surfaces bright to dark green, sharply demarcated laterally from lower white flanks; throat and most of venter pale green; parietal peritoneum yellowish white; edge of upper lip white; ventrolateral borders of arms and tarsus white; small, white warts posterior to cloacal opening; bones green; grey–white iris with thin black reticulation and a horizontal brown stripe. Some individuals have whitish warts on dorsum (QCAZ 22388, 26031–32) or small white spots on legs and forearms (KU 202770)

Color in ethanol (Figure 27): Dorsum of head and body uniform lavender or with minute whitish or unpigmented spots; limbs cream with slight lavender tone and with or without small white spots; conspicuous white border on the upper lip; dorsally, all fingers, Toes I–III, and most of Toe IV unpigmented; ventrolateral borders of arms and tarsus white; cloacal region mostly unpigmented, except for several white warts posterior to cloacal opening; males with cream nuptial pad on Finger I. Three specimens (QCAZ 26031–32, KU 178042) were dissected to observe coloration of internal organs: Anterior half to two-thirds of ventral parietal peritoneum white; pericardium white; hepatic peritoneum transparent; peritonea covering viscera and kidneys translucent to opaque.

Biology and ecology: During the day, Centrolene buckleyi has been found in terrestrial and arboreal bromeliads near and away from streams in secondary forest and pastures (J. D. Lynch, W. E. Duellman field notes). During the night, Centrolene buckleyi were active on terrestrial bromeliads and vegetation 30–160 cm above streams, lakes, and marshes in primary and secondary forests ([20,22], this work). Lynch and Duellman [22] suggested that C. buckleyi might breed in the same situations as other centrolenids—rapid, mountain streams—but also in bromeliads or in ciénegas; they based this statement on the observation of an egg clutch (KU 170221) on the inner leaf of bromeliad in an area where C. buckleyi was abundant. Males fight dangling upside down while holding onto vegetation with their hind limbs, grasping one another venter to venter (Bolívar et al. 1999). A male (QCAZ 22388) was found on the upper side of a leaf close to another leaf with a clutch of eggs, approximately 160 cm above a stream [20]. Females provide short-term parental care to egg clutches; males do not exhibit parental care [25].

Call (Figure 28): Two descriptions of the call of Centrolene buckleyi are available in the literature [20,100]. The differences between these calls, combined with the non-monophyly of C. buckleyi (Figure 24), clearly show that Centrolene buckleyi, as currently recognized, represents a species complex [20,98]. Below we describe a call from the highlands of Ecuador (MZUTI 763; see Examined Specimens); although the call presented herein is not from the neotype locality, genetically, it clusters together with samples from the type locality; thus, apparently corresponding to the same species. Calls are produced in series, every 45–67 s (mean = 53 ± 9.9; n = 10). Each call has a single, highly pulsed note, with a duration of 0.248–0.288 s (mean = 0.264 ± 13.7; n = 10; Figure 23). Calls have a slight frequency modulation, with an initial dominant frequency at 2816–2941 Hz (mean = 2893 ± 38; n = 10) and a final dominant frequency at 3222–3255 Hz (mean = 3229 ± 12; n = 10). First harmonics are at 5385–6393 Hz.

Figure 28. Call of Centrolene buckleyi, MZUTI 763. Recorded at páramo wetland nearby Oyacachi, 3012 m, Napo province, Ecuador.

Tadpole: Not described.

Distribution (Figure 29): The Centrolene buckleyi species complex occurs through the Andes of Colombia and Ecuador to Huacambamba in the Departamento de Piura in northern Peru at elevations between 2450 and 3300 m ([17,101,102], this work). Records below 2450 m are considered dubious. In Ecuador, Centrolene buckleyi is found along the Cordillera Oriental and Cordillera Occidental of the Andes, and inhabits the Western Montane Forest, Andean Shrub, Páramo, and Eastern Montane Forest ecoregions, with a potential distribution of 44,586 km².

Figure 29. Distribution of the species complex currently recognized as Centrolene buckleyi in Ecuador (yellow dots).

Conservation status: Centrolene buckleyi is listed as Vulnerable at a global level by the IUCN [103]. In Ecuador, Centrolene buckleyi was abundant along the high Andes, but recent fieldwork demonstrates population crashes at historical localities (e.g., Pilaló, Cuicocha, Cashca Totoras, Quito, San Pablo de Atenas, Papallacta) ([91], JMG pers. obs.). The last records of the species in Ecuador are from Bosque Protector Cashca-Totoras in 2002 [91], Morán on February 2017 (Diego Batallas, pers. com., Mario Yánez-Muñoz, pers. com.); Yanayacu Biological Station in 2001–2003 [20], Sigchos (0°42′ S, 78°53′ W; 2080 m) on March 2008, Zamorahuaico, near Loja (3°59′ S, 79°12′ W; 2100 m) on August 2008, and nearby Oyacachi (3012 m) on May 2012. The habitat (Páramo, Andean shrub, and montane forest) occupied by the species is heavily impacted by human activities, mainly agriculture and pasture lands. Considering that that populations declines are widespread in the species complex, we suggest the category of Critically Endangered in Ecuador (IUCN criteria A2c,e).

Evolutionary relationships (Figure 24): As defined herein, Centrolene buckleyi is sister to Centrolene ballux. See Taxonomic Remarks.

Taxonomic Remarks: Centrolene buckleyi is a species complex that requires formal subdivision [20,98]. Moreover, morphologically, Centrolene buckleyi is almost identical to four Andean species, namely C. altitudinalis, C. venezuelensis (Venezuela), Centrolene hesperia, and C. lemniscata (Peru). Myers and Donnelly [104] elevated the Venezuelan populations of Centrolene buckleyi (Centrolenella buckleyi venezuelensis Rivero, 1968 [105]) to the species status, without morphological justification; Señaris and Ayarzagüena [106] provided morphological and acoustic data supporting the recognition of Centrolene venezuelensis. The original descriptions of C. hesperia and C. lemniscata did not include a comparison between these species and C. buckleyi; therefore, a reevaluation of their species status is necessary. Genetic [20,98] and morphological [105,106] differences support the validity of Centrolene altitudinalis; for example, the lower two-thirds of the tympanic annulus is visible in C. altitudinalis, whereas the tympanum is completely or mostly concealed in C. buckleyi, C. lemniscata, and C. hesperia. Also, we call attention to the conspicuous geographical barriers that have the potential of limiting gene flow among these species. The Peruvian C. hesperia and C. lemniscata are isolated from other species by the Huacambamba Depression, and isolated from each other by interandean valleys. Likewise, the Venezuelan C. venezuelensis and C. altitudinalis are isolated from other species by the Táchira Depression. The distribution of C. buckleyi is restricted, mostly, to the Andes of Ecuador and Colombia, but these populations, based on genetic and call data, represent several independent evolutionary lineages (Figure 24). Thus, clearly, further work is necessary within this species complex.

Measurements of neotype (in mm): The morphometric data for the neotype (female, KU 202770) are as follows: SVL = 32.6; tibia length = 16.8; foot length = 15.1; head length = 9.3; head width = 10.3; interorbital distance = 3.3; upper eyelid width = 2.6; internarial distance = 2.8; eye-nostril distance = 2.2; snout-eye distance = 4.3; eye diameter = 4.6; tympanum diameter = —; eye-tympanum distance = —; radio-ulna length = 6.72; hand length = 10.4; Finger I length = 6.3; Finger II length = 6.7; disc of Finger III = 2.3; and Finger III width = 1.2.

Specimens examined: Centrolene buckleyi: Ecuador: Provincia del Azuay: 10 km N Girón (3.0833 S, 79.0833 W; 2750 m), KU 202777; 11.5 km SE Gualaceo (2.9333 S, 78.71667; 2940 m); Sigsig (3.05 S, 78.8 W; 2450), QCAZ 1245. Provincia de Bolívar: Bosque Protector Cashca-Totoras (1.71 S, 78.98 W; 2800–3159 m), QCAZ 740, 21231; Guanujo (1.5667 S, 79.01667 W. 2600 m), KU 182214; San Pablo de Atenas (1.8 S, 79.0667 W), QCAZ 372, 2415–16; Santiago (1.7167 S, 78.9833 W), QCAZ 1531. Provincia de Carchi: near Tulcán (0.8 N, 77.7167 W; 2770 m), KU 118005–08; 14 km SW Tulcán (0.72889 N, 77.7958, 3340 m), KU 164516; 5.7 km NW El Carmelo (0.6908 N, 77.63389 W, 2910 m), KU 178053–67; Santa Bárbara (0.61667 N, 77.5833; 2650 m), KU 190017–19; El Goatal, Morán, 0.5 km from Escuela de Morán via La Cortadera, QCAZ 43110; Morán (0.7686 N, 78.056 W, 2785 m), DBR 187, 193. Provincia de Cañar: 4 km W Ingapirca (2.51667 S, 78.9 W; 3000 m), KU 178077. Provincia de Cotopaxi: near Pilaló (0.933 S, 78.9833 W; 2410 m), KU 178034–50, 202780–83. Provincia de Imbabura: La Delicia (0.0667 N, 78.7 W; 2710 m), KU 178079–81, 180311; Lago Cuicocha (0.3 N, 78.3667 W; 3010 m), KU 138822, 178030–33; S shore of Lago Cuicocha (0.29194 N, 78.35389; 3070 m), KU 202703–74; Isla Wolf in Lago Cuicocha (0.3019 N, 78.366 W; 3070 m), KU 202770–72; Mariano Acosta (0.3 N, 77.9833 W; 3000 m), QCAZ 12172. Provincia de Loja: 13 km E Loja, Abra de Zamora (3.9744 S, 79.1114 W; 2800 m), KU 164511–15, 166321; 2 km SSW Saraguro (3.6397 S, 79.24 W; 2560 m), KU 178068–76; 3.7 km S Saraguro (3.6469 S, 79.245 W; 2800 m), KU 202778–79. Provincia de Morona Santiago: 25.5 km WSW Plan de Milagro (3.21667 S, 78.5 W; 2600 m), KU 202775. Provincia de Napo: 9.2 km ESE Papallacta (0.3761 S, 78.0683 W; 2750 m), KU 178052; 11 km ESE Papallacta (0.3869 S, 78.0569; 2660 m), KU 164507–09; 11.2 km ESE Papallacta (0.388 S, 78.055; 2660 m), KU 178051; 12 km ESE Papallacta (0.39194 S, 78.04944 W; 2630 m), KU 155481–92, 164505–06; 31 km N Jondachi (0.5711 S, 77.869 W; 2190 m), QCAZ 2740; Oyacachi–El Chaco road (0.219 S, 78.044 N; 3012 m), MZUTI 763. Provincia de Pichincha: 9.5 km NW Nono (0.026389 S, 78.6403 W; 2530 m), KU 164510; Hacienda El Beaterio, KU 178078; near Machachi (0.5 S, 78.5667 W; 2950 m), KU 148429–30. Provincia de Zamora Chinchipe: 13.5 km E Loja (3.973 S, 79.107 W; 2800 m), KU 142648; Guarumales (3.9405 S, 78.987 W; 2070 m), CJ-11364.

Localities from the literature: Centrolene buckleyi: Ecuador: Provincia del Azuay: Sinicay, 2560 m, AMNH 17464. Provincia de Chimborazo: Pallatanga (1.9833 S, 78.95 W; 1520 m), BMNH 80.12.5.201. Provincia de Imbabura: Intac (0.4 N, 78.6 W), BMNH 78.1.25.16. Provincia de Zamora Chinchipe: Sabanilla (4.033 S, 79.0 W), AMNH 13530 [22,93].

Centrolene charapita Twomey, Delia, and Castroviejo-Fisher 2014 [19] (Figure 30 and Figure 31).
Centrolene charapita Twomey, Delia, and Castroviejo-Fisher, 2014 [19]. Holotype: MHNCP 13933.
Type locality: “near the village of La Oliva, past the village of Muyo (a larger village roughly 49 km N from Bagua), Amazonas, Peru (5°18′3.86″ S, 78°23′44.57″ W, 682 m)”.

Common names: English: Charapita Glassfrog. Spanish: Rana de Cristal Charapita.

Etymology: The specific name charapita derives from a type of yellow chili pepper (ají charapita), which resembles the dorsal ocelli of the species [19].

Identification: Centrolene charapita is the only glassfrog with yellow dorsal ocelli, scalloped and enameled ulnar and tarsal folds (Figure 30), and relatively large size (adult male SVL = 34.7–37.0).

Figure 30. Centrolene charapita in life. Type series from near La Olivia, 682 m, Departamento Amazonas, Peru. Photos by Santiago Castroviejo-Fisher and Evan Twomey.

Diagnosis: Centrolene charapita is recognized by (modified from Twomey et al. [19]): (1) Vomers with 4–10 vomerine teeth; (2) in Peruvian population, snout truncated in dorsal view and truncated to slightly rounded in profile; in Ecuadorian population, snout round in dorsal view and sloping in profile; (3) tympanum small, partially hidden under skin; supratympanic fold present; (4) dorsal skin smooth with microspicules and low enameled warts on ocelli; venter and thighs coarsely areolate, other ventral surfaces smooth; (5) cloacal ornamentation conspicuous, formed by enameled folded skin (flaps) surrounded by lower warts; pair of enlarged subcloacal warts; (6) anterior third-to-half of parietal peritoneum white, posterior portion transparent (state P2); pericardium, hepatic, renal, and gonadal peritonea transparent, visceral peritoneum white; (7) liver trilobed, lacking iridophores (state H0); (8) humeral spines absent; (9) no webbing between Fingers I and II; webbing formula for outer fingers: II 2⁻—3^1/3 III 2—(1⁺–1^1/3) IV; (10) webbing formula on foot: I (1^1/3–1^1/2)—(2⁻–2) II 1—(2⁻–2^1/3) III (1–1⁺)—(2–2⁺) IV (2–2^1/3)—1 V; (11) scalloped enameled ulnar and tarsal folds, extending from fringe on postaxial edge of Finger IV to elbow, and from fringe on postaxial edge of Toe V to ankle, respectively; (12) concealed prepollex; in males, nuptial pad Type I, not pigmented; (13) when appressed, Finger I about equal length or slightly longer than Finger II; (14) eye diameter about 2.5 times the width of disc on Finger III; (15) in life, dorsal and dorsolateral surfaces covered with yellow or pale green spots of different size (ocelli); upper lip and tip of finger and toes I, II, and III white (enameled); ventral surfaces not pigmented; bones green; (16) in preservative, dorsal surfaces and dorsolateral surfaces with cream spots of different size (ocelli), bearing enameled warts and microspicules set in a lavender-greyish reticulum; upper lip and tip of finger and toes I, II, and III white (enameled); ventral surfaces not pigmented; (17) iris in life off-white to light grey with yellow tones and black dots and reticulation, with black ring delimiting iris; in preservative, iris silvery white with black dots and reticulation, with black ring delimiting iris; (18) in life and preservative, tip of fingers and toes white (enameled), Finger IV and Toes IV and V pigmented, Finger II and Toe III not pigmented, but for a small group of melanophores towards the tip, Fingers and Toes I and II not pigmented; (19) males call from upper side of leaves; call not described; (20) fighting behavior unknown; (21) eggs deposition site unknown; parental care unknown; (22) tadpoles undescribed; (23) large size in adult males; SVL 34.7–37.0 mm (n = 5) in males; females unknown.

Color in life (Figure 30): Modified from Twomey et al. [19]. Dorsal and dorsolateral surfaces green, with yellow or pale green spots of different size (ocelli). Upper lip and discs of fingers and toes I, II, and III white (enameled). Ventral surfaces, flanks, upper arms, Fingers I and II, and Toes I and II not pigmented. Iris delimited by a black ring, background coloration off white to light grey with yellow shades and black dots and reticulation. Anterior third-to-half of parietal peritoneum white, posterior portion transparent. Visceral peritoneum white. Hepatic peritoneum transparent.

Color in ethanol: Obtained from Twomey et al. [19]. Dorsal and dorsolateral surfaces with relatively large cream spots bearing enameled warts and microspicules set in a lavender–greyish reticulum, which is formed by minute melanophores that are darker around the ocelli. Upper lip and tips of fingers and toes I, II, and III white (enameled). Ventral surfaces, flanks, upper arms, Fingers I and II, and Toes I and II not pigmented.

Biology and ecology: Very little is known about Centrolene charapita. Twomey et al. [19] report that, at the type locality, individuals were found perched on riverine vegetation during the night, and that sympatric amphibians included Ameerega trivittata, Bolitoglossa altamazonica, Cochranella erminea, Hyloscirtus sp., Rulyrana mcdiarmidi, Pristimantis aff. acuminatus, Pristimantis sp., and Rhaebo glaberrimus. Parental care is unknown.

Call: Unknown.

Tadpole: Not described.

Distribution (Figure 31): Centrolene charapita is known from the type locality, a stream (5°18′3.86″ S, 78°23′44.57″ W, 682 m) near La Oliva, Peru [19], and Reserva Natural Maycu (4.2° S, 78.6° W, 940–1219 m) in southeast Ecuador.

Figure 31. Distribution of Centrolene charapita in Ecuador (yellow dots).

Conservation status: The conservation status of Centrolene charapita has not been evaluated by the IUCN. In Ecuador, populations are within a private reserve (Reserva Natural Maycu). The whole distribution range of the species is within a mining concession. Based on IUCN criteriaB2, Ba, Bb(iii), we suggest that C. charapita should be considered as Critically Endangered in Ecuador.

Evolutionary relationships (Figure 24): Our phylogeny places Centrolene charapita as the sister species to a clade formed by several Centrolene species. A previous analysis had inferred C. charapita as sister to C. geckoidea [19].

Specimens examined: Centrolene charapita: Ecuador: Provincia de Zamora Chinchipe: Reserva Natural Maycu (4.207° S, 78.630° W, QCAZ 66783; 4.222° S, 78.645° W, QCAZ 66786–87; 4.229° S, 78.616° W, QCAZ 66785; 4.226° S, 78.620° W, QCAZ 66784).

Centrolene condor Cisneros-Heredia and Morales-Mite 2008 [107] (Figure 32 and Figure 33).
Centrolene condor Cisneros-Heredia and Morales-Mite, 2008 [107]. Holotype: QCAZ 37279.
Type locality: “Destacamento Militar Cóndor Mirador, western slope of the Cordillera del Cóndor (03°18′25″ S, 78°23′36″ W, between 1750–1850 m elevation), Provincia de Zamora Chinchipe, República del Ecuador”.

Common names: English: Condor Glassfrog. Spanish: Rana de Cristal del Cóndor.

Etymology: The name of this species is in reference to its type locality, Cordillera del Cóndor, a mountain chain shared by Ecuador and Peru [107].

Identification: Centrolene condor can be distinguished from all other glassfrogs by having a green dorsum with many small yellowish–white flecks and dark bluish-black/brown flecks and punctuations (Figure 32), sloping snout in lateral view, light labial stripe, vomerine teeth, small humeral spine in males, enameled ulnar fold, and enameled metatarsal fold followed by a row of distinct enameled tubercles along the outer edge of the tarsus. Among Ecuadorian centrolenids, only C. pipilata shares a similar dorsal pattern, but C. condor has vomerine teeth (absent in C. pipilata) and a small, curved humeral spine (larger and straight in C. pipilata).

Figure 32. Centrolene condor in life. Ecuador, Zamora Chinchipe province, Paquisha, EPN 11343. Photo by Ana Almendáriz.

Diagnosis: Centrolene condor is diagnosed by the following traits (modified from Cisneros-Heredia and Morales-Mite [107]): (1) Vomerine teeth present; (2) snout subacuminate in dorsal view and sloping in profile; nostrils slightly elevated, producing depression in the internarial area; (3) tympanic annulus visible, vertical, with slight dorsolateral orientation; tympanic membrane not differentiated from surrounding skin; (4) dorsal skin shagreen, but with low warts and abundant spicules; (5) ventral skin granular; subcloacal area also granular with several enameled warts; (6) upper two-thirds of parietal peritoneum covered by iridophores (condition P3), pericardium white, all other peritonea not covered by iridophores (condition V1); (7) liver lobed, lacking iridophores (condition H0); (8) small humeral spines present in adult males; (9) webbing absent between Fingers I and II, basal between II and III, moderate between outer fingers: III 2—2⁻ IV; (10) webbing between toes moderate: I 1^1/2—2⁺ II 1—2⁺ III 1—2 IV 2—1 V; (11) enameled ulnar fold; enameled fringe on edge of toe V that continues into thin enameled metatarsal fold and then into row of enameled tubercles along outer edge of tarsus; (12) nuptial excrescences present, Type I; concealed prepollex; (13) first finger shorter than second; (14) eye diameter larger than width of disc on Finger III; (15) in life, green dorsum with abundant yellowish–white flecks and abundant dark flecks; green bones; (16) in preservative, dorsal surfaces greyish with slight lavender hue and abundant light and dark flecks; (17) in life, iris cream–yellow with fine dark reticulation; in preservative, iris light grey with fine dark reticulation; (18) melanophores absent from fingers and toes, except for few on dorsal surfaces of outer fingers and outer toes; (19) males call from upper side of leaves; call composed by two pulsed notes, with dominant frequency at 2.62–2.97 KHz; (20, 21, 22) parental care, fighting behavior, egg clutches, and tadpoles unknown; and (23) medium body size, SVL in adult males 23.2–27.6 mm (X = 25.4 ± 1.826; n = 5); females unknown.

Color in life (Figure 32): Green dorsum with abundant yellowish–white flecks and dark punctuations. Enameled spots and yellowish–white flecks on arms and legs. Whitish–yellow labial line present. Enameled arm and leg folds. Green bones [107].

Color in ethanol: Dorsal surfaces greyish with slight lavender tint, with abundant light and dark flecks. Anterior two-thirds of ventral parietal peritoneum white, pericardium white, all other peritonea lack white lining (modified from Cisneros-Heredia and Morales-Mite [107]).

Biology and ecology: Centrolene condor is a recently described species and little information is available on its natural history. It is nocturnal and males call from amidst the leaves of riverine vegetation a few centimeters over water in mature elfin forest. The species seems to be fairly common at Loma Tigres Bajo, where about 30 males were heard calling [108]. Tadpoles have a bright red coloration and are benthic [108].

Call: The advertisement call has a duration of 650–700 ms (n = 3); it is similar to a trill, composed of two pulsed notes; the first note has a duration of 435–510 ms and is conspicuously longer than the second (101–102 ms); the time between notes is 66–118 ms, and the dominant frequency is 2.62–2.97 KHz [108].

Tadpole: Not described.

Distribution (Figure 33): Centrolene condor is endemic to Ecuador, from localities in the Cordillera del Cóndor, Ecuador, at an elevation of 1737–2920 m [107,108]. The species is distributed within the Eastern Montane Forest ecoregion. Additional searches in the Cordillera del Cóndor have not yielded additional localities for the species [108].

Conservation status: Centrolene condor is currently listed as Data Deficient by the IUCN [109]. Given that the species is known from few localities in an isolated mountain range (Cordillera del Cóndor) with extensive mining activities and associated deforestation and contamination, we suggest considering the species as Endangered—EN B1ab(iii), at the global and local levels, in agreement with Almendáriz and Batallas [108].

Evolutionary relationships (Figure 24): Terminals identified as Centrolene condor are placed in different positions in the tree. Thus, it is likely that more than one species is found in what is currently recognized as C. condor.

Specimens examined: Centrolene condor: Ecuador: Provincia Zamora Chinchipe: Destacamento Militar Cóndor Mirador, western slope of the Cordillera del Cóndor (03°18′25″ S, 78°23′36″ W, 1750–1850 m), QCAZ 37279.

Figure 33. Distribution of Centrolene condor in Ecuador (yellow dots).

Centrolene geckoidea Jiménez de la Espada 1872 [83] (Figure 34, Figure 35 and Figure 36).
Centrolene geckoideum Jiménez de la Espada, 1872 [83]. Holotype: MNCN 1596.
Type locality: “las riberas del río Napo en el Ecuador”. See comment in Distribution.
Centrolene geckoidea—Barrio-Amorós, Rojas-Runjaic, and Señaris, 2019 [85].

Common names: English: Gecko Glassfrog. Spanish: Rana de Cristal Geco.

Etymology: The specific epithet geckoidea refers to the enormous size of the discs on fingers and toes of this species, which resemble those of gecko lizards (Gekkonidae). For almost 150 years, this species was known as C. geckoideum, but its specific ephited was recently modified to geckoidea [85].

Identification: Centrolene geckoidea is unique among centrolenids in having a giant body size (adult males, SVL 70.2–80.7 mm; females, SVL 61.8–72.9 mm) and webbing between the two innermost fingers (Figure 35). Males also have a conspicuous humeral spine (Figure 34), which in some individuals, perforates the skin of the arm. Only C. paezorum, endemic of the high Andes of Colombia, could be confused with C. geckoidea; C. paezorum is known only from a single female and is reported to be smaller (SVL 44.5 mm; see Remarks) and with less hand webbing than C. geckoidea [110].

Figure 34. Centrolene geckoidea in life. Ecuador, Carchi province, Río La Plata (00°48′ N, 78°02′ W; 2525 m), on the Maldonado–Tulcán road, DHMECN 0900. Photo by Doug Wechsler (25 July 1988).

Diagnosis: (1) Each vomer with four or five teeth; (2) snout truncated in dorsal and lateral profiles (Figure 35); (3) tympanum visible and small, oriented almost vertically, its diameter 40.3%–50.0% of eye diameter; supratympanic fold moderate; tympanic membrane completely pigmented, differentiated from surrounding skin; (4) dorsal surfaces of males and females covered with warts, spicules evident only in males; (5) ventral surfaces of thighs below vent lacking pair of enlarged warts; (6) anterior two-thirds or the entire ventral parietal peritoneum white (conditions P3–P4); silvery white pericardium; no iridophores in peritonea covering intestines, stomach, testes, kidneys, gall bladder, and urinary bladder (condition V1); (7) liver tetralobed, lacking iridophores (condition H0); (8) in males, humeral spines present; (9) webbing present, but reduced between Fingers I, II, and III; extensive webbing between Fingers III and IV; hand webbing formula I 2—(2–2⁺) II (1–1⁺)—(2^3/4–3⁺) III (1^1/3–1^2/3)—1 IV (Figure 35); (10) webbing formula on foot I 0⁺—(0⁺–1) II 0⁺—(0⁺–1) III 0⁺—(1–1^1/2) IV (1–1^1/2)—0⁺ V; (11) ulnar fold with small, white tubercles; inner tarsal fold low, short; outer tarsal fold low, with small, white tubercles; (12) concealed prepollex; in males, nuptial pad Type I; (13) Finger I about same length as Finger II (Finger I 91.7%–100.0% of Finger II); (14) disc of Finger III wide, about 115%–130% of eye diameter; (15) in life, dorsum dull green to dark grey; upper lip yellow; bones green; (16) in preservative, dorsal surfaces grey to dark grey; (17) iris greenish gold with fine black reticulation; (18) melanophores covering dorsal surfaces of hands and feet; (19) males call from rocks behind waterfalls or within or near spray zones of fast-flowing streams; call loud, high-pitched, trilled whistle that lacks any consistent pattern of amplitude modulation; weakly to moderately pulsed, duration 155–373 ms; emitted infrequently at intervals of 1.48–5.05 min; dominant frequency modulated, 3468–4187 Hz; (20) fighting behavior unknown; (21) black eggs deposited on rocks in spray zones of streams; males provide prolonged parental care; (22) tadpoles dark, labial tooth row formula (LTRF) 2/3, but see below; (23) giant body size, males SVL 70.2–80.7 mm (X = 75.2, n = 12); females, SVL 61.8–72.9 mm (X = 68.1, n = 9).

Figure 35. Centrolene geckoidea. (A) Head in lateral view, male, KU 116447. (B) Drawing of holotype, not to scale [83]. (C) Hand in ventral view, male, KU 116447; drawing by Juan M. Guayasamin. (D) Humeral spine of adult female; modified from Rueda-Almonacid [111]. (E) Humeral spine of adult male; modified from Rueda-Almonacid [111].

Color in life (Figure 34): Dorsum dull green, with enameled greenish or bluish warts; throat greenish yellow; margin of upper lip yellow; venter cream or yellow cream; ulnar, tarsal, and cloacal tubercles creamy white; white flecks on flanks; iris pale greenish gold with fine black reticulation; palpebrum clear (W. E. Duellman field notes, 9 April 1975; this work); bones green [111]. Males and females are dark grey to brownish green during the night [112].

Color in ethanol: Dorsal surfaces of head, body, and limbs cream grey to dark grey with tips of spicules being white; cloacal, ulnar, and tarsal tubercles white; ventral surfaces cream. Anterior two-thirds of the ventral parietal peritoneum white, posterior third transparent; pericardium white; no iridophores in peritonea covering liver, kidneys, and digestive tract.

Biology and ecology: Males and females of Centrolene geckoidea have been found clinging to vertical or overhanging rock surfaces in the spray zone behind waterfalls [112,113,114]. Males have remarkable call-site fidelity. Egg clutches are deposited on rocks within the spray zone and contain 89–112 black eggs, which are glued to the rocks by a brittle jelly unknown in other centrolenids [112,113]; an “empty space” is evident in the center of the attached clutch [113]. Although oviposition has not been observed, Grant et al. [112] suggested that the ova may be deposited as the female rotates through a circle with the head at the center, a scenario that would explain the origin of the “empty space” near the center of the egg mass. During the night, males have been observed sitting on or near up to four clutches of eggs while calling. During the day, males also have been observed sitting near or on clutches on the rockface behind a waterfall or hidden in spaces between rocks [112]. Thus, all observations by Grant et al. [112] suggest that males provide prolonged parental care to clutches.

Call: Grant et al. [112] described the call of Centrolene geckoidea from El Queremal (3.483 N, 76.7 W), Valle del Cauca, Colombia. The information shown below is based on their study. Males call from behind waterfalls or within or near spray zones of fast-flowing streams. Males have call-site fidelity and have been observed vocalizing at the same site for up to a month. The call is a loud, high-pitched, trilled whistle that lacks any consistent pattern of amplitude modulation. The call is weakly to moderately pulsed and has a duration of 155–373 ms. Calls were emitted infrequently at intervals or 1.48–5.05 min; however, males typically call less frequently (i.e., as little as less than once per hour). The dominant frequency is at 3468–4187 Hz, and it is modulated, beginning at low frequencies (3593–3781 Hz) and rising to a maximum of 3718–4187 Hz, after which it falls, ending at 3468–3718 Hz.

Tadpole: Lynch et al. [113] and Rueda-Almonacid [111] provide descriptions and illustrations of the tadpole of Centrolene geckoidea. Lynch et al. [113] briefly described a tadpole (total length of 22.3 mm) as having a typical centrolenid body form; mouth lacking upper tooth rows, but with two evident lower rows (P₁ with a wide medial gap); upper jaw sheath thick with minute serrations; lower jaw sheath thin and poorly keratinized; one row of large subconical marginal papillae borders the oral disc ventrolaterally and posteriorly. Rueda-Almonacid [111] mentioned that tadpoles in Gosner stage 22, one month after hatching, had two incomplete tooth rows on the anterior labium and three tooth rows on the posterior labium. However, three months after hatching, all the tooth rows were lost, and only dermal ridges were evident [111]. The dorsal surfaces of the body and the tail musculature were black, and the venter lacked pigmentation [111].

Distribution (Figure 36): Centrolene geckoidea occurs across the three Andean Cordilleras of Colombia (Cordillera Occidental, Central, and Oriental; in the departments of Antioquia, Valle del Cauca, Caldas, Quindio, Risaralda, Tolima, Boyacá, and Caquetá), south to the Pacific Andean slope of Ecuador (Carchi and Pichincha provinces) at elevations of 1750–2525 m [111,112,113,114,115,116] (Figure 35). In Ecuador, the potential distribution of the species covers an area of 10,579 km² within the Western Montane Forest region.

Jiménez de la Espada [83] mentioned that the species was found at “las riberas del Río Napo en el Ecuador”. During his trip in South America, Jiménez de la Espada visited several eastern Andean localities (e.g., Basin of Río Quijos, San José de Moti, Cordillera de Guacamayos, Cosanga) before reaching the lowlands near the Río Napo. It has been suggested that the type locality is in error [110], but the holotype of C. geckoidea could have been collected in the headwaters of the Napo river, or at one of the eastern Andean localities [17].

Figure 36. Distribution of Centrolene geckoidea in Ecuador (yellow dots).

Conservation status: Globally, Centrolene geckoidea is listed by the IUCN as a Critically Endangered species [117]. In Ecuador, the species has a severely fragmented distribution, and evidence of population declines. Intensive fieldwork in historical localities such as Quebrada Zapadores has failed in finding the species [91] (JMG, pers. obs.; DFCH, pers. obs.). The last reports of C. geckoidea in Ecuador are from Río La Plata (00°48′ N, 78°02′ W; 2525 m), on the Maldonado–Tulcán road, on 25 July 1988, and from Bosque Protector Río Guajalito in January to May between 1998 and 1999 [17]. A similar situation exists for populations of C. geckoidea found at Valle del Cauca in the Cordillera Occidental (Colombia), as they have not been seen for more than 10 years (W. Bolivar and J.J Sarria-Ospina, pers. com.).

Evolutionary relationships (Figure 24): The molecular tree reported by Twomey et al. [19] places Centrolene geckoidea as the sister species of C. charapita. However, our inferred tree recovers C. geckoidea as sister to all other Centrolene species. Since C. geckoidea is the type species for the genus Centrolene, its phylogenetic placement has fundamental taxonomic implications.

Remarks: The external morphology, osteology, and myology of Centrolene geckoidea was studied in detail by Rueda-Almonacid [111]. Given the remarkable morphological similarly between C. geckoidea and C. paezorum, we consider the possibility that C. paezorum represents a junior synonym of C. geckoidea. Differences between these species include the absence of vomerine teeth in C. paezorum, smaller size, and reduced webbing [110]. Also, the two species occupy different elevations; C. geckoidea is found at elevations between 1750–2525 m, whereas C. paezorum occurs at 3030 m. Additional specimens from the type locality of C. paezorum (Colombia: Departamento del Cauca: km 55–56 on the Popayán–Inza road, 3030 m) are necessary to clarify its status. Until recently, the holotype of Centrolene geckoidea was thought to be lost [118], but in a recent publication, González-Fernández [119] provided dorsal and ventral photographs of the holotype (MNCN 1596).

Specimens examined: Centrolene geckoidea: Ecuador: Provincia de Pichincha: 1 km SW of San Ignacio (0.4486 S, 78.7478 W, 1920 m), KU 178015–17; Quebrada Zapadores, 5 km ESE of Chiriboga on Chiriboga–Quito road (0.245278 S, 78.7261 W; 2010 m), KU 164492; 9 km SE of Tandayapa (0.0167 S, 78.683 W; 2150 m), KU 164490–91. Provincia del Carchi: Río La Plata, on the Maldonado-Tulcán road (0.8 N, 78.033 W; 2525 m), DHMECN 0900.

Centrolene heloderma (Duellman, 1981 [120]; Figure 37, Figure 38, Figure 39 and Figure 40).
Centrolenella heloderma Duellman, 1981 [120]. Holotype: KU 164715.
Type locality: “Quebrada Zapadores, 5 km east-southeast of Chiriboga, 2010 m, Provincia de Pichincha, Ecuador (00°17′ S, 78°47′ W)”.
Centrolene helodermum—Ruiz-Carranza and Lynch, 1991 [6].
Centrolene heloderma—Guayasamin, Castroviejo-Fisher, Trueb, Ayarzagüena, Rada, and Vilà, 2009 [1].

Common names: English: Warty Glassfrog. Spanish: Rana de Cristal Verrugosa.

Etymology: The specific epithet heloderma combines the Greek words “helos” (wart) and “derma” (skin) and is used in allusion to the dorsal texture of the species.

Identification: Centrolene heloderma differs from all glassfrogs by having a green pustular dorsum and males with humeral spines (Figure 34). Centrolene heloderma mostly resembles Centrolene buckleyi; both species have humeral spines in males, snout inclined in lateral profile, and a narrow white labial stripe. The most conspicuous difference between these species is that Centrolene buckleyi lacks a pustular dorsal skin. Further, C. buckleyi differs by having a concealed tympanum (completely visible in C. heloderma; Figure 37).

Figure 37. Centrolene heloderma from Reserva Las Gralarias, Ecuador. (A,B) Adult male. (C) Egg clutch. (D) Egg clutch parasitized by larvae of Drosophilidae fly. Photos by Jaime Culebras.

Diagnosis: (1) Vomers lacking teeth; (2) snout subacuminate in dorsal profile, inclined anteriorly from nostrils to margin of lip in lateral profile (Figure 37 and Figure 38); (3) tympanum completely visible, oriented almost vertically, relatively large, its diameter 38.5%–44.5% of eye diameter; supratympanic fold moderate; tympanic membrane pigmented, clearly differentiated from surrounding skin; (4) dorsal surfaces of males and females pustular; in males, minute spicules evident only on flanks and tympanic region; (5) pair of enlarged subclocacal warts; (6) anterior three-fourths of ventral parietal peritoneum with white iridophores (condition P3); white pericardium; no iridophores in peritonea covering intestines, stomach, kidneys, testes, gall bladder, and urinary bladder (condition V1); (7) liver tetralobed, lacking iridophores (condition H0); (8) in males, humeral spines present; (9) webbing absent between Fingers I, II, and III; moderate webbing between Fingers III and IV; webbing formula III (2–2^1/2)—(1^2/3–2⁺) IV (Figure 37); (10) webbing formula on foot: I (1^1/4–1^1/2)—2 II (1–1⁺)—(2⁺–2^1/4) III (1–1⁺)—(2–2⁺) IV (2⁺–2^1/2)—(1–1^1/4) V; (11) external ulnar fold evident, white; inner tarsal fold low, short; outer tarsal fold long, with low white tubercles; (12) concealed prepollex; in males, nuptial pad Type I; (13) Finger II slightly longer than Finger I (Finger I 91.0%–95.5% of Finger II); (14) disc of Finger III of moderate width, about 59.3%–74.2% of eye diameter; (15) in life, dorsum green with green to bluish white warts; upper lip white; bones green; (16) in preservative, dorsal surfaces dull lavender, sometimes with minute white or cream spots; (17) in life, iris yellow to pale golden yellow with fine black reticulations; (18) melanophores mostly absent from fingers and toes, except for some on Finger IV, and Toes IV and V; (19) males call from upper sides of leaves near streams; call short (133–188 ms, mean = 161 ms, SD = 15.4 ms), with two notes per call; notes strongly pulsed; dominant frequency at 4393–4823 (mean = 4682, SD = 104) Hz; (20) fighting behavior unknown; (21) egg placed on the upper surfaces of leaves; parental care unknown; (22) tadpoles unknown; (23) medium body size, male SVL 26.8–31.5 mm (X = 29.0, n = 17); in one female, SVL 32.3 mm.

Figure 38. Centrolene heloderma, male, KU 164718. (A) Head in ventral view. (B) Hand in ventral view. Illustrations by Juan M. Guayasamin.

Color in life (Figure 37): Dorsum yellow green to dark green with green to bluish white pustules; margin of lip whitish yellow; ventral parietal peritoneum whitish yellow; throat pale greenish yellow; cloacal, heel, and ulnar tubercles white; heart not visible; bones green; iris yellow to pale golden yellow with fine black reticulations [120].

Color in ethanol: Dorsal surfaces of head, body, forearms, thighs, and shanks dull lavender; other surfaces dull cream; flanks white; margin of upper lip and cloacal tubercles white (Duellman, 1981). White parietal peritoneum covering the anterior three-fourths of the venter; pericardium white; no iridophores in peritonea covering liver, intestines, stomach, kidneys, and urinary bladder.

Biology and ecology: During the rainy season (December–April), at night, Centrolene heloderma has been found active on vegetation near streams in primary and disturbed forest. Occasionally, males have been observed away from streams, probably looking for new territories [87]. Males call from the upper surfaces of leaves near streams. Egg clutches, with 25–29 brownish eggs, are placed on the upper side of leaves near streams [87]. At Reserva Las Gralarias, C. heloderma is abundant at Five-frog Creek, but has also been observed at Ballux Creek, Heloderma Creek, and Hercules Creek [88,92]. Parental care is unknown.

Call (Figure 39): The following description is based on the analysis of 58 notes contained within 29 calls from 3 individuals (LBE-C-001, 012, 015); individuals are from Río Alambí (2390 m), and Reserva Las Gralarias (1822–1864 m), in Pichincha province, Ecuador. The typical advertisement call is relatively short (range = 133–188 ms, mean = 161 ms, SD = 15.4 ms), and has two notes per call. Notes are strongly pulsed. The first note is longer than the second note (first note duration: Range = 38–76 ms, mean = 60.2 ms, SD = 9.89 ms; second note duration: Range = 17–44 ms, mean = 27.8 ms, SD = 6.283 ms). The two notes are also separated by an internote duration of 50–99 (mean = 72.9, SD = 9.5) ms. Notes are pulsed and similarly variable in number, with 2–7 (mean = 3.741, SD = 1.906) amplitude peaks throughout the note. The first note tends to have more pulses than the second note. Pulses within a note have a rate of 45.5–117.6 (mean = 83, SD = 15) pulses per second. Notes generally have their peak amplitude in the first 50% of the note (relative peak time: Range = 0.032–0.8697, mean = 0.273, SD = 0.242), but infrequently the first note can have its peak amplitude in the last 50% of the note. Frequencies between both notes are also highly similar. The dominant frequency of a note measured at peak amplitude is 4393–4823 (mean = 4682, SD = 104) Hz and is contained within the fundamental frequency. The fundamental frequency has a lower limit of 4221–4651 (mean = 4513, SD = 138) Hz and a higher limit of 4651–5082 (mean = 4875, SD = 104) Hz.

Figure 39. Call of Centrolene heloderma (LBE-C-012) recorded from Reserva Las Gralarias, Pichincha province, Ecuador. Note that each call has two distinctive pulsed notes.

Tadpole: Not described.

Distribution (Figure 40): Centrolene heloderma occurs on the Pacific slopes of the Cordillera Occidental in Colombia (departments of Antioquia, Cauca, Valle del Cauca, and Risaralda) south to the Tandayapa and Saloya Valleys, in Ecuador (provinces of Imbabura, Pichincha, and Santo Domingo de los Tsáchilas) at elevations of 1850–2575 m ([87,120,121], this work). In Ecuador, Centrolene heloderma has been recorded from seven localities at elevations between 1960 and 2575 m ([87,120,121]) in this work and has a potential distribution of 1067 km², within the Western Montane Forest ecoregion.

Figure 40. Distribution of Centrolene heloderma in Ecuador (yellow dots).

Conservation status: Globally, Centrolene heloderma is currently listed as Vulnerable by the IUCN [122]. In Ecuador, major threats include habitat loss, introduction of exotic predatory fish (Trout), climate change, and emerging diseases [87,123]. The type locality (Quebrada Zapadores), as well as nearby localities, have been visited numerous times in the last years (2000–2015; wet and dry seasons) with no new records of the species [91] (JMG, pers. obs.; DFCH, pers. obs.); in this area, the last record of the species dates from March 1979, when three individuals where collected at Quebrada Zapadores. In 2006, an individual apparently assignable to C. heloderma was photographed in the western versant of the Pichincha volcano (M. H. Yánez-Muñoz, pers. com.). On March 2009, C. heloderma was discovered at Reserva Las Gralarias (0.00806 S, 78.72433 W, 1852 m), Pichincha Province, where it maintains three nearby, reproductive populations. During September 2015, a population of C. heloderma was found at Cordillera de Toisán. Krynak et al. [121] report on a relatively large population (20+ individuals) at Río Alambi observed on 24 April 2017. The potential distribution of the species in Ecuador is 1067 km², 34% of which is affected by human activities. The amphibian chytrid fungus Batrachochytrium dendrobatidis has been found infecting C. heloderma at Reserva Las Gralarias, but no recent declines have been observed [92]. Given the current information of the species, we suggest that it should be considered as Endangered in Ecuador, following IUCN criteria B2, Ba, Bb(iii).

Evolutionary relationships (Figure 24): Centrolene heloderma is sister to a clade formed by several Centrolene species.

Specimens examined: Centrolene heloderma: Ecuador: Provincia de Imbabura: Cordillera de Toisán (0.50276° N, 78.5515° W; 2575 m), MZUTI 4234. Provincia de Pichincha: Quebrada Zapadores, 5 km ESE of Chiriboga on Chiriboga–Quito road (0.2453° S, 78.726° W, 2010 m), KU 164714–15; 9 km SE Tandayapa (0.01667° S, 78.683° W, 2160 m); 8.6 km SE Tandayapa (0.0333° S, 78.7° W, 2000 m), USNM 211218; 13.1 km SW Nono (0.0025° S, 78.659° W, 2140 m), MCZ 97834, USNM 211216–17; Reserva Las Gralarias (0.00806° S, 78.72433° W, 1852 m), QCAZ 40200. Provincia de Santo Domingo de los Tsáchilas: 14 km west of Chiriboga on Chiriboga–Santo Domingo road (0.2653° S, 78.848° W, 1960 m), KU 164716–21.

Centrolene huilensis (Ruiz-Carranza and Lynch, 1995 [26]) (Figure 41).
Centrolene huilense Ruiz-Carranza and Lynch, 1995 [26]. Holotype: ICN 7462.
Type locality: “Colombia, Departamento de Huila, Municipio San José de Isnos, 1 km NW Isnos, vertiente oriental de la Cordillera Central, 1°57′ Latitud N, 76°15′ W Greenwich, 2190 m”.
Centrolene huilensis—Barrio-Amorós, Rojas-Runjaic, and Señaris, 2019 [85].

Common names: English: Huila Glassfrog. Spanish: Rana de Cristal de Huila.

Etymology: The specific epithet huilensis refers to the type locality of the species, within the Huila Department, Colombia [26].

Identification: Centrolene huilensis (Figure 41) can be differentiated from other glassfrogs by having a green dorsum with a combination of dark green to dark lavender and white spots, white tubercles on the ventrolateral edges of the Finger V, forearm, elbow, Toe V, tarsus, and heel, a humeral spine in adult males, and a relatively large body size (SVL = 23.6–26.7 mm in males; SVL = 28.7 mm in 1 female). Species with a similar dorsal color pattern include Centrolene peristicta, C. daidalea, C. condor, C. lynchi, C. muelleri, C. pipilata, C. savagei, C. solitaria, and Nymphargus truebae. Body size of C. huilensis is larger than C. peristicta, C. pipilata, C. savagei, and C. solitaria (in C. peristicta, male SVL = 17.9–22.0 mm, female SVL = 20.8–20.9 mm; in C. pipilata, male SVL 19.7–22.6 mm, female SVL 22.6–23.6 mm; in C. savagei, male SVL = 23.3–23.9 mm, female SVL = 19.8–22.6 mm; in C. solitaria, holotype male SVL = 19.3). Males of C. huilensis have humeral spines, which are absent in males of C. daidalea, C. savegei, C. solitaria, and N. truebae. Additionally, some of the species are located in different biogeographic regions; C. peristicta and C. lynchi are found on the Pacific slopes of the Andes of Ecuador and Colombia (C. peristicta is also found on the western slopes of the Cordillera Central, Colombia), and C. muelleri and N. truebae are only known from the Andes of Peru.

Figure 41. Centrolene huilensis in life. Ecuador, Napo province, Yanayacu Biological Station, QCAZ 45905. Photos by Santiago R. Ron (BioWebEcuador).

Diagnosis: (1) Vomers lacking teeth; (2) snout short, round in dorsal aspect, slightly sloping in lateral view; (3) tympanum moderate, oriented almost vertically, with slight lateral and posterior inclinations, its diameter 31%–40% of eye diameter; tympanic annulus completely visible, except for upper border; supratympanic fold evident; tympanic membrane translucent and pigmented as surrounding skin; (4) dorsal skin shagreen, males with low warts and spicules uniformly distributed; (5) pair of enlarged subcloacal warts (Figure 15); (6) anterior 40%–60% of the ventral parietal peritoneum white, posterior portion translucent (condition P3); white pericardium; no iridophores in peritonea covering intestines, stomach, kidneys, gall bladder, and urinary bladder (condition V1); (7) lobed liver, lacking iridophores (condition H0); (8) males with conspicuous humeral spines; (9) webbing absent between Fingers I, II, and III; moderate between outer fingers; webbing formula: III (2⁺–2^1/2)—(2–2⁺) IV; (10) webbing between toes extensive; foot three/fourths webbed: I (1–1^2/3)—(1^3/4–2⁺) II (1–1⁺)—(2⁻–2^1/2) III (1–1^1/2)—(2⁺–2^2/3) IV (2–3⁻)—(1⁻–1^1/2) V; (11) ulnar and tarsal folds with white tubercles; (12) concealed prepollex; nuptial pad Type I; (13) Finger I slightly shorter than Finger II (Finger I length 94%–97% of Finger II); (14) disc of Finger III width 51%–63% of eye diameter; (15) in life, dorsum green with small diffuse dark green to dark lavender spots and smaller white spots; bones green; (16) in preservative, dorsum cream to light lavender with dark lavender spots and smaller white spots; (17) in life, iris cream with a slight yellow hue and thin black reticulations, yellowish–cream circumpupilar ring; (18) melanophores absent from dorsal surfaces of fingers and toes, except for Finger IV, and Toes IV and V; (19) males call from upper side of leaves; call undescribed; (20) fighting behavior unknown; (21) eggs deposition site unknown; parental care unknown; (22) tadpoles unknown; (23) small to medium body size; in Colombian populations, adult males, SVL 23.6–26.7 mm (X = 25.1 ± 1.703, n = 7 [26]); in Ecuador, SVL 23.8 in one adult male and 28.7 mm in one adult female.

Color in life (Figure 41): Dorsum green with dark lavender and dark green spots of different sizes, and smaller white spots; upper lip white; region below eye with small white warts; bones green. Upper flanks with same color pattern as dorsum; lower flanks with numerous small white warts. Ulnar and tarsal folds with white tubercles; small white cloacal tubercles. Iris cream with a slight yellow hue and thin black reticulations, yellowish–cream circumpupilar ring.

Color in ethanol: Dorsal surfaces of head, body, and limbs cream to light lavender with dark lavender spots and smaller white spots; margin of upper lip white; region below eye with small white warts; white tubercles just posterior to cloaca. White parietal peritoneum covers anterior 40%–60% of venter; white pericardium; iridophores absent from peritonea covering digestive tract, liver, kidneys, and gall and urinary bladders.

Biology and ecology: In Ecuador, individuals were found active during the night on upper surfaces of leaves along slow-flowing streams. Parental care is unknown.

Call: Not described.

Distribution (Figure 42): Centrolene huilensis is known from two localities, one in the Amazonian slope of the Andes of Ecuador (Yanayacu Biological Station, 2000 m, Napo Province), and the type locality in the Cordillera Central of Colombia (near Isnos, Huila Department) at elevations between 2000–2190 m ([26], this work).

Figure 42. Distribution of Centrolene huilensis in Ecuador (yellow dot).

Conservation status: Globally, Centrolene huilensis is currently listed as Endangered by the IUCN [124]. In Ecuador, assessing the conservation status of this species remains challenging because only a single additional locality (Yanayacu Biological Reserve) has been registered since its description in 1995. Yanayacu Biological Reserve is a relatively well-studied site [20,125] and we have been unable to find it during recent surveys (August 2014, June 2016; January 2017); thus, it is possible that C. huilensis spends most of its time in the canopy and/or is extremely rare. We suggest the that C. huilensis should be placed in the Data Deficient category for Ecuador.

Evolutionary relationships (Figure 24): With the current gene and taxon sampling, Centrolene huilensis is inferred as sister to C. muelleri.

Specimens examined: Centrolene huilensis: Ecuador: Provincia de Napo: Yanayacu Biological Station (0°41′ S, 77°53′ W; 2100 m), QCAZ 37230, 45905.

Centrolene lynchi (Duellman, 1980 [126]; Figure 43, Figure 44, Figure 45 and Figure 46)
Centrolenella lynchi Duellman, 1980 [126]. Holotype: KU 164691.
Type locality: “a stream 4 km northeast (by road) of Dos Ríos, Provincia Pichincha, Ecuador, 1140 m (0°21′ S, 78°54′ W)” (now in Provincia de Santo Domingo de los Tsáchilas).
Centrolene lynch—Ruiz-Carranza and Lynch, 1991 [6].
Centrolenella grandisonae—Lynch and Duellman, 1973 [22].
Centrolenella gemmata Flores, 1985 [46]. Holotype: MCZ 104073. Type locality: “San Francisco de las Pampas, 1500 m in elevation, Provincia Cotopaxi, Ecuador (00°25′ S, 78°57′ W, just NW of junction of Río Las Juritas and Río Toachi)”. New synonymy.
Centrolene gemmatum—Ruiz-Carranza and Lynch, 1991 [6]. Guayasamin, Castroviejo-Fisher, Trueb, Ayarzagüena, Rada, and Vilà, 2009 [1].
Centrolenella scirtetes In part, Duellman and Burrowes, 1989 [86]. Holotype: KU 202720. Type locality: “1.4 km (by road) southwest of Tandayapa (00°07 S, 78°40 W), 1820 m, Provincia de Pichincha, Ecuador”. New synonymy.
Centrolene scirtetes—Ruiz-Carranza and Lynch, 1991 [6]. Guayasamin, Castroviejo-Fisher, Trueb, Ayarzagüena, Rada, and Vilà, 2009 [1].

Common names: English: Lynch’s Glassfrog. Spanish: Rana de Cristal de Lynch.

Etymology: The specific epithet lynchi honors Dr. John D. Lynch, who has made multiple and meaningful contributions to the field of amphibian systematics.

Identification: Centrolene lynchi is distinguished from most glassfrogs by having, in life, a green dorsum with minute yellowish–white flecks and diffuse small black spots (Figure 43). Among centrolenids found on the Pacific slopes of the Andes, only C. peristicta has a similar dorsal color pattern; however, body size is clearly different between the two species (in males, SVL 23.3–26.5 mm in C. lynchi; 17.9–21.2 mm in C. peristicta). The Colombian C. antioquiensis lacks black spots on the dorsum and has a white gastrointestinal peritoneum. Centrolene lynchi is also similar to the Colombian C. quindianum, which invariably has iridophores on the digestive tract (iridophores absent in C. lynchi). Other species, such as Nymphargus truebae (from the Amazonian slopes of the Peruvian Andes) and N. garciae (from the Cordillera Central of the Colombian Andes and the Amazonian slope of the Ecuadorian Andes), have a dorsal coloration similar to that found in C. lynchi, but lack humeral spines in males and webbing between Fingers III and IV.

Diagnosis: (1) Vomerine teeth absent; (2) snout round or truncated in dorsal aspect and truncated to slightly inclined in lateral profile; (3) tympanic annulus visible, oriented almost vertically, with slight lateral and posterior inclinations, its diameter about 31.3%–39.4% of eye diameter; supratympanic fold evident; tympanic membrane translucent, partially pigmented, differentiated from surrounding skin; (4) dorsal skin shagreen in males and females, males have low, white warts, and spicules and spiculated warts on sides of head; (5) pair of enlarged subcloacal warts (Figure 15); (6) anterior half to two-thirds of ventral parietal peritoneum white, remaining posterior portion transparent (condition P2–P3); white pericardium; no iridophores in peritonea covering intestines, stomach, and kidneys; transparent peritoneum around gall bladder and urinary bladder (condition V1); (7) liver with four clearly defined lobes, lacking iridophores (condition H0); (8) males with conspicuous humeral spines; (9) webbing absent between Fingers I and II, absent or greatly reduced between Fingers II and III, and moderate between outer fingers; hand webbing formula III (2–2^1/4)—(2⁻–2⁺) IV; (10) webbing between toes extensive; foot webbing formula: I (1–1⁺)—(2–2⁺) II (1–1⁺)—(2–2⁺) III (1–1⁺)—(2–2⁺) IV 2⁺—(1–1⁺) V; (11) ulnar fold present, white; external tarsal fold absent, internal tarsal fold short, low; (12) concealed prepollex, except in a few individuals; nuptial pad Type II; (13) Finger I slightly shorter than Finger II or about equal its length (Finger I 94.5%–100.0% of Finger II); (14) disc of Finger III width about 48.4%–57.0% of eye diameter; (15) in life, dorsum dull green with minute yellowish–white warts and small diffuse black spots (Figure 43); green bones; (16) in preservative, dorsum lavender with numerous minute white spots and larger dark lavender spots (Figure 44); (17) in life, iris greyish–white, with a yellow hue around the pupil; (18) melanophores mostly absent from fingers and toes, except for a few on base of Fingers III and IV, and along Toes IV and V; (19) males call from upper sides of leaves; call is relatively short and consists of a tonal note followed by one to three peaked notes; notes separated by 9.0–138.0 ms; mean tonal note dominant frequency 5296 Hz (SD = 58, range = 4995–5599 Hz); mean peaked note dominant frequency 5264 Hz (SD = 72, range = 4995–5513); notes lack frequency modulation; (20) males fight while hanging upside down, grasping one another venter to venter; (21) eggs placed on upper surfaces of leaves; females provide short-term parental care; prolonged parental care absent; (22) tadpoles unknown; (23) small body size; male SVL 23.3–26.5 mm (X = 24.7, n = 22); in two females SVL 24.6–25.0 mm.

Figure 43. Centrolene lynchi in life from Reserva Las Gralarias, Pichincha province, Ecuador. (A) Adult male, not collected. (B) Adult male in ventral view, QCAZ 40192; photo by Luis A. Coloma. (C) Fight between males; photo by Henry Imba and Rebecca Abuza. (D) Egg clutch.

Color in life (Figure 43): Dorsum yellow green with minute yellowish–white warts (spiculated in males) and small and diffuse black spots; bones green; throat and venter cream white; fingers and toes dull yellow; upper lip white; ulnar and outer tarsal folds with thin white line or low white tubercles; flanks and venter cream white; small white tubercles just posterior to cloaca. White parietal peritoneum covers anterior half to two-thirds of venter; white pericardium; visceral and hepatic peritonea lacking iridophores. Iris greyish white with black reticulation, and a yellow hue surrounding the pupil.

Color in ethanol (Figure 44): Dorsal surfaces of head, body, and limbs lavender with minute white warts and small black spots; margin of upper lip white; region below eye with small white warts; small white tubercles just posterior to cloaca. White parietal peritoneum covering the anterior half to two-thirds of the venter; pericardium white; no iridophores in peritonea covering liver, intestines, stomach, kidneys, gall bladder, and urinary bladder.

Figure 44. Type material of Centrolene lynchi and C. gemmata in preservative. (A,B) C. gemmata, MCZ 104077, paratype. (C) C. lynchi, KU 164698, paratype, dorsal view. (D) C. lynchi, KU 164691, holotype, ventral view.

Variation: The dorsum of most examined specimens except one (KU 164695) have spicules and spiculated warts; it is possible that this variation is a result of the reproductive condition of this particular male individual. Females of Centrolene lynchi lack spicules, normal sexual dimorphism in Centrolenidae.

Biology and ecology: The following description is from Dautel et al. [50]. At Reserva Las Gralarias (Lucy’s Creek), females place clutches of 21–24 neon green eggs on vegetation in males’ territories (n = 4, mean = 22.25, SD = 1.08); most egg clutches are placed on top of large leaves or ferns above a fast-flowing stream at a height of 165–600 cm (n = 14, mean height = 313.3 cm, SD = 78.7). Dautel et al. [50] suggested that males provide long-term parental care; however, a more detailed study by Delia et al. [25] showed that only females provide parental care (short term) and that prolonged parental care is absent in the species.

Calls (Figure 45): The following description is from Dautel et al. [50]. The typical advertisement call is relatively short and consists of a tonal note followed by one to three peaked notes. The first note is tonal and generally longer than the following notes, which are pulsed and show clear frequency peaks. Notes are separated by 9.0–138.0 ms. The dominant frequencies of tonal and peaked notes are similar (mean tonal note dominant frequency = 5296 Hz, SD = 58, range = 4996–5599 Hz; mean peaked note dominant frequency = 5264 Hz, SD = 73, range = 4996–5513) and show no frequency modulation. The aggressive call in this species is markedly different in structure from the advertisement call. It consists of a single short note containing two pulses (n = 5, mean length duration = 150.4 ms, SD = 6.7, range 140.0–156.0 ms), at a dominant frequency significantly lower than the advertisement call (mean frequency = 4892 Hz, SD = 39, range = 4823–4910 Hz); like the advertisement call, the note of the aggressive call is not frequency modulated.

Figure 45. Call of Centrolene lynchi (LBE-C-013) recorded at Reserva Las Gralarias, 1822 m, Pichincha province, Ecuador.

Tadpole: Not described.

Distribution (Figure 46): Centrolene lynchi is known from the Pacific slope of the Cordillera Occidental of the Andes in Ecuador and southern Colombia. In Colombia, the species has been reported in only one locality (Reserva La Planada, 7 km route of Chucunés, 1780 m; as C. scirtetes [86], but see Taxonomic Remarks); specimens cited as C. lynchi by Coloma et al. [126] from Risaralda Department, Colombia, actually corresponds to C. quindianum (Marco Rada, pers. obs.). In Ecuador, Centrolene lynchi is known from seven localities from the Pacific slope of the Cordillera Occidental of the Andes at elevations of 1140–1852 m (see Specimens Examined), with a potential distribution of 1442 km². The habitat of the species in Ecuador is within the Western Foothill Forest and the Western Montane Forest regions.

Conservation status: Globally, Centrolene lynchi is currently listed as Endangered by the IUCN [126]. Arteaga et al. [87] also suggested the Endangered category for Ecuadorian populations. Although the distribution of the species is larger than previously thought (see Distribution and Remarks), the species has not been found in some of its historic localities in the last 20 years, suggesting that the species has suffered population declines (i.e., San Francisco de Las Pampas). In recent years, reproductive populations of C. lynchi have been found at Reserva Las Gralarias (2009–2019) and at Bosque Protector Río Guajalito (2006) ([87], JMG pers. obs., DFCH pers. obs., Mario Yánez-Muñoz pers. comm.). The amphibian chytrid fungus Batrachochytrium dendrobatidis has been found infecting C. lynchi at Reserva Las Gralarias, but no recent declines have been observed [92].

Figure 46. Distribution of Centrolene lynchi in Ecuador (yellow dots).

Evolutionary relationships (Figure 24): Centrolene lynchi is sister to a clade formed by C. sabini and an unidentified species of Centrolene.

Taxonomic remarks: Centrolene lynchi was described by Duellman [127] from a locality 4 km NE (by road) of Dos Ríos, Provincia de Pichincha. After a few years, Flores [46] described Centrolene gemmata from San Francisco de Las Pampas, Provincia de Cotopaxi. Flores [46] mentioned that Centrolene lynchi differs from Centrolene gemmata mainly by: (i) Having a snout truncated in dorsal view and round in profile (snout round in dorsal view and slightly anteroventrally sloping in C. gemmata); (ii) having slightly more webbing on the hands (webbing formula of IV 2—2 V in Centrolene lynchi; webbing formula of IV (2–2^1/3)—(2⁻–2⁺) V in C. gemmata); and (iii) in overall head shape, with a squatter, more semicircular head shape when viewed from above, with little or no post-cephalic constriction, and nostrils only very slightly protuberant, in contrast to the more elongated, circular head shape of gemmata when viewed from above, with a marked post-cephalic constriction and very protuberant nostrils. We have examined the complete type series of Centrolene lynchi and three paratypes of C. gemmata (MCZ A-104397, A-104074, A-104077) and, as noted by Cisneros-Heredia and McDiarmid [17], find that the differences mentioned above are not consistent. Several of the characteristics present in C. gemmata fall into the variation observed in Centrolene lynchi, and others are the product of preservation artifacts. It is clear that the type material of C. gemmata was poorly preserved, causing dehydration of the specimens, which usually produces changes in head shape (i.e., post-cephalic constriction and protuberant nostrils present in the type series of C. gemmata). The examined type series of C. gemmata has a snout that is truncated to slightly sloping in profile; the exact same variation is present in the type material of Centrolene lynchi. Also, the hand webbing formula in the two species are nearly identical [III (2–2^1/4)—(2⁻–2⁺) IV in Centrolene lynchi, and III (2–2^1/3)—(2⁻–2⁺) IV in C. gemmata]. Therefore, none of the characters listed by Flores (1985) to differentiate C. gemmata from Centrolene lynchi are valid. Based on the evidence mentioned above, we consider these to represent the same species and place Centrolene gemmata in the synonymy of Centrolene lynchi.

Duellman and Burrowes [86] described Centrolenella scirtetes from a locality 1.4 km SW Tandayapa (Ecuador, male holotype KU 202720) and from Reserva La Planada (Colombia, two female paratypes IND-AN 1405 and 1533). For unknown reasons, C. scirtetes was never compared with specimens of Centrolene lynchi. As pointed out by Cisneros-Heredia and McDiarmid [17], the holotype of C. scirtetes is undistinguishable from the holotype of C. lynchi (KU 164691). Consequently, herein we formally place C. scirtetes, as defined by its holotype, in the synonymy of Centrolene lynchi. We have not examined the paratypes of C. scirtetes, but material from Colombia identified as C. scirtetes (ICN 12172–74) and resembling the description and photograph of the paratypes provided by Duellman and Burrowes (1989) are indistinguishable from individuals of Nymphargus griffithsi, in which the dorsum has black flecks and the humeral crista ventralis presents a distal prolongation. Accordingly, we conclude that the Colombian material assigned to C. scirtetes is in fact representative of N. griffithsi.

Specimens examined: Centrolene lynchi: Ecuador: Provincia de Cotopaxi: San Francisco de Las Pampas, just NW of junction of río Las Juritas and río Toachi (0.433 S, 78.9667 W; ca. 1500 m), MCZ A-104397, A-104074, A-104077; Provincia de Pichincha: 1.4 km SW of Tandayapa (0.033 S, 78.7667 W; 1820 m), KU 202720; Tandapi (0.4164 S, 78.7989 W; 1460 m), KU 118036, 118047–50, 178095–104; 2.1 km E Tandapi (0.4258 S, 78.7853 W; 1500 m), MCZ A-93313–14, 95742; Reserva Las Gralarias (0.01675 S, 78.73165; 1852 m), QCAZ 40191–2, 40194. Provincia de Santo Domingo de los Tsáchilas: stream 4 km northeast (by road) of Dos Ríos (0.3028 S, 78.8678 W; 1140 m), KU 164691 (holotype), KU 164692–99 (paratypes); 14.4 km ENE La Palma on the road La Palma-Chiriboga (0.25 S, 78.846 W; 1380 m), MCZ A-91455.

Centrolenella medemi Cochran and Goin, 1970 [96]. Holotype: USNM 152277.
Type locality: “Puerto Asís, upper Río Putumayo, [Comisaría] Putumayo, Colombia” (apparently in error; see Distribution).
Centrolene medemi—Ruiz-Carranza and Lynch, 1991 [6].
“Centrolene” medemi—Guayasamin, Castroviejo-Fisher, Trueb, Ayarzagüena, Rada, and Vilà, 2009 [1].

Common names: English: Medem’s Glassfrog. Spanish: Rana de Cristal de Medem.

Etymology: The epithet medemi honors Dr. Fred Medem, who discovered the species [96].

Identification: “Centrolene” medemi is unique among Ecuadorian glassfrogs by having a dark olive-green dorsum with large greenish–cream spots (Figure 47), wide disc on Finger III (>80% of eye diameter), small tympanum (<20% of eye diameter), and fully webbed foot. Adults are robust and relatively large (SVL 25.5–44.3 mm).

Figure 47. “Centrolene” medemi in life. Adult female (KU 164493) from 2 km SSW of junction between Río Reventador and Baeza-Lumbaqui road, Ecuador. Photo by William E. Duellman.

Diagnosis: (1) Teeth on dentigerous process of the vomer present or absent, each process bearing zero to four teeth; (2) snout round in dorsal profile, truncated to slightly protruding in lateral profile (Figure 48); (3) tympanum small, tympanum diameter 17.1%–18.5% of eye diameter, dorsal third of tympanum covered by supratympanic fold, tympanic membrane pigmented and not clearly differentiated from surrounding skin; (4) dorsal surfaces of males and females smooth to shagreen; males with small spicules on dorsum and flanks; (5) pair of slightly enlarged subcloacal warts; (6) anterior half of the ventral parietal peritoneum white (condition P2); silvery white pericardium; no iridophores in peritonea covering the intestines, stomach, testes, kidneys, gall bladder, and urinary bladder (condition V1); (7) liver tetralobed, two large ventral lobes covering two smaller lobes; hepatic peritoneum lacking iridophores (condition H0); (8) in males, small humeral spines present; (9) hand webbing: Webbing between Fingers I, II, and III absent or basal, extensive webbing between Fingers III and IV; webbing formula as follows: II (1⁺–2)—3⁺ III (1^1/4–2)—(0–1^1/4) IV; (10) fully webbed foot: I (0–0⁺)—(0⁺–1) II (0–0⁺)—(0–1) III (0–0⁺)—(0⁺–1) IV (1⁻–1)—0⁺ V; (11) ulnar and tarsal folds low; (12) concealed prepollex; in males, nuptial pad Type I; (13) Finger I slightly shorter than Finger II (Finger I about 91%–98% length of Finger II); (14) disc of Finger III large, 80%–91% of eye diameter; (15) in life, dorsal surfaces of head, body, and limbs olive green to greyish brown with large (up to 2.8 mm) cream spots (Figure 47); bones bluish green or green; (16) in ethanol, dorsal surfaces of head, body, and limbs pale brown with large cream spots; (17) iris greenish brown with black reticulation; (18) melanophores covering dorsal surfaces of fingers and toes; (19) calling behavior unknown; (20) fighting behavior unknown; (21) eggs deposited on rocks along streams; parental care unknown; (22) tadpoles unknown; (23) medium body size; in adult males, SVL 25.5–30.8 mm; in adult females, SVL 34.7–44.3 mm.

Figure 48. “Centrolene” medemi. (A) Head in lateral view, KU 164493. (B) Hand in ventral view, KU 164494. Illustrations by Juan M. Guayasamin.

Variation: The size of the nuptial pad varies in males and can be restricted to the dorsolateral area of the base of Finger I or extend until reaching the typical Type I morphology.

Color in life (Figure 47): At night, dorsum black with bluish green spots; venter dull blue. By day, dorsum dark olive-brown with pale green spots; flanks cream. Lining of mouth and tongue pale blue. Bones green. Iris dull greyish–bronze with minute black flecks. Webbing pale yellowish tan (W. E. Duellman field notes, 19 March 1975).

Color in ethanol: Dorsal surfaces of head, body, and limbs pale brown with large cream spots. White lining on the anterior half of the ventral parietal peritoneum; silvery white pericardium; no iridophores in peritonea covering the liver, intestines, stomach, testes, kidneys, gall bladder, and urinary bladder.

Biology and ecology: During the night, adults have been found on rocks, in rock crevices, and on rock cliffs along streams; juveniles were found on palm leaves or in rock crevices [128]. The oviductal eggs of one female (KU 164493) are dark brown. It is likely that females of “Centrolene” medemi deposit their eggs on rocks, as reported in species with similar microhabitats (e.g., Centrolene geckoidea, “Centrolene” petrophilum [113,129]). Parental care is unknown.

Call: Not described.

Tadpole: Not described.

Distribution (Figure 49): “Centrolene” medemi is known from localities on the eastern and western slopes of the Cordillera Oriental of the Colombian Andes and the Amazonian slopes of the Cordillera Oriental of Ecuador at elevation between 790 and 1800 m [101,128,130]. In Ecuador, “Centrolene” medemi has been registered from a single stream nearby Volcán Reventador at 1490 m (Specimens Examined), within the Eastern Montane Forest ecoregion. Although this species was described from a locality in lowlands of Amazonian Colombia (Puerto Asís, upper Río Putumayo at about 280 m [96]), all subsequent records are from Andean localities. Based on this evidence, Ruiz-Carranza et al. [101] rejected the record from Amazonian Colombia.

Figure 49. Distribution of “Centrolene” medemi in Ecuador (yellow dot).

Conservation status: Globally, “Centrolene” medemi is currently listed as Endangered by the IUCN [131]. In Ecuador, the only record is from a stream nearby Volcán Reventador on 19 March 1975 [130], and area that has been visited several times during the last 10 years. We suggest that the species should be locally listed as Critically Endangered.

Evolutionary relationships (Figure 24): Based on morphological traits, “Centrolene” medemi was placed in the Centrolene geckoidea species group by Ruiz-Carranza and Lynch [6]. Guayasamin et al. [1] considered the generic placement of “Centrolene” medemi as incertae sedis within the subfamily Centroleninae. No genetic data are available for this species.

Specimens examined: “Centrolene” medemi: Ecuador: Provincia de Napo: Stream 2 km SSW of junction between Río Reventador and Baeza-Lumbaqui road (0.1 S; 77.6 W, 1490 m), KU 164493–94.

Centrolene peristicta (Lynch and Duellman, 1973 [22]; Figure 50, Figure 51, Figure 52 and Figure 53).
Centrolenella peristicta Lynch and Duellman, 1973 [22]. Holotype: KU 118051.
Type locality: “Tandapi, 1460 m, Provincia Pichincha, Ecuador”.
Centrolene peristictum—Ruiz-Carranza and Lynch, 1991 [6]. Guayasamin, Castroviejo-Fisher, Trueb, Ayarzagüena, Rada, and Vilà, 2009 [1].
Centrolene peristicta—Barrio-Amorós, Rojas-Runjaic, and Señaris, 2019 [85].

Common names: English: Dappled Glassfrog. Spanish: Rana de Cristal Punteada.

Etymology: The specific epithet is derived from the Greek peristiktos, meaning dappled, and refers to the spotted color pattern of the species [22].

Identification: Centrolene peristicta is distinguished from most glassfrogs by its minute body size (SVL < 21.3 mm) and having, in life, a yellowish–green dorsum with minute white flecks and small diffuse black spots (Figure 50). Additionally, adult males of C. peristicta have a small, curved humeral spine that is morphologically different from the spines present in most glassfrogs (i.e., not curved). Similar species include C. antioquiensis, C. lynchi, C. pipilata, and Nymphargus truebae. Centrolene antioquiensis, an endemic species from Colombia, is remarkably similar to C. peristicta (see Taxonomic Remarks), but C. antioquiensis has less hand webbing: hand webbing formulae in C. antioquiensis, III (2⁺–2⁻)—(2–2⁻) IV; in C. peristicta, III (2⁺–2⁻)—(1^1/2–1⁺) IV. Centrolene lynchi is larger than C. peristicta (in males, SVL 23.3–26.5 mm in Centrolene lynchi) and has a very different call. Centrolene pipilata occurs on the Amazonian slopes of the Andes, whereas C. peristicta is restricted to the Pacific slopes of the Andes. Nymphargus truebae, a species only known from the Andes of southern Peru, has the same color pattern as C. peristicta, but N. truebae lacks humeral spines and webbing between Fingers III and IV (present in C. peristicta).

Diagnosis: (1) Vomers lacking teeth; (2) snout round in dorsal aspect, round or truncated in lateral profile (Figure 51); (3) tympanum large, oriented almost vertically, with slight lateral and posterior inclinations, its diameter 44.0%–51.9% of eye diameter; tympanic annulus visible, supratympanic fold evident; tympanic membrane translucent, partially pigmented, differentiated from surrounding skin; (4) dorsal skin shagreen with small warts corresponding to yellowish–white spots, males and females lack spicules; (5) pair of enlarged subcloacal warts (Figure 15); (6) anterior half of the ventral parietal peritoneum white, posterior half transparent (condition P2); white pericardium; iridophores partially or completely covering stomach and colon; no iridophores in peritonea covering kidneys, gall bladder, and urinary bladder (condition V2); (7) liver with four clearly defined lobes, lacking iridophores (condition H0); (8) males with conspicuous humeral spines; (9) webbing absent between Fingers I and II, reduced between Fingers II and III, and moderate to extensive between outer fingers (Figure 51); webbing formula: II (1^3/4–2⁻)—(3⁺–3^1/4) III (2⁺–2⁻)—(1^1/2–1⁺) IV; (10) webbing between toes extensive; webbing formula on foot I 1—(1^1/2–2) II 1—(2⁻–2⁺) III (1⁻–1⁺)—(2⁻–2⁺) IV (2⁻–2⁺)—(1–1^1/3) V; (11) ulnar fold present, with low white tubercles; outer tarsal fold present, with low white tubercles; internal tarsal fold low and short; (12) prepollex usually exposed; nuptial pad Type I or Type III; (13) Finger I slightly shorter or as long as Finger II (Finger I length 90.9%–102.7% Finger II); (14) disc of Finger III width about 45.6%–67.3% of eye diameter; (15) in life, dorsum green with minute yellowish–white spots and larger black spots (Figure 50); green bones; (16) in preservative, dorsum lavender with numerous minute white spots and larger diffuse dark lavender spots; (17) in life, iris white grey with a yellow hue and thin black reticulation; thin yellow line surrounds the pupil; (18) melanophores mostly absent from fingers and toes, except for a few on dorsal surfaces of Finger IV and proximal portion of Finger III; (19) males call from mostly from the lower sides of leaves; each call has one pulsed note with a duration of 0.036–0.087 s and a dominant frequency of 6471–7278 Hz; (20) fighting behavior unknown; (21) egg clutches usually deposited on the underside of leaves; short-term maternal care absent; prolonged parental care provided by males; (22) tadpoles undescribed; (23) minute body size; in adult males, SVL 17.9–21.2 mm (X = 19.9, n = 14); in two adult females SVL 20.8–20.9 mm.

Figure 50. Centrolene peristicta in life. (Top row): Male guarding two egg clutches; Reserva Las Gralarias. (Bottom left): Adult male guarding eggs, from Mindo Biological Reserve, QCAZ 22313; photo by Martín Bustamante. (Bottom right): Individual from Reserva Las Gralarias; photo by Marco Rada.

Diagnosis: (1) Vomers lacking teeth; (2) snout round in dorsal aspect, round or truncated in lateral profile (Figure 51); (3) tympanum large, oriented almost vertically, with slight lateral and posterior inclinations, its diameter 44.0%–51.9% of eye diameter; tympanic annulus visible, supratympanic fold evident; tympanic membrane translucent, partially pigmented, differentiated from surrounding skin; (4) dorsal skin shagreen with small warts corresponding to yellowish–white spots, males and females lack spicules; (5) pair of enlarged subcloacal warts (Figure 15); (6) anterior half of the ventral parietal peritoneum white, posterior half transparent (condition P2); white pericardium; iridophores partially or completely covering stomach and colon; no iridophores in peritonea covering kidneys, gall bladder, and urinary bladder (condition V2); (7) liver with four clearly defined lobes, lacking iridophores (condition H0); (8) males with conspicuous humeral spines; (9) webbing absent between Fingers I and II, reduced between Fingers II and III, and moderate to extensive between outer fingers (Figure 51); webbing formula: II (1^3/4–2⁻)—(3⁺–3^1/4) III (2⁺–2⁻)—(1^1/2–1⁺) IV; (10) webbing between toes extensive; webbing formula on foot I 1—(1^1/2–2) II 1—(2⁻–2⁺) III (1⁻–1⁺)—(2⁻–2⁺) IV (2⁻–2⁺)—(1–1^1/3) V; (11) ulnar fold present, with low white tubercles; outer tarsal fold present, with low white tubercles; internal tarsal fold low and short; (12) prepollex usually exposed; nuptial pad Type I or Type III; (13) Finger I slightly shorter or as long as Finger II (Finger I length 90.%–102.7% Finger II); (14) disc of Finger III width about 45.6%–67.3% of eye diameter; (15) in life, dorsum green with minute yellowish–white spots and larger black spots (Figure 50); green bones; (16) in preservative, dorsum lavender with numerous minute white spots and larger diffuse dark lavender spots; (17) in life, iris white grey with a yellow hue and thin black reticulation; thin yellow line surrounds the pupil; (18) melanophores mostly absent from fingers and toes, except for a few on dorsal surfaces of Finger IV and proximal portion of Finger III; (19) males call from mostly from the lower sides of leaves; each call has one pulsed note with a duration of 0.036–0.087 s and a dominant frequency of 6471–7278 Hz; (20) fighting behavior unknown; (21) egg clutches usually deposited on the underside of leaves; short-term maternal care absent; prolonged parental care provided by males; (22) tadpoles undescribed; (23) minute body size; in adult males, SVL 17.9–21.2 mm (X = 19.9, n = 14); in two adult females SVL 20.8–20.9 mm.

Figure 51. Centrolene peristicta, KU 178148. (A) Head in lateral view. (B) Hand in ventral view. (C) Dorsal view of Finger I, showing nuptial pad. Illustrations by Juan M. Guayasamin.

Color in life (Figure 50): Dorsum green with minute yellowish–white flecks and small dark grey spots; upper lip white; region below eye with small white warts; bones green; vocal sac green; upper flanks green with minute white spots; lower flanks cream with minute white spots; venter yellowish cream; fingers and toes dull yellow green; ulnar fold with thin white line or with low white ulnar tubercles; outer tarsal fold with low white tubercles; small white tubercles just posterior to cloaca.

Color in ethanol: Dorsal surfaces of head, body, and limbs lavender with minute white spots and small black spots; margin of upper lip white; region below eye with small white warts; white tubercles just posterior to cloaca. White parietal peritoneum covers anterior half of venter; white pericardium; clear peritoneum on liver and kidneys; digestive tract partially or completely covered with white lining.

Variation: One individual (QCAZ 16313) has considerably less webbing on the hands (III 2^1/2—2⁺ IV). Individuals from streams nearby the town of Mindo lack iridophores on the gastrointestinal peritonea.

Biology and ecology: The information shown below is from Salgado and Guayasamin [132]. At Reserva Las Gralarias, during the night, Centrolene peristicta is active on vegetation 40–600 cm above permanent streams in primary evergreen lower-montane forests and cloudforests. Its breeding season is in the rainy (December–April), but peaks in February–April. Males vocalize to advertise themselves and defend territories in which females place clutches containing 6–41 eggs. Most of the time, egg clutches are placed on the underside of leaves and ferns 70–500 cm directly above streams, or under dead leaves nearby streams. Males are polygynous and exhibit high site fidelity; some males were observed simultaneously guarding egg clutches at different stages of development. Parental care by males was demonstrated experimentally; unattended clutches have a significantly lower eclosion rate than attended clutches, and prolonged hatching time. Clutch mortality was mainly because of desiccation, predation, and parasitism. Embryos develop for 17–27 days, then hatching as free-swimming larvae. At Reserva Las Gralarias, the species is abundant at Lucy’s Creek, but it is also found at Kathy’s Creek and Santa Rosa River [88].

Call (Figure 52): The information shown below is from Salgado and Guayasamin [132]. Advertisement call of males of Centrolene peristicta consists of one short and pulsed note that resembles the sound of a cricket chirp. The note has a duration of 0.036–0.087 s (X + SE = 0.062 + 1.07, N = 35) and each call has one note. The dominant frequency is at 6471–7278 Hz (X + SE = 6878 + 177 Hz., N = 35). The fundamental frequency is the same as the dominant frequency; there is one (range = 3235–6198 Hz) or two harmonics (range = 6471–7278 Hz). There is no conspicuous frequency modulation during the call. The repetition rate of the advertisement call varies from two to 20 calls per minute.

Tadpole: Not described.

Distribution (Figure 53): Centrolene peristicta is known from the Pacific slope of the Cordillera Occidental of the Andes in Ecuador and Colombia at elevations between 1380 and 1900 m ([22,87,133,134], this work). In Ecuador, C. peristicta is known from localities in the provinces of Carchi, Pichincha, and Santo Domingo de los Tsáchilas, at elevations of 1400–1852 m, with a potential distribution of 12,603 km². All localities are within the Western Montane Forest ecoregion.

Conservation status: Globally, Centrolene peristicta is currently listed as Least Concern by the IUCN [135]. In Ecuador, Arteaga et al. [87] suggested the category of Near Threatened. Several recent observations show that the species has a wider distribution than previously thought. Reproductive populations have been observed at Reserva Las Gralarias (December 2009–May 2018; JMG, pers. obs.), Mindo Biology Station (February 2002; December 2014; M. R. Bustamante and L. Bustamante, pers. com.), and at Río Pachijal on August 2001 (I. Tapia, pers. com.). The potential distribution of the species covers an area of 12,603 km², 44.6% of which is affected by human activities. At Reserva Las Gralarias, the species is infected by the amphibian chytrid fungus Batrachochytrium dendrobatidis [92], but populations look healthy and there is no evidence of declines. The conservation category of Least Concern seems too optimistic, given the fragmented distribution of the species. We suggest that the species should be considered as Near Threatened, in agreement with Arteaga et al. (2013).

Figure 52. Call of Centrolene peristicta (LBE-C-047), recorded at Reserva Las Gralarias, 1800 m, Pichincha province, Ecuador. (A) Series of calls. (B) Single, pulsed call.

Figure 53. Distribution of Centrolene peristicta in Ecuador (yellow dots).

Evolutionary relationships (Figure 24): Molecular evidence places Centrolene peristicta and C. antioquiensis as sister species.

Taxonomic Remarks: Centrolene antioquiensis and C. peristicta are almost identical in morphology and could represent a single species. The subtle differences found between these two species might be a consequence of incomplete sampling or intraspecific geographic variation. The analysis of calls from C. antioquiensis is critical to assess the validity of the species status of C. peristicta.

Specimens examined: Centrolene peristicta: Ecuador: Provincia de Carchi: Maldonado (0.9 N, 78.1 W, 1410 m), KU 178137–44, 178145–51, 180325; Provincia de Pichincha: Bosque Protector Río Guajalito (0.233 S, 78.817 W; 1900 m), QCAZ 6446; stream near Mindo Biology Station (0.07805 S, 78.7319 W; 1600 m), QCAZ 22757–59, 22312–14; Tandapi (0.416389 S, 78.7989 W, 1520 m), KU 118051–52, 121053; 1.6 km W of Tandapi (0.40472 S, 78.8058 W; 1400 m), KU 178152; 5 km W of Tandapi on the Tandapi–Atenas road (0.3954 S, 78.8326 W; 1670 m), QCAZ 15901, 15922; Río Pachijal (0.2 S, 78.75 W; 1740 m), QCAZ 16316; Reserva Las Gralarias (0.00806 S, 78.72443 W; 1852 m), QCAZ 47298. Provincia de Santo Domingo de los Tsáchilas: Río Faisanes, ca. 15 km NE of La Palma, on the Quito–Chiriboga–Santo Domingo road (0.3167 S, 78.817 W; 1380 m), USNM 286714. Colombia: Departamento de Nariño, Reserva La Planada, ICN 12114 (PR 7871).

Centrolene pipilata (Lynch and Duellman 1973 [22]; Figure 54, Figure 55 and Figure 56).
Centrolenella pipilata Lynch and Duellman, 1973 [22]. Holotype: KU 143278.
Type locality: “16.5 km NNE of Santa Rosa, 1700 m, on Quito-Lago Agrio road, Provincia de Napo, Ecuador”.
Centrolene pipilatum—Ruiz-Carranza and Lynch, 1991 [6].
Centrolene pipilata—Barrio-Amorós, Rojas-Runjaic, and Señaris, 2019 [85].

Common names: English: Peeping Glassfrog. Spanish: Rana de Cristal Piadora.

Etymology: The specific epithet pipilata is an adjectival derivative of the Latin verb pipila, meaning to peep, and refers to the call of this centrolenid frog [22].

Identification: Centrolene pipilata is distinguished from most glassfrogs by having, in life, a green dorsum with yellowish–white flecks and diffuse dark green marks, and a distinct prepollex (Figure 54 and Figure 55). On the Amazonian slopes of Ecuador, the only species that have a similar dorsal color pattern are C. sanchezi and C. huilensis. Centrolene sanchezi has a short, thin humeral spine (large, wide, and usually with the tip projected anteriorly in C. pipilata) and lacks a distinct prepollex. Centrolene huilensis is conspicuously larger than C. pipilata (in C. huilensis, male SVL = 23.6–26.7 mm; SVL = 28.7 mm in 1 female; in C. pipilata, male SVL 19.7–22.6 mm, female SVL 22.6–23.6 mm). Nymphargus truebae, a species only known from the Andes of southern Peru, has the same color pattern as C. pipilata, however, males in N. truebae lack humeral spines (present in males of C. pipilata). The Colombian Centrolene hybrida has a white gastrointestinal peritoneum (iridophores absent in C. pipilata), and lacks a distinct prepollex.

Figure 54. Centrolene pipilata. (Top row): Coloration in life, adult male, holotype, KU 143278; photos by William E. Duellman. (Bottom row): Coloration in preservative of KU 178155 (left) and KU 143287 (right); photos by Juan M. Guayasamin.

Diagnosis: (1) Vomers lacking teeth; (2) snout round in dorsal aspect, round or truncated in lateral profile (Figure 55); (3) tympanum moderate, oriented almost vertically, with slight lateral and posterior inclinations, its diameter 31.0%–39.4% of eye diameter; tympanic annulus completely visible; supratympanic fold evident; tympanic membrane translucent, partially pigmented, clearly differentiated from surrounding skin; (4) dorsal skin shagreen, males with spicules coinciding with white spots; (5) pair of enlarged subcloacal warts (Figure 15); (6) anterior 50%–60% of the ventral parietal peritoneum white, posterior portion transparent (condition P2–P3); white pericardium; no iridophores in peritonea covering intestines, stomach, kidneys, gall bladder, and urinary bladder (condition V1); (7) liver with four clearly defined lobes, lacking iridophores (condition H0); (8) males with conspicuous humeral spines; (9) webbing absent between Fingers I, II, and III; moderate between outer fingers (Figure 55); webbing formula: III (2⁺–2^1/2)—(2–2⁺) IV; (10) webbing between toes extensive; webbing formula on foot: I (1⁺–2⁻)—(2–2⁺) II (1–1^1/3)—(2–2^1/3) III (1–1^1/2)—2⁺ IV (2–2^1/2)—(1–1^1/2) V; (11) white ulnar fold present; outer tarsal margin with low white tubercles: Internal tarsal fold low; (12) exposed prepollex; nuptial pad Type II; (13) Finger I as long as Finger II or slightly shorter (Finger I 90.0%–100% of Finger II); (14) disc of Finger III width 44.8%–56.5% of eye diameter; (15) in life, dorsum green with small diffuse black spots and yellowish–white flecks (Figure 54); green bones; (16) in preservative, dorsum lavender with numerous minute white spots and larger dark lavender spots (Figure 54); (17) in life, iris greyish–white with thin black reticulations and a yellow hue around pupil; (18) melanophores mostly absent from fingers and toes, except for a few on proximal half of Finger IV, and along Toes IV and V; (19) males call from upper side of leaves; call undescribed; (20) fighting behavior unknown; (21) eggs placed on upper surface of leaves; parental care unknown; (22) tadpoles unknown; (23) minute body size; in adult males, SVL 19.7–22.6 mm (X = 21.5, n = 9); in two adult females SVL 22.6–23.6 mm.

Color in life (Figure 54): Dorsum green with minute yellowish–white flecks and larger, diffuse black spots; upper lip white; region below eye with small white warts that are spiculated in males; bones green; upper flanks green with minute white spots; lower flanks whitish cream; ulnar fold with thin white line; outer tarsal fold with low white tubercles; small white tubercles just posterior to cloaca; iris greyish–white with thin black reticulations and a yellow hue around pupil.

Color in ethanol: Dorsal surfaces of head, body, and limbs lavender with small white spots and larger dark lavender spots (Figure 54); margin of upper lip white; region below eye with small, spiculated white warts; white tubercles just posterior to cloaca. White parietal peritoneum covers anterior 50%–60% of venter; white pericardium; no iridophores in peritonea covering digestive tract, liver, kidneys, gall bladder, and urinary bladder.

Variation: In preservative, some individuals lack dorsal dark spots.

Figure 55. Centrolene pipilata, KU 143278, holotype. (A) Head in lateral view. (B) Hand in ventral view. Illustrations by Juan M. Guayasamin.

Biology and ecology: All individuals have been found at night on vegetation along cascading mountain streams. In captivity, a female deposited a clutch of 18 eggs. Males call from the upper side of leaves. At Río Azuela, Centrolene pipilata was found in sympatry with Nymphargus anomalus, N. megacheirus, N. siren, and Hyalinobatrachium pellucidum. At 16.5 NNE of Santa Rosa, the species was found with Espadarana audax, N. megacheirus, and N. siren [22]. Eggs have clear jelly and pale green yolks [22]. Parental care is unknown.

Call: Not described.

Tadpole: Not described.

Distribution (Figure 56): Centrolene pipilata is endemic to the cloud forest on the Amazonian slope of the Ecuadorian Andes at elevations between 1300–1910 m ([17,22], this work). The species has been recorded from four localities in the provinces of Napo and Sucumbíos. The habitat of the species is within the Eastern Montane Forest region.

Figure 56. Distribution of Centrolene pipilata in Ecuador (yellow dots).

Conservation status: Globally, Centrolene pipilata is listed by the IUCN as Endangered [136] because of its limited distribution and the continuing decline in the extent and quality of suitable habitat. The last confirmed report of C. pipilata was at 14.7 km NE of Río Salado on February 1979 [17]. Recent surveys at Río Azuela [91] and the type locality (JMG, pers. obs.) have failed to find the species. In Ecuador, based on the IUCN criteria A2c,e, we suggest that the species should be considered as Critically Endangered.

Evolutionary relationships (Figure 24): Given the current taxon and gene sampling, Centrolene pipilata and C. hybrida are sister species.

Specimens examined: Centrolene pipilata: Ecuador: Provincia de Napo: 16.5 km NNE Santa Rosa (0.2186 S, 77.732 W, 1700 m), KU 143278–83, 143554; 3.2 km NNE Oritoyacu (0.4597 S, 77.8672 W; 1910 m), KU 178153; Río Azuela (0.11667 S, 77.61667 W; 1740 m), KU 143284–87, 155498, 166331; Río Salado, 1 km upstream from Río Coca (0.19167 S, 77.6997 W; 1420 m), KU 178154–55.

Centrolene sanchezi Ruiz-Carranza and Lynch, 1991 [129] (Figure 57, Figure 58, Figure 59, Figure 60 and Figure 61).
Centrolene sanchezi Ruiz-Carranza and Lynch, 1991 [129]. Holotype: ICN 24293.
Type locality: “Departamento de Caquetá, municipio de Florencia, vereda Gabinete, 3.1 Km por carretera abajo del Alto Gabinete, vertiente oriental, Cordillera Oriental, 1°4′ latitud N, 75°4′ W de Greenwich, 2190 m”, Colombia.
Centrolene bacatum Wild, 1994 [137]. Holotype: KU 202803. Type locality: “11.2 km WSW Plan de Milagro (03°02′ S, 78°35′ W, 2350 m), Provincia Morona Santiago, Ecuador”. New synonymy.
Centrolene guanacarum Ruiz-Carranza and Lynch, 1995 [26]. Holotype: ICN 11686. Type locality: “Departamento del Cauca, municipio de Inzá, Km. 84 carretera Popayán a Inzá, Río Guanacas, quebrada afluente, Internado Indígena Río Guanacas, vertiente oriental Cordillera Central, 2°34′ Latitud N, 76°05′ W de Greenwich, 1800–1900 m”, Colombia. New synonymy.
“Centrolene” guanacarum—Guayasamin, Castroviejo-Fisher, Trueb, Ayarzagüena, Rada, and Vilà, 2009 [1]. New synonymy.
Centrolene bacata—Barrio-Amorós, Rojas-Runjaic, and Señaris, 2019 [85].

Common names: English: Sánchez’s Glassfrog. Spanish: Rana de cristal de Sánchez.

Etymology: The specific epithet sanchezi honors Ricardo Sánchez, who, with John D. Lynch, discovered the species.

Identification: Centrolene sanchezi is easily distinguished from other glassfrogs by its minute body size (SVL 18.9–22.3 mm), green dorsum with small white spots, and the presence of white warts in an area that extends from below the eye to the insertion of the arm (Figure 57 and Figure 58A). Additionally, adult males have conspicuous humeral spines. The only other centrolenid from eastern Ecuador that can be confused with Centrolene sanchezi is Centrolene pipilata, which has a green dorsum with small, diffuse black spots and yellowish–white flecks (Figure 54), and a distinct prepollex. Also, the two species occupy different elevations on the Amazonian slope of the Andes; C. sanchezi is found at elevations of 1950–2350 m, whereas C. pipilata occurs at 1420–1910 m.

Diagnosis: (1) Vomerine teeth absent; (2) snout rounded in dorsal aspect, bluntly rounded or truncated in lateral profile (Figure 58); (3) tympanum oriented almost vertically, its diameter 31.4%–37.8% of eye diameter; tympanic annulus visible except for dorsal border covered by supratympanic fold; tympanic membrane partially pigmented, but differentiated from surrounding skin; (4) dorsal surfaces shagreen, with small spicules evident in most males; (5) pair of enlarged subcloacal warts (Figure 15); (6) anterior half of ventral parietal peritoneum white, posterior half translucent (condition P2); white pericardium, translucent visceral peritoneum (condition V1); (7) liver tetralobed, lacking iridophores (condition H0); (8) humeral spines present in adult males; (9) no webbing between Fingers I and II; webbing formula on hand: II 2—3^1/3 III 2^1/2—2^1/4 IV (Figure 58); (10) webbing formula on foot: I 1^1/2—2⁺ II (1–1^1/2)—(2–2^1/3) III 1⁺—(2⁺–2^1/4) IV 2^1/2—(1–1⁺) V (Figure 58); (11) ulnar and inner tarsal folds low or absent; outer tarsal fold absent; (12) nuptial pad Type I, concealed prepollex; (13) Fingers I and II about equal in length (FII/FI = 91.3%–104.5%); (14) disc of Finger III of moderate size, about 40.3%–50.6% of eye diameter; (15) in life, dorsum dark green with small white spots (Figure 57); conspicuous white warts below the eye and tympanum; upper lip white; bones green; (16) in preservative, dorsum lavender with small white spots; (17) iris pale bronze with black reticulation; (18) melanophores mostly absent from fingers and toes, except for a few on Toes IV and V and on base of outer fingers; (19) males call from the upper side of leaves; calls are produced in series; each call has one or two notes, with a duration of 6–21 ms (mean = 11, SD = 2.8); dominant frequency at peak amplitude is 5719–6188 Hz (mean = 5996, SD = 131); (20) fighting behavior unknown; (21) eggs deposited on upper side of leaves; short-term maternal care unknown; parental care by males absent; (22) tadpoles unknown; (23) minute body size, SVL 18.9–22.3 mm in males (n = 15); 20.9 mm in one female.

Figure 57. Centrolene sanchezi in life. (A,B) Male and female from Yanayacu Biological Station, Ecuador. (C) Holotype of C. sanchezi, ICN 24293. (D) Paratype of C. guanacarum, ICN 11685. Photos by A. Arteaga (A,B) and P. Ruiz-Carranza (C,D).

Figure 58. Centrolene sanchezi, KU 202803 (holotype of Centrolene bacata). (A) Head in lateral view. (B) Head in dorsal view. (C) Hand in ventral view. (D) Foot in ventral view. Drawings not to scale. Modified from Wild [137].

Color in life (Figure 57): Based on field notes by W. E. Duellman (4 March 1984), reported in Wild [137], and observation by authors. Dorsum dark green; series of white–cream tubercles under eye; throat and ventral surfaces of limbs green; digits pale green; cloacal region with white warts; ventral parietal peritoneum white anteriorly and translucent posteriorly; visceral peritoneum translucent; bones green; iris pale bronze with thin black reticulation.

Color in ethanol (Figure 59): Dorsal surfaces lavender with small, unpigmented spots and white warts; limbs cream lavender with numerous small, unpigmented spots and few white warts; white warts on lateral surface of head; upper lip white; tympanum pigmented with purple specks; cloacal region with white or cream warts; iris silvery white with dark purple reticulation. Fingers I and II and Toes I–III dorsally unpigmented; some pigmentation visible on Fingers III and IV, and Toes IV and V. White parietal peritoneum covering anterior half of venter, posterior half translucent; silvery-white pericardium; translucent peritonea covering liver, gastrointestinal track, and renal capsules (dissected male: QCAZ 22386).

Figure 59. Types of Centrolene sanchezi and C. bacata in preservative. (A,B) Centrolene bacata, holotype KU 202803. (C,D) C. sanchezi, paratype ICN 24294. Photos by Juan M. Guayasamin.

Biology and ecology: During the night, Centrolene sanchezi has been found on leaves approximately 130–300 cm above streams in primary and secondary forest. Males call from the upper sides of leaves. A male (QCAZ 22728) was found nearby two egg clutches, one on the underside and the other on the upper side of a single leaf; egg clutches have 14–18 pale yellowish–green eggs. During February 2013, males have been heard calling at Yanayacu Biological Station. At this locality, Centrolene sanchezi (reported as C. bacata) is the most abundant centrolenid; other sympatric species at Yanayacu include Centrolene aff. buckleyi, C. huilensis, Nymphargus posadae, N. siren, and N. wileyi [20,125]. Short-term maternal care unknown; parental care by males absent ([25], as C. bacata).

Call (Figure 60): We analyzed 63 notes contained within 11 calls from 1 individual (LBE-C-023). Calls are produced in series, which can be relatively long (range = 618–3085 ms, mean = 1462.6 ms, SD = 639.5 ms); each can have one or two notes; note repetition rate is 3–9 (mean = 5.7, SD = 1.6) notes per call. Notes sometimes occur in pairs (i.e., much shorter note interval than compared to the rest of the call). Each call is very short, with a duration of 6–21 (mean = 11, SD = 2.8) ms. Notes are strongly pulsed and have one or two (mean = 1.4, SD = 0.5) amplitude peaks throughout the note, where the second amplitude peak is generally weaker than the first. Pulses within a note have a rate of 83–200 (mean = 122, SD = 29) pulses per second. Notes have their peak amplitude in the first 50% of the note (relative peak time: Range = 0.0987–0.5497, mean = 0.216, SD = 0.076). The dominant frequency measured at peak amplitude is 5719–6188 (mean = 5996, SD = 131) Hz and is contained within the fundamental frequency. The fundamental frequency has a lower limit of 4688–6000 (mean = 5714, SD = 222) Hz and a higher limit of 5906–6656 (mean = 6268, SD = 144) Hz.

Figure 60. Call of Centrolene sanchezi (LBE-C-023), recorded at Reserva Yanayacu, 2150 m, Napo province, Ecuador. (A) Series of calls. (B) Single, pulsed call.

Tadpole: Not described.

Distribution (Figure 61): Centrolene sanchezi is known from a few localities on the Amazonian slopes of the Andes of Ecuador and Colombia and from one locality on the eastern slope of the Colombian Cordillera Central (see Specimens Examined), at elevations between 1800 and 2350 m [20,101,129,137]. In Ecuador, the habitat of the species is within the Eastern Montane Forest region.

Figure 61. Distribution of Centrolene sanchezi in Ecuador (yellow dots).

Conservation status: Globally, Centrolene sanchezi is listed as Data Deficient by the IUCN [138], but the evaluation does not account for the synonym presented herein. The habitat of the species is fragmented by agriculture and pastureland and threatened by mining (mainly in southern Ecuador). We suggest that the species should be considered as Endangered, following IUCN criteria B2, Ba, Bb(iii).

Evolutionary relationships (Figure 24): Centrolene sanchezi is the sister species to a clade formed by C. pipilata plus C. hybrida. These three species are found on the Amazonian slopes of the Andes.

Taxonomic Remarks: Examination of the type material of Centrolene sanchezi, C. bacata, and C. guanacarum reveals no morphological differences among them. Moreover, all specimens share two distinctive traits; a small, laminar humeral spine in males and the presence of a lateral row of enameled warts that extends from below the eye to just posterior to the insertion of the arm (Figure 51). Therefore, we place Centrolene bacatum Wild 1994 [137] and Centrolene guanacarum Ruiz-Carranza and Lynch 1995 [26] under the synonymy of Centrolene sanchezi Ruiz-Carranza and Lynch, 1991 [129].

Specimens examined: Centrolene sanchezi: Ecuador: Provincia de Morona Santiago: 11.2 km WSW Plan de Milagro (03°07′ S, 78°30′ W; 2350 m), KU 202803 (holotype of Centrolene bacata), 202804, 202807–12 (paratypes of C. bacata); Provincia de Napo: Yanayacu Biological Station (0°41′ S, 77°53′ W; 2100 m), QCAZ 16212, 17807, 22386–87, 22728, 26025–27, 26056, 27438. Colombia: Departamento de Caquetá: Municipio de Florencia, Vereda Gabinete, 3.1 Km por carretera abajo del Alto Gabinete, vertiente oriental, Cordillera Oriental, 1°4′ latitud N, 75°4′ W de Greenwich, 2190 m, ICN 24293 (holotype of C. sanchezi); Departamento del Cauca: Municipio de Inzá, Km. 84 carretera Popayán a Inzá, Río Guanacas, quebrada afluente, Internado Indígena Río Guanacas, vertiente oriental Cordillera Central, 2°34′ Latitud N, 76°05′ W de Greenwich, 1800–1900 m, ICN 11685 (paratype of C. guanacarum).

Genus Chimerella Guayasamin, Castroviejo-Fisher, Trueb, Ayarzagüena, Rada, & Vilà, 2009 [1].
Etymology: The name Chimerella comes from the Greek Khímaira and the suffix–ella diminutive. In Greek mythology, the Chimera is a creature composed of parts of multiple animals; the name refers to the peculiar combination of morphological characteristics of Chimerella mariaelenae [1].
Chimerella mariaelenae (Cisneros-Heredia and McDiarmid, 2006 [139]; Figure 62, Figure 63, Figure 64 and Figure 65).
Centrolene mariaelenae Cisneros-Heredia and McDiarmid, 2006 [139]. Holotype: DFCH-USFQ D125.
Type locality: “small stream, tributary of the Jambue River, ca. 16 km S from Zamora, Podocarpus National Park (ca. 04°15′ S, 78°56′ W, 1820 m), on the western slope of Contrafuerte de Tzunantza, Cordillera Oriental, eastern slopes of the Andes, Provincia de Zamora Chinchipe, Republic of Ecuador”.
Cochranella parabambae (in part)—Goin, 1961 [97].
Chimerella mariaelenae—Guayasamin, Castroviejo-Fisher, Trueb, Ayarzagüena, Rada, and Vilà, 2009 [1].

Common names: English: María Elena’s Glassfrog. Spanish: Rana de Cristal de María Elena.

Etymology: The specific epithet mariaelenae is a noun in the genitive case and a patronym for María Elena Heredia, Diego F. Cisneros-Heredia’s mother [139].

Identification: Chimerella mariaelenae (Figure 62 and Figure 63), Chimerella corleone, and Vitreorana gorzulae are the only known glassfrog that have the following combination of traits: Humeral spine in adult males, transparent ventral parietal peritoneum, and white pericardial, hepatic, and visceral peritonea. Vitreorana gorzulae, an endemic to the Guiana Shield, is distinguished by lacking dark spots on the dorsum (present in C. mariaelenae). Chimerella mariaelenae is easily differentiated from the Peruvian C. corleone by having an orange to reddish iris (silvery white in C. corleone) and a dorsum with small black dots (dorsum with yellow dots in C. corleone [19]). On the Amazonian slopes of Ecuador, only species in the genus Hyalinobatrachium and Teratohyla amelie could be confused with C. mariaelenae. However, these species lack the small dark spots that characterize the dorsum of C. mariaelenae. Additionally, adult males of C. mariaelenae have small humeral spines, which are absent in all Hyalinobatrachium species and Teratohyla amelie.

Diagnosis: (1) Vomers lacking teeth; (2) snout truncated in dorsal aspect, truncated to slightly protruding in lateral profile (Figure 63); (3) tympanum oriented posterolaterally with slight dorsal inclination, its diameter 23.0%–27.4% of eye diameter; tympanic membrane translucent, clearly differentiated from surrounding skin; (4) dorsal skin shagreen; males with minute dorsal spicules (only visible under magnification ×250); (5) pair of enlarged subcloacal warts; (6) ventral parietal peritoneum transparent (condition P0); iridophores in pericardium and peritonea covering digestive tract and testes; kidneys and urinary bladder lacking iridophores (condition V5); (7) liver bulbous and covered by iridophores (condition H2); (8) males with small humeral spines; (9) webbing absent or basal between inner fingers, moderate between outer fingers (Figure 63); webbing formula IV (2⁺–2^1/2)—(2⁺–2^1/2) V; (10) webbing between toes extensive; webbing formula on foot I (1^1/2–2⁻)—(2–2^1/2) II 1—2⁺ III (1⁺–1^1/2)—(3⁻–3⁺) IV (2^1/2–3^1/4)—(1⁺–1^1/4) V; (11) ulnar and tarsal folds absent; (12) concealed prepollex; nuptial pad Type I; (13) Finger I slightly longer than Finger II (Finger II length 95.6%–97.3% of Finger I); (14) disc of Finger III relatively narrow, its width about 22.3%–25.7% of eye diameter; (15) in life, dorsum yellowish green with small grey to black spots (Figure 62); bones light green; (16) in preservative, dorsum pale lavender with small, dark, lavender spots; (17) in life, iris white with abundant dark flecks, pupil surrounded by a rufous to orange ring, outlined by a dark grey to blue ring (Figure 62); (18) dorsal surfaces of fingers and toes lacking melanophores, except for some on base of Toe V; (19) males call from the upper surface of leaves; typical call with two notes; each note extremely short at 4–7 (mean = 6, SD = 0.9) ms; dominant frequency measured at peak amplitude is 6718–8010 (mean = 7510, SD = 408) Hz; (20) fighting behavior unknown; (21) eggs laid on the upper surfaces of leaves; short-term maternal care present; parental care by males absent; (22) tadpoles with elongated body, head with dorsoventral compression; labial tooth row formula 2(1)(2)/3; (23) minute body size; adult males, SVL 17.9–19.7 mm (n = 3); one adult female SVL 20.8 mm.

Color in life (Figure 62): Dorsal surfaces yellowish green, with several grey to black spots. No white coloration is evident on upper lip or ulnar and tarsal regions. Iris bicolored, with dark brown circumpupilar area separated from pupil by orange to red–brown ring; whitish external background with abundant dark flecks on uppermost and lowermost portions of iris. Venter completely transparent (clear ventral parietal peritoneum), showing white heart (covered by white pericardium); white digestive tract, liver, testes, and gall bladder. Transparent peritoneum covering urinary bladder. Bones white to light green.

Color in ethanol: Dorsal surfaces of head, body, and limbs pale lavender, with small dark lavender spots. Ventral parietal peritoneum and kidneys lack iridophores. White pericardial, hepatic, and visceral peritonea.

Figure 62. Chimerella mariaelenae in life. (Top row): Adult male from Bigal River Biological Station, 931 m, Ecuador; photos by Ross Maynard. (Bottom row): Amplectant pair, Quebrada Pangayaku, 930 m, Napo province, Ecuador, MZUTI 1680–81; photo by Eduardo Toral.

Figure 63. Chimerella mariaelenae. (A) Head in lateral view, QCAZ 31729. (B) Head in dorsal view, QCAZ 21252. (C) Hand in ventral view, QCAZ 22363. Illustrations by Juan M. Guayasamin.

Biology and ecology: The biology of Chimerella mariaelenae is poorly known. All individuals have been found on the upper surfaces of leaves along small streams and ditches in cloud forest. At the type locality, C. mariaelenae is sympatric with Nymphargus cochranae and Hyloscirtus phyllognathus [139]. Reproductively active individuals have been observed nearby Cascada San Rafael in Quebrada Pangayacu in August 2012. Short-term maternal care present; parental care by males absent [25].

Tadpole: The following information is summarized from Terán-Valdez and Guayasamin [140] and is based on a specimen at Gosner Stage 39 raised in laboratory conditions. Body elongated and oval-depressed, wider (body width = 5.1 mm) than higher (body height = 3.8 mm). Chondrictial elements not visible. Snout rounded in dorsal and lateral views. Lateral line system visible, formed by several stitches parallel or perpendicular to longitudinal axis of body. Short, single, sinistral spiracle located at posterolateral region of body; spiracular aperture with dorsoposterior orientation, with inner wall present as a low ridge. Vent tube short and abdominal, free posteriorly, opening directed posteriorly. Tail long, with subacute tip. Myotomes visible throughout length of tail; straight medial line visible, separating dorsal and ventral myotomes. Dorsal fin originating at about mid-length of tail; height relatively uniform until distal end, where it decreases abruptly. Proximally, ventral fin originating at base of tail muscle, reaching its maximum height after mid-length of tail. Oral disc non-emarginated and surrounded by 49 marginal uniserial papillae. Only ventral and lateral papillae present, lacking dorsal papillae. Upper and lower jaw sheaths nearly straight and fully keratinized, with serrated edge. Labial tooth row formula 2(1)(2)/3; gap in tooth row A-1 could be artificial (because of teeth loss). In life, dorsally, tadpoles at Stage 39 brownish with two areas without pigmentation at anterolateral border of eye. Reddish coloration visible on anterior half of dorsum because of its transparency. Anterior-most part of head is grey. Iridophore aggregations present on dorsum, along vertebral column. Iris bronze. Ontogenetic variation is provided in Terán-Valdez and Guayasamin [140].

Call (Figure 64): We analyzed 26 notes contained within six calls from one individual (LBE-C-021). The typical advertisement call is variable from being short to a moderate length call (range = 231–1761 ms, mean = 679 ms, SD = 623.3 ms). The typical call has two notes, but some call can have up to 10 (mean = 4.3, SD = 3.4) notes. Each note is extremely short at 4–7 (mean = 6, SD = 0.9) ms. Notes have a single amplitude peak. The dominant frequency measured at peak amplitude is 6718–8010 (mean = 7510, SD = 408) Hz and is contained within the fundamental frequency. The fundamental frequency has a lower limit of 6460–7752 (mean = 7222, SD = 387) Hz and a higher limit of 7063–8441 (mean = 7828, SD = 398) Hz.

Figure 64. Call of Chimerella mariaelenae (LBE-C-021), recorded at Pangayaku creek, 929 m, Napo province, Ecuador. (A) Series of calls. (B) Single, pulsed note.

Distribution (Figure 65): Chimerella mariaelenae occurs on the Amazonian slopes of the Ecuadorian Andes at elevations between 813 and 1820 m ([139,141,142], this work), and was recently reported from Peru [143]. The species is known from a few localities in the provinces of Napo, Morona Santiago, Orellana, Tungurahua, and Zamora Chinchipe (Specimens Examined). The potential distribution of the species is 25,472 km² within the Eastern Foothill and Montane Forest regions.

Figure 65. Distribution of Chimerella mariaelenae in Ecuador (yellow dots).

Conservation status: Globally, Chimerella mariaelenae is currently listed as Least Concern by the IUCN [144]. The species is threatened by human activities, although it tolerates some habitat disturbance and has a relatively large distribution, including populations within protected areas (i.e., Bigal River Biological Station, Parque Nacional Podocarpus, Parque Nacional Sangay, Reserva Ecológica Cayambe-Coca, Reserva Narupa). In Ecuador, major threats for the species include deforestation and mining; thus, we suggest that the species should be considered as Near Threatened at the local level.

Evolutionary relationships (Figure 19 and Figure 66): In the original description and based on morphological data, Chimerella mariaelenae was assigned to the Centrolene gorzulai group, which, otherwise, consisted of species from the Guiana Shield. Under this hypothesis, the distribution of C. mariaelenae was thought to support a biogeographical connection between the Andes and the Guiana Shield [139]. Subsequently, Cisneros-Heredia and McDiarmid [17], based on differences in the chromatophore organization, considered that the relationships of C. mariaelenae were uncertain. Phylogenetic trees based on molecular data show that Chimerella mariaelenae and C. corleone are sister species [19] (Figure 66); both species are restricted to the Amazonian versant of the Andes, with no close relationship with species from the Guiana Shield. The genus Chimerella is herein inferred as sister to Espadarana (Figure 19), but with low nodal support.

Remarks: One of the specimens reported by Goin [97] as Centrolenella parabambae from the Topo River is an adult female Chimerella mariaelenae.

Specimens examined: Chimerella mariaelenae: Ecuador: Provincia de Napo: Río Hollín (ca. 00°58′ S, 77°45′ W, ca. 1400 m), QCAZ 18618–19, 22363. Quebrada Pangayaku (0.78255 S, 77.791 W, 929 m), MZUTI 1680–81; Reserva Narupa (0.684 S, 77.741 W, 1179–1208 m), ZSFQ 0437–38; Reserva Narupa (0.671 S, 77.774 W, 1502 m), ZSFQ 0439–41. Provincia de Morona Santiago: 6.7 km W of 9 de Octubre (02°13′30.5″ S, 78°17′25.6″ W, 1715 m), QCAZ 32643. Provincia de Tungurahua: near Río Negro (01°24′ S, 78°15′ W, 1423 m), on the Río Negro–Río Verde road, QCAZ 21252, 31729; Río Topo, BM 1912.11.1.69. Provincia de Zamora Chinchipe: stream tributary of the Río Jambue (ca. 4°15′ S, 78°56′ W, ca. 1820 m), ca. 16 km S from Zamora, Parque Nacional Podocarpus, DFCH-USFQ D125; gravel road E to Sarsa (3.80783976 S, 78.60593076 W; 1500 m), QCAZ 47053.

Photographic records: Provincia de Orellana: Reserva Río Bigal (0.532913° S, 77.423228° W; 981 m). Photo by Morley Read.

Genus Cochranella Taylor, 1951 [15].
Etymology: Named in honor of Doris M. Cochran, herpetologist and curator of the Smithsonian Institution, Washington, D.C.
Cochranella granulosa (Taylor, 1949 [145]; Figure 67 and Figure 68).
Centrolenella granulosa Taylor, 1949 [145]. Holotype: FMNH 178269.
Type locality: “Los Diamantes, one mile south of Guápiles, (Cantón de Pococí, Provincia Limón), Costa Rica”.
Cochranella granulosa—Taylor, 1951 [15].

Common names: English: Granular Glassfrog. Spanish: Rana de Cristal Granulosa.

Etymology: The specific epithet granulosa comes from the Latin word granum (grain), in allusion to the granular dorsal skin texture of the species.

Identification: Cochranella granulosa can be distinguished from most glassfrogs by having a dark green to bluish–green dorsum with whitish granules and faint to well-defined dark spots (Figure 67), yellowish–green hands and feet, a reduced white ventral parietal peritoneum (which covers less than 40% of the venter), white digestive tract, and by lacking humeral spines. Another distinctive trait is the granular texture of its dorsal skin. Species that might have a sympatric distribution with C. granulosa and that present a similar dorsal coloration include Cochranella litoralis and some populations of Espadarana prosoblepon. However, C. granulosa has a granular skin (shagreen in E. prosoblepon and C. litoralis) and lacks humeral spines (present in males of E. prosoblepon and C. litoralis). Also, C. litoralis has an orange to red iris (yellow to golden in C. granulosa) and E. prosoblepon has a digestive tract that lacks white iridophores (present in C. granulosa).

Figure 67. Cochranella granulosa in life. (Top row): Reserva Guayacán, Costa Rica; photos by Brian Kubicki. (Bottom row): Reserva El Jardín de los Sueños, 461 m, Cotopaxi province, Ecuador, MZUTI 4811; photos by Jaime Culebras (dorsolateral) and Javier Aznar (ventral).

Diagnosis: (1) Vomerine teeth present, each vomer with one to three teeth; (2) snout truncated in dorsal aspect, slightly sloping in lateral profile; (3) tympanum oriented dorsolaterally, with slight posterior inclination, its diameter about 35% of eye diameter; tympanic annulus visible, low supratympanic fold evident, tympanic membrane partially pigmented and clearly differentiated from surrounding skin; (4) dorsum granular, lacking spicules; (5) venter areolate, lacking pair of enlarged subcloacal warts; (6) white lining on the anterior 20%–40% of the ventral parietal peritoneum (condition P1); peritoneum with white iridophores covering digestive tract (condition V2); (7) liver tetralobed; hepatic peritoneum lacking iridophores (condition H0); (8) humeral spines absent; (9) webbing absent between Fingers I–II, vestigial to moderate between Fingers II–III, extensive between III–IV; webbing formula on hand: II (1⁺–2)—(3–3⁺) III (1^1/3–2^1/2)—(1–2⁺) IV; (10) webbing on foot extensive, formula: I 0⁺—1^1/2 II 0⁺—1^2/3 III (1–1⁺)—2⁻ IV 2—1⁻ V; (11) ulnar fold low; inner and outer tarsal folds low and thin; (12) concealed prepollex; nuptial pad Type I; (13) Finger I about same length as Finger II; (14) disc of Finger III narrow, about 40% of eye diameter; (15) in life, dorsum dark green to bluish green with fine, yellowish–white granules and faint to clearly marked dark spots (Figure 67); bones green; (16) in preservative, dorsum cream to light lavender with faint to clearly defined dark spots; (17) iris yellow to golden with few, faint brownish streaks; (18) melanophores absent from dorsal surfaces of hands and toes; (19) males call from the upper side of leaves; each call consists of three to five notes, the first note being slightly longer that the others; average duration for a three-note call is 0.8–0.9 s; dominant frequency at 3700 Hz; (20) males fight venter to venter, hanging from the vegetation by the tips of their toes; (21) dark egg are placed on the upper sides of leaves; short-term maternal care present; parental care by males absent; (22) tadpoles with a flattened body and slender tail; tadpole in early stages have a brown pigmentations that is lost in older, red larvae; jaw sheaths are strongly serrated, the upper jaw forms a smooth M-shape; the lower sheath is V-shaped; labial tooth row formula 2/3; (23) medium body size; in males, SVL 22.5–29.0 mm; in females, SVL 29–32 mm.

Color in life (Figure 67): Dorsal surfaces green to bluish green, with yellowish or bluish granules and faint to clearly marked blue to black spots. Anterior 20%–40% of venter white, posterior portion translucent. Iris yellow to golden. Pale or whitish lip in populations from Central America (Savage, 2002; Kubicki, 2007). One individual from Ecuador (MZUTI 4811) lacks dark dorsal spots.

Color in ethanol: Dorsum cream to light lavender to dark lavender with faint to clearly marked dark spots (the Ecuadorian specimen lacks dark spots, MZUTI 4811); iridophores on the anterior 20%–40% of the ventral parietal peritoneum; pericardium and gastrointestinal peritonea with white iridophores; hepatic peritoneum lacking iridophores.

Biology and ecology: The information below is from Kubicki [24] and RWM (pers. obs.) Cochranella granulosa is active at night in riparian habitats of streams through pristine tropical forest, but also sometimes along streams in disturbed areas. Males fight venter to venter, hanging from the vegetation by the tips of their toes. Reproduction usually occurs in riparian habitats, but Kubicki [24] also reported reproductive activity in a small forest pond about 10 m away from the nearest stream. Egg clutches consist of 40 to 104 eggs [24,146,147] that are attached on masses on the upper distal half of a leaf overhanging the stream. Following hydration, the eggs form a pendent mass (i.e., drip tip) that hangs off the leaf and thereby ensures a constant flow of water over the developing larvae [4,146]. There is no long-term parental care in this species [148]. Delia et al. [25] experimentally demonstrated that females provide short-term parental care (brooding) to their egg clutches just after oviposition; this behavior, although brief (i.e., few hours), reduces embryo mortality from dehydration.

Call: Males often call in a chorus, with one male initiating calling and then being answered by several males. The call consists of a rapid high-pitched pulsed trill, usually composed by three notes (range three to five notes; [24]). Usually, the first note is longer (average duration = 0.3 s) than the subsequent notes (average duration = 0.15 s). The mean duration for a three-note call is 0.8–0.9 s [24]. The dominant frequency is 3700–4500 Hz [24,149,150]. In Ecuador (Los Laureles, Cotopaxi province) males were actively calling in February 2016 (Jaime Culebras, pers. obs.).

Tadpoles: The tadpole of Cochranella granulosa was first described by Starrett [146]. Hoffmann [147] recently provided an additional and detailed description; below, we present a summary based on Hoffmann’s description. Tadpoles of C. granulosa and C. euknemos share a particular slim shape. Recently hatched tadpoles (one to five days old) are markedly slender and dark brown. In the laboratory, older tadpoles are slender with a very long tail; they lack much body pigment and their bright red coloration is from the highly vascularized internal anatomy anterior to the body cavity and visible through the body wall. The oral disc is not emarginate and is bordered laterally and along the posterior margin with 28–41 marginal papillae (mean = 33 ± 4 papillae). Jaw sheaths are strongly serrate; the upper jaw forms a smooth M-shape, and the lower sheath is V-shaped. Generally, all five tooth rows are well developed; LTRF is 2(2)/3; the A2 row has a large central gap with the two halves extending as short wings on each side of the lateral descendent ends of the upper jaw sheath.

Distribution (Figure 68): The distribution of Cochranella granulosa includes the humid lowland and premontane slopes from the Atlantic drainage of eastern Honduras to central Panama and on the Pacific versant in humid upland or gallery forests from northern Costa Rica to northern Ecuador, 40–1500 m elevation ([148,151], this work). Records of Cochranella granulosa in Ecuador are from El Jardín de los Sueños reserve, Cotopaxi province [151], and Durango, 238 m, Esmeraldas province.

Conservation status: Globally, Cochranella granulosa is listed as Least Concern by the IUCN [152]. We suggest Data Deficient in Ecuador, since it is known from two recent records (Figure 68).

Evolutionary relationships (Figure 69): Cochranella granulosa is sister to C. resplendens, being an example of allopatric speciation mediated by the uplift of the Andes.

Figure 68. Distribution of Cochranella granulosa in Ecuador (yellow dot).

Figure 69. Evolutionary relationships among species in the genus Cochranella, inferred using maximum likelihood and Bayesian criteria.

Remarks: Karyotype of Cochranella granulosa is 2N = 20 [153].

Specimens examined: Cochranella granulosa: Ecuador: Provincia de Esmeraldas: 4 km W Durango (1.042° N, 78.1081° W, 253 m), QCAZ 32769. Provincia de Cotopaxi: Reserva El Jardín de los Sueños (0.8416° S, 79.2006° W, 461 m), MZUTI 4811.

Cochranella litoralis (Ruiz-Carranza and Lynch, 1996 [154]; Figure 70, Figure 71, Figure 72 and Figure 73).
Centrolene litoralis Ruiz-Carranza and Lynch, 1996 [154]. Holotype: ICN 13821.
Type locality: “Departamento de Nariño, municipio de Tumaco, La Guayacana, Litoral Pacífico, 1°49.8′ Latitud N, 78°46.2′ W de Greenwich, 100 m, (Colombia)”.
Centrolene litorale—Frost, 2004 [155].
Cochranella litoralis—Guayasamin, Castroviejo-Fisher, Trueb, Ayarzagüena, Rada, and Vilà, 2009 [1].

Common names: English: Litoral Glassfrog. Spanish: Rana de Cristal del Litoral.

Etymology: The specific epithet litoralis makes reference to the proximity of the type locality to the ocean [154].

Identification: Cochranella litoralis is the only species in Centrolenidae that has a bright orange to red iris (Figure 70 and Figure 71) and adult males with humeral spines and a nuptial pad that partially covers the two innermost fingers (Figure 72). Additionally, its dorsum is pale yellow green with small grey to black dots and faint yellow–cream dorsolateral stripes. Males are small (SVL < 20 mm) and have humeral and distinct prepollex. In the Chocoan lowlands, only some populations of Espadarana prosoblepon and Nymphargus griffithsi have a green dorsum with dark dots and humeral spines. Both E. prosoblepon and N. griffithsi are considerably larger, have a cream to white iris, and lack prepollical spines. The Peruvian Cochranella guayasamini also has a red iris and humeral spines, but lacks dark dorsal spots [19].

Figure 70. Cochranella litoralis in life. Adult male, QCAZ 27693, from near Durango, 220 m, Esmeraldas province, Ecuador. Photos by Luis A. Coloma.

Diagnosis: (1) Vomers lacking teeth; (2) snout truncated in dorsal and lateral profiles (Figure 72); (3) tympanum of moderate size, oriented almost vertically, with slight lateral and posterior inclinations, its diameter about 30% of eye diameter; tympanic annulus mostly visible, with supratympanic fold covering its posterodorsal margin; tympanic membrane translucent, partially pigmented, clearly differentiated from surrounding skin; (4) dorsal skin smooth anteriorly and shagreen in sacral region, lacking spicules; (5) lacking pair of enlarged subcloacal warts; (6) anterior half of ventral parietal peritoneum with iridophores, posterior half translucent (condition P2); iridophores in pericardium and gastrointestinal peritoneum; no iridophores on testes, gall bladder, urinary bladder, and kidneys (condition V2); (7) liver with clearly defined lobes covered by transparent peritoneum (condition H0); (8) males with small and pointy humeral spines; (9) webbing absent between Fingers I and II, absent or vestigial between Fingers II and III, moderate between outer fingers; webbing formula IV 2—2⁻ V; (10) webbing between toes moderate; webbing formula on foot I 1—2 II 1—2⁺ III 1—2^1/2 IV 2^1/2—1 V; (11) ulnar and tarsal folds absent or low and inconspicuous, lacking white coloration; (12) distinct prepollex; nuptial pad Type V; (13) Finger I as long as Finger II; (14) disc of Finger III width about 33% of eye diameter; (15) in life, dorsum green usually with small dark spots and faint yellow–cream dorsolateral stripes (Figure 70 and Figure 71); pale green bones; (16) in preservative, dorsum lavender with dark spots; (17) in life, iris orange to red; (18) dorsal surfaces of fingers and toes lacking melanophores, except for some on base of Toe V; (19) calling behavior unknown; call undescribed; (20) fighting behavior unknown; (21) egg deposition site unknown; parental care unknown; (22) tadpoles unknown; (23) minute body size; in two adult males, SVL 19.4–20.0 mm; females unknown.

Figure 71. Cochranella litoralis in life. Male from Tundaloma Lodge (1.182 N, 78.749 W; 74 m), Esmeraldas province, Ecuador (3 January, 2014; not collected). Photo by Lucas Bustamante/Tropical Herping.

Color in life (Figure 70 and Figure 71): Cochranella litoralis has a pale yellowish–green dorsum with dark spots that vary in size and conspicuousness; the anterior half of the venter is white, turning transparent posteriorly; the iris is bright orange to red.

Color in ethanol: Dorsal surfaces cream lavender with small dark spots, and two faint white dorsolateral lines. White parietal peritoneum covering anterior half of venter. White pericardium; white peritonea covering stomach and lower colon. Liver, testes, kidneys, and urinary and gall bladders covered by translucent peritonea. Iris silvery white, with black punctuations.

Biology and ecology: A male of Cochranella litoralis and an egg clutch presumed to be from the same species were found on riverine vegetation (L. Bustamante, pers. comm.). The egg clutch was placed on the upper side of a leaf and contained 25 embryos (Figure 71). Parental care unknown.

Call: Not described.

Tadpole: Not described.

Figure 72. Cochranella litoralis, holotype, adult male, ICN 13821. (A) Head in lateral view. (B) Head in dorsal view. (C) Fingers I and II in dorsal view; note that nuptial pad extends on the two fingers; illustrated by Juan M. Guayasamin. (A,B) Modified from Ruiz-Carranza and Lynch [154].

Distribution (Figure 73): Cochranella litoralis has been reported from the type locality in Colombia and four localities in Ecuador at elevations below 260 m ([130,154], this work). In Ecuador, this species has a potential distribution of 5784 km² within the Chocoan Tropical Forest region.

Figure 73. Distribution of Cochranella litoralis in Ecuador (yellow dots).

Conservation status: Globally, Cochranella litoralis is currently listed as Vulnerable by the IUCN [156]. In Ecuador, it has a distribution restricted to the provinces of Esmeraldas and Cotopaxi (Chocó ecoregion), where logging is causing continuous habitat reduction and fragmentation. The most recent reports of this species are from Río Cachabí (September 2005), Tundaloma (January 2014), and Jardín de los Sueños (2018). In Ecuador, because of habitat loss and mining, we suggest that the species should be considered as Endangered, following IUCN criteria B2a, B2(iii).

Evolutionary relationships (Figure 69): Cochranella litoralis is sister to all other member of the genus Cochranella, but with low nodal support. Twomey et al. [19] recovered a sister relationship between C. litoralis and C. nola.

Specimens examined: Cochranella litoralis: Colombia: Departamento de Nariño: Municipio de Tumaco, La Guayacana (1.83 N, 78.77 W; 100 m), ICN 13821. Ecuador: Provincia de Esmeraldas: stream near Durango (1.047 N, 78.618 W; 220 m), QCAZ 27693; Pichiyacu, Comunidad Chachi, Río Cayapas (0.9397° N, 79.005° W; 260 m), QCAZ 31705; Río Cachabí (1.03 N, 78.77 W, 200 m), 2 km NE Urbina on the San Lorenzo—Lita road, DHMECN 3198; Tundaloma Lodge (1.17868° N, 78.7497° W; 74 m), MZUTI 3481.

Localities from the literature: Cochranella litoralis: Ecuador: Provincia de Esmeraldas: Tsejpu, Río Zapallo (0.7 N, 78.9 W, 150 m) [156].

Cochranella mache Guayasamin and Bonaccorso, 2004 [157] (Figure 74, Figure 75 and Figure 76).
Cochranella mache Guayasamin and Bonaccorso, 2004 [157]. Holotype: QCAZ 22412.
Type locality: “Riachuelo La Ducha (0°20′41″ N, 79°42′36″ W; 510 m), tributary of Río Aguacatal, Reserva Biológica Bilsa, 27.4 km W (airline distance) of the town of Quinindé, Montañas del Mache, Provincia Esmeraldas, Ecuador”.

Common names: English: Mache Glassfrog. Spanish: Rana de Cristal de Mache.

Etymology: The specific name mache is a noun in apposition and refers to the Montañas de Mache, the type locality of the species [157].

Identification: Cochranella mache is differentiated from most glassfrogs by having a snout gradually inclined in profile and dermal folds with white tubercles on the ventrolateral edges of Finger V, forearm, elbow, Toe V, tarsus, and heel (Figure 74 and Figure 75). Species sharing similar characteristics are: Centrolene daidalea, C. savagei, C. solitaria, Cochranella resplendens, and C. euknemos. Three of these species (C. daidalea, C. savagei, and C. solitaria) are restricted to the Andes, and Cochranella resplendens is found only in the Amazon Basin. The closely related C. euknemos lacks the tubercles that are characteristic in C. mache [157].

Figure 74. Cochranella mache in life. Adult male, QCAZ 27764, from Río La Carolina, 500 m, Esmeraldas province, Ecuador. Photos by Luis A. Coloma.

Diagnosis: (1) Each vomer with three or four teeth on dentigerous process; (2) snout subacuminate in dorsal aspect and gradually inclined in lateral profile (Figure 75); (3) tympanum oriented almost vertically, with slight lateral and posterior inclinations, its diameter about 33%–37% of eye diameter; tympanic annulus visible, low supratympanic fold evident, tympanic membrane translucent and pigmented as surrounding skin; (4) dorsal skin shagreen with warts that usually correspond to light spots; males with numerous minute spicules; (5) ventral skin granular; several round, enameled warts around cloaca; cloacal fold present; pair of enlarged subcloacal warts; (6) anterior one-third of parietal peritoneum covered by iridophores (condition P1); iridophores over pericardium and visceral peritonea (digestive tract, gonads); renal capsules covered by iridophores in some individuals (condition V2); (7) liver tri- or tetralobed, hepatic peritoneum clear or with small, isolated patches of iridophores (condition H0); (8) humeral spines absent; (9) webbing absent between Fingers I and II, reduced between Fingers II and III, and extensive between outer fingers (Figure 75); webbing formula II (1⁻–1⁺)—(3⁻–3⁺) III (2⁻–2)—(1–1⁺) IV; (10) webbing between toes extensive (Figure 75); webbing formula on foot I (1⁻–1)—(2–2⁻) II 1—2 III (1–1⁻)—2⁻ IV (2⁻–2)—(1–1⁻) V; (11) ventrolateral edges of Finger IV, forearm, elbow, Toe V, tarsus, and heel with dermal enameled folds and tubercles; (12) concealed prepollex; nuptial pad large; (13) Finger I about same length as Finger II (Finger I about 94%–98% of Finger II); (14) disc of Finger III width about 44%–54% of eye diameter; (15) color in life, dark olive-green to lavender–blue or pale blue dorsum with numerous small yellow to orange spots, large dull to bright yellow patch on top of head (Figure 74); (16) color in preservative, dorsal surfaces pale lavender with small white or cream spots; tubercles on dermal folds of limbs, fingers, and toes cream-white; inner fingers and toes white or unpigmented; (17) in life, iris whitish cream to beige with thin brown reticulation; white to golden circumpupilary ring; (18) melanophores covering dorsal surfaces of Fingers III and IV and Toes IV and V; (19) males call from the upper sides of leaves; each call formed by two notes; dominant frequency at 5383–5426 Hz; (20) fighting behavior unknown; (21) brown eggs placed on upper sides of leaves; parental care unknown; (22) tadpoles unknown; (23) small body size; in adult males, SVL 22.0–27.0 mm (n = 43); in adult females, SVL 28.0–33.4 mm (n = 4).

Figure 75. Cochranella mache. (A) Head in lateral view, paratype, adult male, KU 291176. (B) Nuptial excrescences on dorsal surface of Finger I, paratype, adult male, QCAZ 22413. (C) Foot in ventral view, KU 291176. (D) Hand in ventral view, KU 291176. Hand and foot not drawn to scale. Modified from Guayasamin and Bonaccorso [157].

Color in life (Figure 74): Dorsal surfaces vary from green to lavender blue or pale blue, with small yellow to orange spots. Upper lip with a thin, white margin. White tubercles visible on dermal folds of hind- and forelimbs. Innermost fingers and toes (Fingers I and II, Toes I and II) white in males and partially white in females. Throat and ventral surfaces of limbs blue green. White parietal peritoneum covering anterior half of venter, posterior portion translucent; white pericardial and visceral peritonea. Transparent hepatic peritoneum, but in some individuals, a small patch of white iridophores covers part of liver. White warts surround cloaca. Iris whitish cream to beige with thin brown reticulation; white to golden ring surrounding pupil. Bones green ([157,158], this work).

Color in ethanol: Dorsum of head, body, and limbs lavender with small white or cream spots. Anterior one-third of parietal peritoneum covered by iridophores; iridophores also cover pericardium and visceral peritonea (digestive tract and gonads); hepatic peritoneum lacking iridophores or with small, isolated patches of iridophores; renal capsules covered by iridophores in some specimens ([157,158], this work).

Biology and ecology: The information shown below was obtained mainly from Ortega-Andrade et al. [159] and, to a lesser extent, from Guayasamin and Bonaccorso [157]. Cochranella mache seems to be restricted to small streams and rivulets in primary and secondary forests in lowlands and piedmont forest. Abundance of the species is correlated with rainfall, being higher during rainy season. Males call from the upper surface of leaves or branches on trees and bushes; reproduction seems to be restricted to the rainy season. Although individuals have been observed on different strata of the forest up to 6 m high, C. mache prefers the midstory vegetation. Amplexus is axillary and a gravid female was observed to have about 30 eggs. The diversity of amphibians and reptiles at the type locality (Reserva Biológica Bilsa) of C. mache is described by Ortega-Andrade et al. [160]. Parental care unknown.

Call: The advertisement call of Cochranella mache was described by Ortega-Andrade et al. [159]. The recorded male was calling from the upper side of a dead Heliconia leaf, about 2.5 m above ground, horizontally separated from the stream by about 3m. A call consists of two notes, separated by an interval of 0.107–0.130 s. Each note has a duration of 0.038 ± 0.008 (0.029–0.049) s. The dominant frequency is at 5410.2 ± 17.9 (5383–5426) Hz. The harmonics are not visible. The call rate is 1.46 calls per minute.

Tadpole: Not described.

Distribution (Figure 76): Cochranella mache has been reported from few localities in northwestern Ecuador, provinces of Esmeraldas, Imbabura, and Manabí, at elevations between 38 and 800 m ([157,158,159], this work), and from the Pacific lowlands of Colombia, Departments of Antioquia and Valle del Cauca, at elevations of 750–1030 m [161]. Three localities lie within protected areas (Reserva Biológica Bilsa, Reserva Biológica Canandé, Reserva Jama-Coaque). In Ecuador, the potential distribution of C. mache is 27,433 km² (see also Ortega-Andrade et al. [159]) within the Chocoan Tropical Forest and the Western Foothill Forest regions. When deforestation is taken into account, ~70% of the predicted distribution is reduced [159].

Figure 76. Distribution of Cochranella mache in Ecuador (yellow dots).

Conservation status: Listed as Endangered by the IUCN at a global level [162]. Other studies suggest that the species should be classified into the category of Critically Endangered [159,163]. The distribution of the species is likely to correspond to the remaining lowland forest in northwestern Ecuador and southwestern Colombia, an area under the constant pressure by wood companies. Although the range of C. mache is partially within the Mache-Chindul Ecological Reserve and three private reserves (Reserva Biológica Bilsa, Reserva Biológica Canandé, Reserva Jama-Coaque), most conservation measures are ineffective because of institutional and funding restrictions and a lack of law enforcement. Some of the larger fragments are preserved by private organizations but many remain unprotected. Habitat degradation is mainly caused by unsustainable timber extraction, uncontrolled expansion of the agricultural frontier, and replacement by non-native plantations [159,163]. Records from Colombia, however, expand considerably the distribution of C. mache. More than half of this distribution area of the species is affected by human activities [159]. Under scenarios of climate changes, the potential distribution of C. mache suffers a reduction of 13%–21% of its predicted range [159].

Evolutionary relationships (Figure 69): Cochranella mache is the sister species of C. euknemos.

Specimens examined: Cochranella mache: Ecuador: Provincia de Esmeraldas: Montañas de Mache, Reserva Biológica Bilsa, Riachuelo La Ducha (0°20′41″ N, 79°42′36″ W; 510 m), tributary of Río Aguacatal, 27.4 km W (airline distance) of the town of Quinindé, QCAZ 22412 (holotype), QCAZ 22413, KU 291176; Río Balthazar (0.9745 N, 78.61675 W; 645 m), QCAZ 27747, 31327; Monte Saíno, Punta Galeras region (0.700 N, 80.017 W; 100 m), DHMECN 2611; 3 km NW of Quinindé (0.350 N, 79.483 W; 150 m), DFCH-USFQ LQ23; Reserva Biológica Canandé (0.433 N, 79.133 W; 270 m), DHMECN 3560. Provincia de Imbabura: Río La Carolina (0.70449 N, 78.20115 W; 500 m), on the Ibarra–San Lorenzo Road, nearby Jijón y Caamaño, QCAZ 27764. Colombia: Departamento de Antioquia: Municipio Dabeiba, Río Amparradó, Quebrada Iotó, 805 m, ICN 10689–90, 8665; Municipio Frontino, Vereda Venados, PNN Las Oriquideas, Quebrada La Miguera, 1030 m, ICN 19638–9.

Records from the literature: Ortega-Andrade et al. [159]: Ecuador: Comunidad San Salvador (0.496710 N, 79.85298 W; 38 m); Hacienda Shangrilá (0.18630 N, 79.03019 W; 499 m).

Photographic record: Ryan Lynch: Ecuador: Provincia de Manabí: Reserva Jama-Coaque (0.0978 S, 80.147 W).

Cochranella resplendens Lynch and Duellman, 1973 [22] (Figure 77, Figure 78, Figure 79, Figure 80, Figure 81 and Figure 82).
Cochranella resplendens Lynch and Duellman, 1973 [22]. Holotype: KU 118053.
Type locality: “Santa Cecilia, 340 m, Provincia de Napo (Sucumbíos), Ecuador”.
Cochranella resplendens—Ruiz-Carranza and Lynch, 1991a. Guayasamin, Castroviejo-Fisher, Trueb, Ayarzagüena, Rada, and Vilà, 2009 [1].
Cochranella phryxa Aguayo and Harvey, 2006 [164]. Holotype: CBG 778. Type locality: “approximately 20 km west of Población de la Cascada (Territorio Comunitario de Origen y Reserva de la Biósfera Pilón Lajas), La Paz Department, Bolivia (15°22′37″ S, 67°12′4″ W), 1000 m”. New synonymy.

Common names: English: Resplendent Glassfrog. Spanish: Rana de Cristal Resplandeciente.

Etymology: The specific name resplendens is derived from the Latin verb resplendo, meaning to glitter, and is used in allusion to the jewel-like appearance of this frog [22].

Identification: Cochranella resplendens is distinguished from most glassfrogs by having a snout gradually inclined in profile and ventrolateral edges of Finger IV, forearm, elbow, Toe V, tarsus, and heel with dermal folds and enameled tubercles (Figure 77, Figure 78, Figure 81). Additionally, the species lacks humeral spines and has a white venter except for the posterior one-fourth, which is transparent. The following species could be confused with Cochranella resplendens: Centrolene daidalea, Cochranella mache, Centrolene savagei, and Centrolene solitaria. However, none of these species is found in the Amazon basin, where Cochranella resplendens occurs; C. mache is distributed in the Pacific lowlands of Ecuador, whereas C. daidalea, C. savagei, and C. solitaria are endemic to the Colombian Andes.

Figure 77. Cochranella resplendens in life. (Left): Amplectant pair. (Right): Female brooding clutch. Locality: 2 km N from HW 20 on a dirt road out of Guagua Sumaco, Napo province, Ecuador. Photos by Jesse Delia.

Diagnosis: (1) Each vomer with three or four teeth on dentigerous process; (2) snout round in dorsal aspect and gradually inclined in lateral profile (Figure 78); (3) tympanum oriented almost vertically, with slight lateral and posterior inclinations, its diameter about 38% of eye diameter; tympanic annulus partially visible, low supratympanic fold evident, tympanic membrane barely translucent, pigmented as surrounding skin; (4) dorsal skin shagreen with elevated and spiculated warts corresponding to white spots; (5) venter granular; pair of slightly enlarged subcloacal warts; enlarged cloacal fold present; (6) white lining (iridophores) on the anterior three-fourths of the ventral parietal peritoneum, posterior one-fourth transparent (condition P3); iridophores in pericardium and peritonea covering intestines and stomach; renal capsules and gall bladder and urinary bladder lacking iridophores (condition V2); (7) liver lobate, covered by transparent peritoneum (condition H0); (8) humeral spines absent; (9) webbing absent between inner fingers, and extensive between outer fingers (Figure 78); webbing formula III (1^1/2–2⁻)—(1–1^1/2) IV; (10) webbing between toes extensive (Figure 81); webbing formula on foot I (1–1⁺)—(1^2/3–2) II (0–0⁺)—(2–2⁺) III (1–1^1/2)—(2–2⁺) IV (2⁻–2⁺)—(1⁻–1⁺) V; (11) ventrolateral edges of Finger IV, forearm, elbow, Toe V, tarsus, and heel with dermal folds and enameled tubercles (Figure 77, Figure 79 and Figure 81); (12) concealed prepollex; nuptial pad not evident; (13) Finger I about same length as Finger II; (14) disc of Finger III width about 55% of eye diameter; (15) in life, dorsum dark green with numerous small white spots (Figure 77 and Figure 79); bones green; (16) in preservative, dorsum lavender with small white spots (Figure 81); (17) iris whitish cream to beige, with a fine grey to brown reticulation; pale yellow circumpapilar ring; (18) dorsal surfaces of fingers and toes with few small, enameled spots; melanophores only on surfaces of Finger IV and Toe V; (19) calling behavior unknown; call undescribed; (20) fighting behavior unknown; (21) egg clutches placed on upper sides of leaves; short-term maternal care present; parental care provided by males absent; (22) tadpoles with non-emarginate oral apparatus; tooth row formula 2(2)/3; upper jaw slightly curved; (23) medium body size; in two adult males, SVL 26.5–26.6 mm; females unknown.

Figure 78. Cochranella resplendens, holotype KU 118053. (A) Head in dorsal view. (B) Head in lateral view. (C) Hand in ventral view. Drawings of head not to scale. Illustrations by Juan M. Guayasamin.

Color in life (Figure 77 and Figure 79): Dorsum green with numerous, small white to bluish–white spots. Ventral surfaces mostly white, except posterior portion, which is translucent. Upper lip, tubercles on forearm and foot, and cloacal warts white. Bones green. Iris whitish cream to beige, with fine grey to brown reticulation; pale yellow circumpapilar ring ([22,164], this work).

Color in ethanol (Figure 81): Dorsal surfaces of head, body, and limbs slate grey to lavender with small white spots, often corresponding to small, flat tubercles. Upper lip white; tympanum pigmented as surrounding skin. White tubercles present on Fingers III and IV and Toes II–V. White on dermal folds of forearm and tarsus and on tips of Finger IV and Toe V. White cloacal tubercles. Pericardium and anterior three-fourths of the ventral parietal peritoneum white. White peritonea covering intestine, stomach, and colon. Transparent peritonea covering kidney, gall bladder, and urinary bladder. Hepatic peritoneum clear in holotype and most known specimens, but with a patch of iridophores in two Ecuadorian specimens and in Bolivian specimen ([22,164], this work).

Biology and ecology: Cochranella resplendens is a rare species. Since its description in 1973, only eight additional individuals have been reported (Specimens Examined). The scarcity of Cochranella resplendens is not related to low sampling effort. At the type locality (Santa Cecilia), fieldwork from 1967 through 1972 resulted in the collection of 7765 specimens of amphibians and reptiles, but no additional specimens of Cochranella resplendens [165]. However, canopy surveys may reveal that its rarity is an artifact of microhabitat sampling [166]. Sixty-six species of amphibians, including Teratohyla midas and Hyalinobatrachium munozorum, are known to occur at Santa Cecilia [165]. At the Bolivian locality (20 km W of Población La Cascada), Cochranella resplendens was found sympatric with Rulyrana spiculata and Cochranella sp. [164]. At the Tiputini Biodiversity Station, Cochranella resplendens was found sympatric with Teratohyla midas, Hyalinobatrachium munozorum, and Vitreorana ritae ([134], this work). Females provide short-term parental care; male parental care is absent [25].

Figure 79. Life cycle in Cochranella resplendens. (A) Juvenile, QCAZ 38088. (B) Ontogenetic variation of tadpoles. (C) Egg clutch. Photos by Luis A. Coloma. Figure modified from Terán-Valdez et al. [167].

Tadpole (Figure 79 and Figure 80): At Río Napinaza, a clutch with 74 embryos was found on the upper side of a leaf [167]. According to Terán-Valdez et al. [167], the tadpole of Cochranella resplendens has the following traits (based on tadpole in Gosner Stage 36): Body elongated, oval-depressed (sensu [168]), wider than high; snout rounded in dorsal and lateral views. Eyes located on dorsal surface of head; in early stages, eyes C-shaped; after Stage 35, eyes become round. Short, single, sinistral spiracle, at the posterolateral region of the body; vent tube short and abdominal, free posteriorly, opening directed posteriorly; myotomes visible throughout length of tail; dorsal fin originating at about mid-length of tail; ventral fin originating almost at base of tail muscle and reaching its maximum height posterior to mid-length of tail. Oral disc non-emarginate; marginal papillae uniserial, distributed around oral disc, but larger on the lower labium; upper labium with papillae only on lateral extremes. Upper jaw sheath completely keratinized with serrated edge; slightly curved. Lower jaw sheath keratinized, U-shaped, and with serrated edge. Labial tooth row formula 2(2)/3; A-2 with medial gap. For ontogenetic variation, see Terán-Valdez et al. [167]. Dorsally, the tadpole is mostly red, with some aggregations of iridophores that form an interorbital line and a middorsal band from behind the eyes to nearly the end of the body (based on a tadpole in Gosner Stage 36). Dorsally, the anterior-most part of the body is pale yellow. Laterally, the tadpole is translucent. The iris is mostly black with a yellow ring around the pupil [167].

Figure 80. Oral apparatus of different species of Centrolenidae (from Terán-Valdez et al. [167]). Cochranella granulosa and Hyalinobatrachium ibama are in Gosner Stage 24, Cochranella resplendens in Stage 36, H. cappellei in Stage 25, H. aureoguttatum in Stage 35. Gosner stages of the remaining species are not provided in the original descriptions. Figures modified from: (A,G) Rada et al. [169]; (B,D,F,I,J) Starrett [146]; (C,E) Terán-Valdez et al. [167], (H) Noonan and Bonett [170].

Taxonomic Remarks: Aguayo and Harvey [164] described Cochranella phryxa from the Amazonian slopes of the Bolivian Andes. These authors mentioned that C. phryxa can be distinguished from Cochranella resplendens (characteristics in parenthesis) by having a hidden tympanum (tympanum visible), a first finger longer than the second (second finger longer than the first), and a straight cloacal fold (U-shaped cloacal fold). After examining the holotype and six additional specimens (QCAZ 38099, MHNSM 19507, DFCH-USFQ D103–4; FHGO 1305, 1324) of Cochranella resplendens, we find that the tympanic membrane in Cochranella resplendens is only slightly differentiated from the skin surrounding the tympanum. Therefore, one could interpret the tympanum in Cochranella resplendens as hidden, which is the character state that Aguayo and Harvey [164] described for C. phryxa. The relative length of the two innermost fingers has been used for many authors when comparing centrolenid species. Although we agree that the character is useful when the differences are obvious (e.g., comparing species of Hyalinobatrachium), it is problematic when differences are minor, as in C. resplendens. We have tried to minimize the error in the assessment of this character by measuring the fingers with a digital caliper. Consistently, we found that these fingers have almost the same length in Cochranella resplendens (Finger I length 96%–102% of Finger II). The last character used by Aguayo and Harvey [164] is the shape of the cloacal fold, which they described as straight in C. phryxa and U-shaped in Cochranella resplendens. Differences in shape of the cloacal ornaments may be either artificial, associated with how the specimens are preserved (i.e., posterior extremities fixed with an anterior, posterior, or parallel orientation in relation to the sagittal axis of body), or due to intraspecific variation. The cloacal ornamentation (i.e., white warts and folds) is the same in the two species (compare Lynch and Duellman [22]: Figure 2c; Aguayo and Harvey [164]: Figure 4). Cochranella mache, a species with U-shaped cloacal folds, shows the same intraspecific variation observed between C. resplendens and C. phryxa. The only difference that we find between the holotype of Cochranella resplendens and the Bolivian specimen is the presence of a small patch of iridophores on the hepatic peritoneum of the latter. However, it is possible that this patch is either an artifact of preservation (i.e., iridophores from the ventral parietal peritoneum can be attached to the liver) or intraspecific variation (i.e., the presence/absence of patches of iridophores in the peritoneum has been observed in other centrolenids (e.g., Cochranella mache [17,158]). For the reasons discussed above, we formally place Cochranella phryxa Aguayo and Harvey, 2006 [164], in the synonymy of Centrolenella resplendens Lynch and Duellman, 1973 [22].

Figure 81. Cochranella resplendens in preservative. Holotype (KU 118053) from Santa Cecilia, Sucumbíos province, Ecuador. (A) Dorsal view. (B) Ventral view. (C) Lateral view. Photos by Martín Bustamante.

Distribution (Figure 82): Cochranella resplendens in known from the Amazon basin of Colombia, Ecuador, Peru, and Bolivia, at elevations between 190 and 1100 m ([22,134,164,167,171], this work).

A disjunct population of the species was recently found on the eastern slope of the central Andes in the Departamento Antioquia, Colombia, at much higher elevations (1309–1699 m) [172]. In Ecuador, this species is known from localities in the provinces of Morona Santiago, Napo, Orellana, Pastaza, and Sucumbíos at elevations between 250–1100 m. In Ecuador, the potential distribution of C. resplendens is 77,793 km² within the Amazonian Tropical Rainforest and Eastern Foothill Forest ecoregions.

Figure 82. Distribution of Cochranella resplendens in Ecuador (yellow dots).

Conservation status: Globally, Cochranella resplendens is currently listed as Least Concern by the IUCN [173]. Although it is evident that C. resplendens is rare in collections (see above), we presume that this is a consequence of inadequate sampling [166]. The potential distribution of this species in Ecuador is 77,793 km², 14% of which is affected by human activities. Given the broad distribution of C. resplendens and the lack of immediate threats, the category of Least Concern is justified.

Evolutionary relationships (Figure 69): With the current genetic and taxon sampling, the sister species of C. resplendens is C. granulosa. These two species are an example of vicariant speciation, mediated by the uplift of the Andes.

Specimens examined: Cochranella resplendens: Colombia: Departamento de Putumayo: Santa María de Sucumbíos (ca. 00°16′ N, 76°55′ W), AMNH 88083 (Lynch and Duellman, 1973). Ecuador: Provincia de Sucumbíos: Santa Cecilia (00°03′ N, 76°58′ W; 340 m), KU 118053 (holotype). Provincia de Morona Santiago: Río Napinaza (2.927° N, 78.407° W; 1100 m), QCAZ 38088. Provincia de Napo: Reserva Yachana (00°52′21.71″ S, 77°14′13.43″ W; 300–350 m), QCAZ 38099; Provincia de Orellana: San José Viejo de Sumaco (0.5333 S, 77.4167 W; ca. 810 m), USNM 288460; Tiputini Biodiversity Station (00°37′ S, 76°10′ W; 190–270 m), DFCH-USFQ D103–04; Provincia de Pastaza: Pozo Garza, Oryx (01°26′ S, 77°03′ W; 300 m), FHGO 1305, 1324. Peru: Departamento de San Martín: Cainarachi Valley (6°43′10 S, 76°29′13 W; 550 m), Km 33, Carretera Tarapoto-Yurimaguas, MHNSM 19507. Bolivia: Departamento de La Paz: approximately 20 km west of Población de la Cascada (15°22′37″ S, 67°12′4″ W; 1000 m), CBG 778 [164].

Photographic records: Ecuador: Provincia de Napo: ca. 2 km north from HW 20 on a dirt road out of Guagua Sumaco (0.688° S, 77.6° W; ca. 900 m); photographs by Jesse Delia (Figure 77).

Genus Espadarana Guayasamin, Castroviejo-Fisher, Trueb, Ayarzagüena, Rada, & Vilà 2009 [1].
Etymology: The name Espadarana honors Marcos Jiménez de la Espada, a Spanish zoologist who was part of the Comisión Científica del Pacífico that explored America between 1862 and 1865. Jiménez de la Espada described the first centrolenid frog, Centrolene geckoideum in 1872. In Spanish, the word Espada means sword, in allusion to the humeral spines present in males of species in this genus. Espadarana is a combination of the words Espada and rana (frog) and is feminine in gender [1].
Espadarana audax (Lynch and Duellman, 1973 [22]; Figure 83, Figure 84, Figure 85, Figure 86 and Figure 87).
Centrolenella audax Lynch and Duellman, 1973 [22]. Holotype: KU 146624.
Type locality: “Salto de Agua, 2.5 km NNE of Río Reventador on Quito–Lago Agrio road, 1660 m, Provincia de Napo, Ecuador”.
Centrolene audax—Ruiz-Carranza and Lynch, 1991 [6].
“Centrolene” audax—Guayasamin, Castroviejo-Fisher, Trueb, Ayarzagüena, Rada, Vilà, 2009 [1].
Centrolene fernandoi—Duellman and Schulte, 1993 [174]. Synonymy by Cisneros-Heredia and Guayasamin, 2014 [175].
Espadarana audax—Twomey, Delia, and Castroviejo-Fisher, 2014 [19].

Common names: English: Daring Glassfrog. Spanish: Rana de Cristal Audaz.

Etymology: The specific name audax is Latin, meaning daring, and is used in allusion to the precipitous regions inhabited by this species [22].

Identification: Espadarana audax is unique among Ecuadorian glassfrogs by having small yellow spots on the dorsum, short and distally curved humeral spines in males, and extensive webbing only between Fingers III and IV (Figure 83, Figure 84 and Figure 85). In Ecuador, species with a similar dorsal coloration include Nymphargus buenaventura, N. siren, N. humboldti sp. nov., Rulyrana flavopunctata, and Teratohyla midas; however, males of these species lack humeral spines. In addition, Teratohyla midas has visceral peritonea covered by iridophores; N. buenaventura, N. siren, and N. humboldti sp. nov. have considerably less hand webbing; and R. flavopunctata has more webbing between Fingers III and IV. The Colombian Centrolene notosticta has a similar dorsal color pattern but lacks vomerine teeth (present in E. audax) and has less webbing between the outer fingers. Espadarana audax is most similar to E. durrellorum (see Taxonomic Remarks), but the latter differs by lacking yellow dorsal spots. Also, the two species are found at slightly different, although overlapping, elevations; E. audax inhabits the Amazonian slopes of the Andes (800–1900 m), whereas E. durrellorum is found mostly in the Amazonian lowlands and foothills (220–1150 m).

Figure 83. Espadarana audax in life. Adult male, QCAZ 37871, from Gral. Leonidas Plaza Gutiérrez (Limón), Quebrada del Río Napinaza, Morona Santiago, Ecuador. Photos by Luis A. Coloma.

Diagnosis: (1) Dentigerous process of the vomer bearing two to four teeth; (2) snout rounded in dorsal profile, rounded to truncated in lateral profile; (3) tympanum of moderate size, tympanum diameter 23%–29% of eye diameter, low supratympanic fold evident, tympanic membrane partially pigmented, clearly differentiated from surrounding skin; (4) dorsal surfaces of males and females shagreen, with minute spicules in males; (5) pair of enlarged subcloacal warts, other cloacal ornamentation absent; (6) anterior two-thirds of the ventral parietal peritoneum covered with white iridophores, posterior third transparent (condition P2); white pericardium; no iridophores in peritonea covering intestines, stomach, testes, kidneys, gall bladder, and urinary bladder (condition V1); (7) liver tetralobed, hepatic peritoneum lacking iridophores (condition H0); (8) in adult males, humeral spines present; (9) webbing absent between Fingers I, II, and III; webbing formula for outer fingers III (2⁻–2^1/3)—(2⁻–2⁺) IV; (10) extensively webbed foot: I (1–1⁺)—(2–2⁺) II (1⁻–1⁺)—(2–2^1/4) III (1–1^1/4)—(2⁺–2^1/3) IV (2⁺–2^1/3)—(1–1⁺) V; (11) ulnar fold present, inner tarsal fold present, outer tarsal fold absent; (12) concealed prepollex; in males, nuptial pad Type I; (13) Finger I about same length as Finger II (Finger II 95.6%–102% length of Finger I); (14) disc of Finger III moderate, its width 43%–56% of eye diameter; (15) in life, dorsal surfaces of head, body, and limbs green with small yellow spots; bones green; (16) in ethanol, dorsal surfaces of head, body, and limbs lavender with small white spots; (17) in life, iris white with yellowish hue and thin black reticulations; (18) melanophores on outer fingers and outer toes; (19) males call from upper sides of leaves; calls are produced in series, with each series having four or five calls; each call is composed by a single, pulsed note; dominant frequency is at 5426–6718 (mean = 6146, SD = 368) Hz; (20) fighting behavior unknown; (21) eggs deposited in moss on branches; short-term maternal care present; parental care by males absent; (22) tadpoles unknown; (23) small body size; in adult males, SVL 21.6–25.5 mm (X = 23.5 ± 0.722, n = 41); in adult females, SVL 24.5–28.8 (X = 27.0 ± 1.468, n = 7).

Figure 84. Espadarana audax, adult males. (A) Head in lateral view, KU 164503. (B) Hand in ventral view, KU 178018. Illustrations by Juan M. Guayasamin.

Figure 85. Humeral spines of Espadarana audax and E. durrellorum. (A) Espadarana audax, KU 164500. (B) Espadarana durrellorum, QCAZ 47909. Photos by Juan M. Guayasamin.

Variation: In several males, the dorsal spicules are minute and only visible under magnification. A population from Leonidas Plaza Gutiérrez (Limón) has a dorsum with very few and small dorsal yellow spots.

Color in life (Figure 83): Dorsum green, with small yellow spots; fingers and toes pale yellow; parietal peritoneum white; heart not visible; visceral peritoneum lacking iridophores. Ventral surfaces of limbs unpigmented; bones green; iris pale bronze to mustard with thin black reticulation ([22], this work).

Color in ethanol: Dorsal surfaces of head, body, and limbs lavender with small white dots; Fingers I–III and Toes I–III cream (without melanophores); upper lip white. White lining on the anterior two-thirds of ventral parietal peritoneum; white pericardium; no iridophores in peritonea covering liver, intestines, stomach, testes, kidneys, gall bladder, and urinary bladder. Some preserved individuals (KU 164497–98) have lost their dorsal white spots; however, close examination revealed the presence of collapsed chromatophores (apparently iridophores).

Biology and ecology: By day, one individual (KU 146624) was found in a bromeliad. During the night, adults are found on the upper sides of leaves along streams, a site from which males call (W. E. Duellman field notes, 19 March 1975; this work). On 18 July 1981, 12 individuals were found in Río Salado; the males were calling and one female (KU 178023) had dark brown oviductal eggs visible through the body wall. Females provide short-term parental care; male parental care is absent [25].

Call (Figure 86): We analyzed 14 notes contained within four calls from two individuals (MZUTI 1492–93). Calls are emitted in series; a typical call series has four or five calls (mean = 4.3, SD = 0.6) and is relatively long (range = 2398–3285 ms, mean = 2711 ms, SD = 497.8 ms). Each call has a single note with a duration of 26–53 (mean = 38, SD = 8.2) ms. Notes have clearly defined pulses with 5–11 (mean = 7.4, SD = 1.7) amplitude peaks throughout the note. Pulses within a note have a rate of 171–208 (mean = 189, SD = 10) pulses per second. Notes have their peak amplitude in the first 50% of the note (relative peak time: Range = 0.0144–0.0722, mean = 0.043, SD = 0.02), whereby the first note usually has the highest amplitude. The dominant frequency of a note measured at peak amplitude is 5426–6718 (mean = 6146, SD = 368) Hz and is contained within the fundamental frequency. The fundamental frequency has a lower limit of 5254–6288 (mean = 5925, SD = 374) Hz and a higher limit of 6115–7063 (mean = 6608, SD = 297) Hz.

Figure 86. Calls of Espadarana audax, MZUTI 1492, recorded at Río Hollín, 1038 m, Napo province, Ecuador. (A) Call series. (B) Single call.

Tadpole: Not described.

Distribution (Figure 87): Espadarana audax occurs through the Eastern Montane region of southeastern Colombia, Ecuador, and northern Peru at elevations between 800 and 1900 m. This species has been recorded from the provinces of Napo, Morona Santiago, Sucumbíos, and Zamora Chinchipe in Ecuador, the departments of Huila (eastern slope of Cordillera Central), Putumayo, Cauca, Caquetá, and Boyacá (eastern slope of Cordillera Oriental) in Colombia, and the Department of San Martín in Peru ([19,22,174,175,176], this work). In Ecuador, the species is found within the Eastern Foothill Forest and Eastern Montane Forest ecoregions. The potential distribution of E. audax is 6738 km², 9% of which is affected by human activities.

Figure 87. Distribution of Espadarana audax in Ecuador (yellow dots).

Conservation status: Globally, Espadarana audax is currently listed as Least Concern by the IUCN [177]. In Ecuador, E. audax has not been found in some of its historical localities (e.g., Río Azuela, Río Salado ([91], JMG, pers. obs.)), but has been discovered recently at several new localities (Miazi Alto, General Leonidas Plaza Gutiérrez, Zamora); see Specimens Examined. The distribution of the species is fragmented because of agriculture, pasture lands, and mining. We suggest that, in Ecuador, the species should be considered as Near Threatened.

Evolutionary relationships (Figure 88): Given the current gene and taxon sampling, Espadarana audax and E. durrellorum are sister species.

Taxonomic Remarks: The morphological differences that separate Espadarana durrellorum and E. audax are subtle, but given the available genetic and morphological data, we consider them as valid species. “Centrolene” fernandoi was recently placed in the synonymy of E. audax [175].

Specimens examined: Espadarana audax: Ecuador: Provincia Napo: 2.5 km NNE of Río Reventador on Quito–Lago Agrio road (0.133 S, 77.633 W, 1600 m), KU 146624 (holotype); 16.5 km NNE Santa Rosa (0.2186 S; 77.7319 W, 1700 m), KU 143290, 143292; 2 km SSW of junction between Río Reventador and Baeza-Lumbaqui road (0.1 S, 77.6 W, 1700 m), KU 164497–504; 7 km SW Río Azuela on Quito–Lago Agrio road (0.1667 S, 77.667 W), KU 155502–03; Río Salado (0.19167 S, 77.6997 W, 1420 m), KU 178018–27; 43 km NE Santa Rosa (0.1186 S, 77.6003 W, 1490 m), KU 190015; 8.9 km NE Santa Rosa on Quito–Lago Agrio road (0.1667 S, 77.667 W), KU 190016; Río Hollín (0.72 S, 77.639 W; 1100 m), QCAZ 6898; Reserva Narupa (0.6848 S, 77.742 W, 1164 m), ZSFQ 382–86. Provincia Sucumbíos: Río Azuela (0.11667 S, 77.6167 W, 1740 m), KU 164496. Provincia Morona Santiago: Quebrada del Río Napinaza, 6.6 km N in the road Limón–Macas (2.92665 S, 78.40701 W; 1100 m), QCAZ 37871, 29439; La Y (3.43236 S, 78.60449 W, 835 m), QCAZ 23910. Provincia Zamora Chinchipe: Alto Miazi (4.25026 S, 78.61746 W; 1250 m), QCAZ 41653; Cordillera del Cóndor, Centro Shuar El Tink, ca. 1050 m, QCAZ 48202. Colombia: Departamento Huila: Parque Arqueológico San Agustín, 3 km SW of San Agustín, 1750 m. Peru: Departamento San Martín: W slope of Abra Tangarana, 7 km (by road) NE of San Juan de Pacaysapa (6.2 S, 76.733 W; 1080 m), KU 211770–75.

Espadarana callistomma (Guayasamin and Trueb 2007 [9]; Figure 89, Figure 90, Figure 91 and Figure 92).
Centrolene callistommum—Guayasamin and Trueb, 2007 [9]. Holotype: QCAZ 25832.
Type locality: “stream affluent of Río Bogotá (1°05′13.8″ N, 78°41′25.8″ W, 83 m), nearby San Francisco de Bogotá, Provincia de Esmeraldas, Ecuador.”
Espadarana callistomma—Guayasamin, Castroviejo-Fisher, Trueb, Ayarzagüena, Rada, and Vilà, 2009 [1].

Common names: English: Beautiful eyes Glassfrog. Spanish: Rana de cristal de ojos bellos.

Etymology: The specific name callistomma is derived from the Greek kallistos–, meaning “most beautiful” and omma, meaning “eye”, as a reference to the fantastic iris pattern in this species [9].

Identification: Espadarana callistomma is easily distinguished from most glassfrogs by its large size (in males, SVL 26.7–29.6 mm; in females, SVL 29.5–31.8 mm), uniform green dorsal coloration, and silvery white iris with black reticulations (Figure 89). Espadarana callistomma closely resembles Sachatamia ilex; however, males of E. callistomma have a conspicuous humeral spine, whereas males of S. ilex have a small humeral spine that is embedded in the arm musculature (Figure 201). Some populations of Espadarana prosoblepon have the same color pattern as E. callistomma, but they have a significantly smaller body size (in males, SVL 23.2–27.5 mm; in females, SVL 25.3–27.8 mm) and a two-note call (three to four notes in E. callistomma). See Taxonomic Remarks.

Figure 89. Espadarana callistomma in life from Durango, Esmeraldas province, Ecuador. Adult male, QCAZ 32055. Photo by Martín Bustamante.

Diagnosis: (1) Vomerine teeth present, each vomer with two to seven teeth; (2) snout truncated in dorsal and lateral views (Figure 90); (3) tympanum small (tympanum diameter 20%–31% eye diameter), oriented vertically, with lateral inclination; tympanic annulus visible except for upper border, which is covered by supratympanic fold; tympanic membrane partially pigmented, differentiated from surrounding skin; (4) dorsal surfaces of males and females shagreen, with minute spicules evident in males (visible under magnification); (5) pair of enlarged subcloacal warts (Figure 90); (6) anterior two-thirds of ventral parietal peritoneum covered with white iridophores, posterior third transparent (condition P3); white pericardium; translucent peritoneum covering intestines, stomach, gall bladder, testes, and urinary bladder; kidneys covered with translucent peritoneum (condition V1); (7) liver tetralobed, covered by transparent peritoneum (condition H0); (8) humeral spines present in adult males; (9) no webbing between Fingers I and II, webbing between Fingers II and III reduced, webbing between Fingers III and IV extensive (Figure 90), webbing formula: II (1^2/3–2)—(3⁺–3^1/4) III (1^1/2–1^2/3)—(1–1^1/2) IV; (10) webbing on foot extensive (Figure 90), webbing formula: I (0⁺–1)—(2⁻–2⁺) II (0⁺–1)—(2⁻–2⁺) III (0⁺–1)—2⁻ IV (2⁻–2^1/3)—(1⁻–1⁺) V; (11) ulnar and inner tarsal folds low; outer tarsal fold absent; (12) concealed prepollex; in males, nuptial pad Type I; (13) Finger II slightly longer than Finger III (Finger III 93.3%–100% length of Finger II); (14) disc of Finger III of moderate size, about 29%–34% eye diameter; (15) in life, dorsum olive green (Figure 89); bones green; (16) in preservative, dorsum dark lavender; (17) iris silvery white with thin black reticulations; (18) melanophores covering dorsal surfaces of Fingers III and IV and Toes IV and V; (19) males call from upper side of leaves; each call consists of three to four pulsed notes, dominant frequency at 5343–5812 Hz; (20) fighting behavior unknown; (21) eggs deposited on upper side of leaves; parental care unknown; (22) tadpoles unknown; (23) in males, SVL 26.7–29.6 mm (X = 27.8 ± 0.762; n = 19); in females, SVL 29.5–31.8 mm (X = 30.5 ± 0.796; n = 13).

Figure 90. Espadarana callistomma. (A) Head in lateral and dorsal views, paratype, adult male, QCAZ 28803. (B) Ventral surfaces of thighs showing enlarged subcloacal tubercles, holotype, adult male, QCAZ 25832. (C) Hand and foot in ventral view, paratype, adult male, QCAZ 28803. (D) Nuptial pad and thumb in dorsal view, holotype, adult male, QCAZ 25832. Modified from Guayasamin and Trueb [9].

Osteology: A detailed osteological description of Espadarana callistomma was provided by Guayasamin and Trueb [9]; they mentioned that the frontoparietals were exceedingly slender along the orbital margin of the braincase. After reexamining the cleared-and-stained material, it seems that the frontoparietals extend medially and their medial margins partially delimit the frontal fontanelle (area shown as cartilage in [9]: Figure 8A).

Color in life (Figure 89): Dorsum uniform olive green, without spots; upper lip with whitish–cream coloration; iris silvery white with marked black reticulations; flanks white; parietal peritoneum white, covering anterior two-thirds of abdomen (heart not visible); bones green [9].

Color in preservative: Dorsal surfaces of head, body, and limbs uniform lavender; upper lip cream; iris white with marked dark lavender reticulations; nuptial pad on Finger II cream; dorsally, Fingers II and III and Toes I–III unpigmented; venter cream [9].

Biology and ecology: The natural history of Espadarana callistomma is poorly known. The species is active during the night and has been found on leaves along streams. Males call from the upper side of leaves, and females deposit pigmented eggs on the upper side of leaves. It is unknown if males or females provide parental care [9].

Call (Figure 91): Description based on a call (LBE-C-022) of a male E. callistomma recorded by JMG on 17 May 2010 at Reserva Otokiki, Ecuador. Each call consists of three to four pulsed notes (X= 3.7 ± 0.483, n = 10) and lasts 0.28–0.44 s (X= 0.38 ± 0.066, n = 10). Notes are produced at a rate of 9.1–11.0 notes/s (X= 9.76 ± 0.596, n = 10); each note has a duration of 0.014–0.04 s (X = 0.03 ± 0.007, n = 10). The call is not frequency modulated; dominant frequency at 5343–5812 Hz (n = 10).

Figure 91. Calls of Espadarana. (A) Call of Espadarana callistomma from Reserva Otokiki, 706 m, Esmeraldas province, Ecuador, LBE-C-022. (B) Call of E. prosoblepon from Cordillera de Chontilla, 908 m, Pichincha province, Ecuador, MZUTI 601.

Tadpole: Not described.

Distribution (Figure 92): Espadarana callistomma is known from four localities in the lowland rainforest of northwestern Ecuador (provinces of Esmeraldas and Carchi) at elevations below 500 m ([9], this work) and the Pacific lowlands of Colombia [178]. The habitat of the species is within the Chocoan Tropical Forest and the Western Foothill Forest regions (see Biogeographic Regions).

Figure 92. Distribution of Espadarana callistomma in Ecuador (yellow dots).

Conservation status: Globally, Espadarana callistomma is currently listed as Least Concern by the IUCN [179]. At present, E. callistomma is known from a few localities in the Pacific lowlands of Ecuador and Colombia [9,178]. The most likely scenario, however, is that E. callistomma is restricted to portions of the Chocó ecoregion, an area with high rates of deforestation and mining; we suggest that the species is considered as Endangered in Ecuador, following IUCN criteria B1, B2a, B2b(iii).

Evolutionary relationships (Figure 88): Espadarana prosoblepon is paraphyletic with respect to E. callistomma, although E. callistomma is monophyletic. The most likely explanations to the observed pattern are: (i) Divergence between the two species is recent and reciprocal monophyly was not yet achieved, (ii) E. callistomma is a synonym of E. prosoblepon, or (iii) E. prosoblepon is a species complex that requires further subdivision. Given the morphological and acoustic data at hand, we favor the first and third hypotheses.

Taxonomic Remarks: The specific status of Espadarana callistomma is uncertain because of the conflict among different sets of data. Genetically, Espadarana callistomma and E. prosoblepon are not reciprocally monophyletic (Figure 88). On the other hand, body size of males and females of E. callistomma is significantly larger than those of E. prosoblepon (t-test, P < 0.001), and the dorsal and iris color patterns of E. callistomma are different from most E. prosoblepon (Figure 89 and Figure 95). In terms of vocalization, E. callistomma has a call with more notes (three to four notes per call) than E. prosoblepon (two notes per call; Figure 91). Given that we find biologically meaningful differences among the species (i.e., body size, vocalizations), we recognize both species as valid. The lack of molecular divergence could be the product of a recent divergence, but this is a hypothesis that remains untested. Another pending taxonomic problem is that the type of Hylella parabambae matches the spotless coloration of E. callistomma and some populations of E. prosoblepon.

Specimens examined: Espadarana callistomma: Ecuador: Provincia de Esmeraldas: stream affluent of Río Bogotá (1°05′13.8″ N, 78°41′25.8″ W, 83 m), nearby San Francisco de Bogotá, QCAZ 25832 (holotype), 27776–8, 28555–56, 28557 (C&S), 28558; stream affluent of Río Bogotá (1°05′9.06″ N, 78°41′8.7″ W, 77 m), 2 km E of San Francisco de Bogotá on the San Francisco–Durango road, QCAZ 28803; 2.1 km E of Durango (1.02477 N, 78.61746 W; 284 m), QCAZ 32169. Provincia de Carchi: Río La Carolina (0°42′16.16″ N, 78°12′4.14″ W, 500 m), on the Ibarra–Lita road, nearby Jijón y Caamaño, QCAZ 27744–45, 27768.

Espadarana durrellorum (Cisneros-Heredia, 2007 [180]; Figure 85, Figure 93 and Figure 94).
Centrolene durrellorum Cisneros-Heredia, 2007 [180]. Holotype: DFCH-USFQ D131.
Type locality: “small rivulet tributary of the Jambue River, ca. 6 km S from Zamora (ca. 04°03′ S, 78°56′ W, 1150 m), on the western slope of Contrafuerte de Tzunantza, Cordillera Oriental, eastern slopes of the Andes, Provincia de Zamora Chinchipe, República del Ecuador”.
“Centrolene” durrellorum—Guayasamin, Castroviejo-Fisher, Trueb, Ayarzagüena, Rada, and Vilà, 2009 [1].
Espadarana durrellorum—Twomey, Delia, and Castroviejo-Fisher, 2014 [19].

Common names: English: Durrell’s Glassfrog. Spanish: Rana de Cristal de Durrell.

Etymology: The specific name durrellorum honours Gerald Durrell and Lee Durrell (Durrell Wildlife Conservation Trust) for their contributions to the conservation of biodiversity [180].

Identification: Espadarana durrellorum can be distinguished from all other glassfrogs by having a green dorsal coloration (Figure 93), humeral spines short and straight in adult males (Figure 85), moderate body size (25.7–26.1 mm adult males), and lacking iridophores on the visceral peritonea. Among Ecuadorian centrolenids, E. durrellorum is most similar to E. audax. The main trait to differentiate between the two species is the absence of yellow dorsal spots in E. durrellorum (present in E. audax). E. audax inhabits higher elevations (800 to 1900 m) than E. durrellorum (220–1150 m).

Figure 93. Espadarana durrellorum in life. Ecuador, Napo province, Comunidad Ñukanchi Allpa, 403 m, QCAZ 47909. Photos by Luis A. Coloma.

Diagnosis: (1) Vomerine teeth present; (2) snout rounded in dorsal view, bluntly truncated in lateral view; (3) tympanic annulus evident, oriented dorsolaterally; very weak supratympanic fold above tympanum; (4) dorsal skin shagreen, with minute spicules uniformly distributed; (5) ventral skin granular; cloacal area granular, with two large, rounded, flat subcloacal tubercles, and two folds on the sides of the cloacal opening; distinct cloacal sheath; (6) upper two-thirds of the parietal peritoneum covered by iridophores (condition P3), pericardium white, all other peritonea lacking iridophores (condition V1); (7) liver lobed, lacking iridophores (condition H0); (8) in adult males, humeral spine short and straight, with sharp point; (9) webbing absent between Fingers I and II, basal or absent between II and III, moderate between outer fingers: In males, III 2—2⁻ IV; in one female III 1^2/3—(1^1/2–2⁻) IV; (10) webbing formula on feet: In males, I 2—2⁻ II 1—2 III 1⁺—2⁻ IV 2^1/3—1⁺ V; in one female, I 1—1^3/4 II 1—2⁻ III 1⁺—2 IV 2—1 V; (11) no dermal folds on hands, forearms, feet, or tarsus; (12) nuptial excrescences present, Type I; concealed prepollex; (13) first finger slightly longer than second; (14) eye diameter larger than width of disc on finger III; (15) in life, dorsal surfaces uniform green; (16) in preservative, dorsal surfaces uniform pale lavender; (17) in life, iris golden with thin dark reticulations; in preservative, lavender with darker reticulation; (18) melanophores widespread on outer fingers and outer toes; (19) males call from upper surfaces of leaves; call undescribed; (20, 21, 22) fighting behavior, egg clutches, tadpoles, and parental care unknown; (23) small body size; SVL 24.8–26.1 mm in adult males (n = 3); 27.0 in one female.

Color in life (Figure 93): Uniform green dorsum without flecks or spots. Iris golden with dark reticulations. Green bones.

Color in ethanol: All dorsal surfaces pale lavender (no light or dark spots). Ventral surfaces cream. Parietal peritoneum covered by iridophores to the level of the lower stomach; pericardium white, all other peritonea lacking iridophores.

Biology and ecology: Espadarana durrellorum is a recently described species and little information is available on its natural history. It is nocturnal, and males call from the tops of leaves of riverine vegetation in mature Foothill and Lowland Evergreen forests. Parental care unknown.

Call: Not described.

Tadpole: Not described.

Distribution (Figure 94): Espadarana durrellorum is known from few localities on the foothills of Cordillera Oriental and the Amazonian lowlands of Ecuador, at elevations of 300–1267 m [176,180].

Figure 94. Distribution of Espadarana durrellorum in Ecuador (yellow dots).

Conservation status: Globally, E. durrellorum is Least Concern by the IUCN [181]; the species is known from few confirmed records, and a re-evaluation of its conservation status may be required.

Evolutionary relationships (Figure 88): Espadarana durrellorum and E. audax are sister species.

Taxonomic Remarks: See Taxonomic Remarks under Espadarana audax.

Specimens examined: Espadarana durrellorum: Ecuador: Provincia de Napo: ca. 45 km E of Narupa, on the Hollín–Loreto road (ca. 800 m), DFCH-USFQ D291; Parroquia Chontapunta, comunidad Ñukanchi Allpa, cabecera del río Canoayacu (0.99965° S, 77.39619° W; 403 m), QCAZ 47909; Reserva Yachana (00°52′21.7″ S, 77°14′13.4″ W; 300–350 m), DHMECN 03492, 06790–91; Pungarayacu (00°42′27.8″ N, 77°44′26.2″ W; 1267 m), DHMECN 03476. Provincia de Sucumbíos: Zábalo (0.3181333° S, 75.76625° W; 220 m), QCAZ 27832; Shuara (00°00′26″ N, 76°33′55.1″ W; 300 m), DHMECN 06794–95; Plataforma Shushufindi (00°05′14.8″ S, 76°40′03.8″ W; 330 m), DHMECN 06793. Provincia de Zamora Chinchipe: tributary of the Jambue River, ca. 6 km S from Zamora (ca. 04°03′ S, 78°56′ W; 1150 m), on the western slope of Contrafuerte de Tzunantza, Cordillera Oriental, eastern slopes of the Andes, DFCH-USFQ D131.

Espadarana prosoblepon (Boettger, 1892 [182]; Figure 91, Figure 95, Figure 96 and Figure 97).
Hyla prosoblepon Boettger, 1892 [182]. Syntypes: SMF 3756 (formerly 1400.1a, according to Boettger, 1892 [182]), and ZMB 28019, according to Duellman, 1977 [118]; SMF 3756 designated lectotype by Mertens, 1967.
Type locality: “Plantage Cairo (La Junta) bei Limon, (Cantón de Siquirres, Provincia de Limón) atlantische Seite von Costa Rica”. Savage, 1974 [183], commented on the type locality.
Hylella puncticrus Boulenger, 1896 [184]. Syntypes: BM 96.10.8.70–71. Type locality: “La Palma” San José Province, Costa Rica. Placed in synonymy by Günther, 1901 [185].
Hyla parabambae Boulenger, 1898. Holotype: BM 98.4.28.163. Type locality: “Paramba” Imbabura Province, Ecuador. Noted elsewhere in the original publication as “Paramba, a farm on the W. bank of the River Mira, at 3500 feet altitude; it is still in the forest region, but the open country commences two or three miles higher up the Mira”. Placed in synonymy by Lynch and Duellman, 1973 [22].
Centrolene prosoblepon—Noble, 1924 [186].
Cochranella parabambae—Taylor, 1951 [15].
Centrolenella parabambae—Goin, 1964 [187].
Centrolenella prosoblepon—Goin, 1964 [187].
Hyla ocellifera Boulenger, 1899. Holotype: BM 98.5.19.3. Type locality: “Paramba, (Provincia Imbabura,) N. W. Ecuador”. Placed in synonymy by Cisneros-Heredia and McDiarmid, 2007 [17].
Espadarana prosoblepon—Guayasamin, Castroviejo-Fisher, Trueb, Ayarzagüena, Rada, and Vilà, 2009 [1].

Common names: English: Variable Glassfrog. Spanish: Rana de Cristal Variable.

Etymology: The specific name prosoblepon is apparently derived from the Greek words proso (forward, onward, in front) and blepo (look or see), probably referring to the forward orientation of the eyes in centrolenid frogs [84].

Identification: Espadarana prosoblepon is distinguished from most glassfrogs from the Pacific versant of the Andes by having a large, flat, and projected humeral spine, and green dorsum with small black spots (Figure 95); however, E. prosoblepon has a highly variable dorsal coloration, including individuals that have both black and yellows spots, only yellow spots, only black spots, or absence of spots all together. In Ecuador, the only species that have a green dorsum with dark spots are Cochranella litoralis, Vitreorana ritae, Nymphargus cochranae, and N. megacheirus. In three of these species (V. ritae, N. cochranae, N. megacheirus), males lack humeral spines; additionally, they occur on the Amazonian lowlands or Amazonian slope of the Andes and are never in sympatry with E. prosoblepon. Cochranella litoralis has a bright orange iris and is smaller than E. prosoblepon (in males, SVL < 22.1 mm in C. litoralis; 23.2–27.5 mm in E. prosoblepon). Unspotted E. prosoblepon can be confused with E. callistomma and C. buckleyi; E. callistomma is distinguished by having a larger body size (in males, SVL 26.7–29.6 mm; in females, SVL 29.5–31.8 mm), contrasting black-and-white iris, and a call with three to four notes (call with two notes in E. prosoblepon; see Taxonomic remarks). Centrolene buckleyi has an inclined snout in lateral view (Figure 26) and is only found at elevations above 2000 m. Individual of E. prosoblepon with uniform dorsum and white iris with black reticulations resemble Sachatamia ilex; however, males of E. prosoblepon have a conspicuous humeral spine, whereas males of S. ilex have a small humeral spine that is embedded in the arm musculature (Figure 201). The closely related E. andina is similar to E. prosoblepon by having dark spots on the dorsum but has a pointy humeral spine that is almost parallel to the humerus, whereas in E. prosoblepon, the spine is broad, laminar, and projected at an angle of about 45° from the humerus.

Figure 95. Espadarana prosoblepon in life. Note variation of dorsal patterns. Frogs from Mindo and surroundings, Pichincha province, Ecuador; not collected. Photos by Alejandro Arteaga/Tropical Herping.

Diagnosis: (1) Each vomer with two to seven teeth on dentigerous process; (2) snout usually truncated in dorsal and lateral profiles (Figure 96); (3) tympanum small, oriented almost vertically, with slight lateral and posterior inclinations, its diameter 20.5%–30.8% of eye diameter; tympanic annulus mostly visible, with supratympanic fold covering its posterodorsal margin; tympanic membrane translucent, partially pigmented, clearly differentiated from surrounding skin; (4) dorsal skin shagreen; males and females may possess minute spicules on flanks and tympanic region; (5) pair of enlarged subcloacal warts; (6) white iridophores on the anterior 50%–70% of the ventral parietal peritoneum, posterior portion transparent (condition P2–P3); white pericardium; translucent peritonea covering intestines, stomach, kidneys, gall bladder, and urinary bladder (condition V1); (7) liver with four clearly defined lobes covered by transparent peritoneum (condition H0); (8) males with conspicuous humeral spines; (9) webbing absent between Fingers I and II, absent or basal between Fingers II and III; moderate between outer fingers (Figure 96); webbing formula III (1^2/3–2)—(1–2) IV; (10) webbing between toes moderate; webbing formula on foot I (0⁺–1)—(2⁻–2⁺) II (0⁺–1)—(2⁻–2⁺) III (0⁺–1)—(2⁻–2⁺) IV (2⁻–2^1/3)—(1–1⁺) V; (11) ulnar and tarsal folds absent or low and inconspicuous, lacking white coloration; (12) concealed prepollex; nuptial pad Type I; (13) Finger I as long as Finger II or slightly longer (Finger II length 93.8%–101% Finger I); (14) disc of Finger III moderate, width 28.6%–49.0% of eye diameter; (15) in life, dorsum green usually with small dark spots (Figure 95; but see Color in life section); bones green; (16) in preservative, dorsum lavender with or without cream and/or dark spots; (17) in life, iris varies from grey with fine dark reticulations to white with thick black reticulations; (18) melanophores covering dorsal surfaces of Fingers III and IV and Toes IV and V; (19) males call from upper side of leaves or small branches; (20) males fight upside down, grasping one another venter to venter; (21) black eggs are deposited on the upper surface of leaves, moss-covered rocks, or branches; short-term maternal care present; males do not provide parental care; (22) tadpoles with non-emarginated oral apparatus; tooth row formula 2/3; upper jaw curved; (23) small to medium body size; in adult males, SVL 23.2–27.5 mm (X = 24.8 ± 0.982, n = 53); in adult females, SVL 25.3–27.8 mm (X = 26.6 ± 0.859, n = 15).

Figure 96. Espadarana prosoblepon, KU 132462. (A) Head in lateral view. (B) Hand in ventral view. Illustrations by Juan M. Guayasamin

Color in life (Figure 95): In Costa Rica, dorsal surfaces green dorsum with dark spots [148]. About one-half of Honduran specimens lack small dark dorsal spots [84]. In Ecuador, several color patterns have been reported: (1) Uniform green dorsum; (2) dorsum green with dark spots; (2) dorsum green with yellow spots; (3) dorsum green with dark and yellow spots, separated from each other; and (4) dorsum green with dark spots surrounding yellow spots, forming false ocelli. Iris varies from greyish white to pale bronze with fine dark reticulation, to silvery white with thick black reticulations. Green bones; whitish cream upper lip; white venter that is transparent posteriorly; ventrolateral margins of forearm and tarsus lacking white coloration.

Color in ethanol: As described above, but green turns into lavender and yellow spots turn white. Some individuals (e.g., KU 132463–64) present black and smaller white spots on dorsum. White parietal peritoneum covering about anterior 60% of venter. Translucent peritonea cover digestive tract, liver, kidneys, and urinary and gall bladders; white pericardium.

Biology and ecology: The following information is based on studies by Jacobson [188], Hayes [189], Savage [148], Hoffmann [147], and Arteaga et al. [87]. At night, during the breeding season, Espadarana prosoblepon is found on vegetation along margins of streams in primary to slightly degraded evergreen forests and, sporadically, even in pastures. Reproduction peaks in the rainy season, when males become more vocal and often engage in aggressive territorial combats. Males initiate amplexus by jumping on the back of approaching gravid females. Females lay 20–52 black eggs on a variety of substrates and directly above the water. Females provide short-term parental care; male parental care is absent [25]. After hatching, tadpoles usually hide under leaf litter or sand at the bottom of streams. Males are territorial, and the advertisement call is used to space them along the stream. If an intruder moves into an occupied territory, the resident male produces a rapid series of calls; if the intruder approaches too closely, a fight ensues with both males dangling upside down while holding onto vegetation with their legs and grappling with their arms and humeral spines. The combat concludes when the loser drops from the fight site or signals submission by flattening his body against the leaf surface. Espadarana prosoblepon feeds on a variety of small arthropods including beetles, moths, spiders, and mites. As a defense mechanism, this glassfrog produces pungent odors when grabbed. Espadarana prosoblepon is the only glassfrog species for which survival information is available; McCaffery and Lips [190] report that mean annual survival probability is 0.46 (0.41–0.52, 95% CI). These authors also provide abundance data on several populations from Panama.

Call (Figure 91): The following description is based on a call of a male Espadarana prosoblepon (MZUTI 601) recorded by Italo Tapia during the night of 4 July 2012 at Río Sune Chico (908 m, Cordillera Chontilla, Pachijal-Mashpi) Pichincha province, Ecuador. Air temperature at the moment of the recording was 18.8 °C. Calls are produced every 4–7 min. Each call consists of two pulsed notes and lasts 0.207–0.223 s (X = 0.212 ± 0.007, n = 4). Each note has a duration of 0.032–0.054 notes/s (X = 0.040 ± 0.008, n = 8); time between the two notes of each call is 0.130–0.137 s (X = 0.134 ± 0.004, n = 4). Calls are frequency modulated; the dominant frequency at the beginning of each note is 5712–5825 Hz (X = 5759 ± 56.1, n = 4), whereas at the end of the note is at 6196–6394 Hz (X = 6252 ± 95.7, n = 4). Harmonics are also frequency modulated, showing complex spectral patterns.

The call of E. prosoblepon from Costa Rica shows some differences from the Ecuadorian call described above. The following information is from Jacobson [188] and describes a call from Monteverde, Costa Rica. Males of Espadarana prosoblepon call at irregular intervals (17 calls/h) from the upper surfaces of leaves during the night. The advertisement call consists of two to five short “beeps”. When encountering another frog within 15 cm, the resident males often give a series of rapid short beeps. Males of Espadarana prosoblepon call vigorously while in amplexus and also immediately after egg deposition. Kubicki [24] described the call of E. prosoblepon as a rapid three-note series, but sometimes males emit four to five notes; each note has an average duration of 0.03 s; the dominant frequency is 5.5–5.8 kHz. Typical call series have a total duration lasting 0.5 s (average three-note series) or 0.75 s (average four-note series). Because males call from exposed sites on the upper surfaces of leaves and twigs, these periodic calling bouts, often synchronized among a few, nearby males, may make it more difficult for predatory bats (i.e., Trachops cirrhosus) to locate and eat calling male E. prosoblepon (RWM, pers. obs.).

Tadpole: Females lay egg clutches on the upper side of leaves, moss-covered rocks, or branches; clutch size varies from 20 to 52 eggs that have a dark brown to black coloration; females remain with the eggs for some time (up to 131 min), but once they leave neither parent returns to the clutch [87,147,148,188,189]. Embryos are black but quickly turn yellow. By the time the tadpoles hatch and fall into the stream, they are red colored [148]. At Stage 25 [191], larvae have a sinistral spiracle, a relatively simple oral disc with medium-sized jaw sheaths, 2/3 tooth rows, and a single row of marginal papillae with a wide dorsal gap. The second anterior (upper) tooth row (A-2) also has a wide gap [148]. Drawings of the mouth and body can be found in Starrett [146], Savage [148]), and McCranie and Wilson [84].

Distribution (Figure 97): Espadarana prosoblepon is found from eastern Honduras, Nicaragua, Costa Rica, and Panama, south along the Pacific slopes of the Andes of Colombia and Ecuador from sea level to about 1600 m elevation. In Colombia, it occurs on the northern and eastern flanks of the Eastern Cordillera south to Caldas, and in the Magdalena Valley [192]. In Ecuador, Espadarana prosoblepon has been reported from several localities below 1620 m [87] (Specimens examined). The habitat of the species in Ecuador is mainly within the Chocoan Tropical Forest and the Western Foothill Forest regions, but it is also found in Deciduous Forest and the lower limit of the Western Montane Forest region. In Ecuador, it has a potential distribution of 41,612 km².

Figure 97. Distribution of Espadarana prosoblepon in Ecuador (yellow dots).

Conservation status: Globally, Espadarana prosoblepon is listed as Least Concern [192] in view of its large distribution and presumed large populations. In Ecuador, this species is common and is found even in habitats with moderate alterations; the category of Least Concern is justified.

Evolutionary relationships (Figure 88): Espadarana prosoblepon is paraphyletic, containing populations currently assigned to E. callistomma, a pattern that can be explained, mainly, by recent species divergence or the two species actually representing a single evolutionary lineage. See Taxonomic Remarks below.

Internal morphology: A detailed osteological and myological description of Espadarana prosoblepon was provided by Eaton [193]. The hyoid apparatus illustrated by Eaton lacks the anterolateral processes that are evident in cleared-and-stained material from Ecuador (KU 178163) and Costa Rica (KU 65178). We assume that the normal condition is the presence of these processes.

Taxonomic remarks: While the general morphology of Espadarana prosoblepon is mostly conservative, its color pattern is extremely variable, and because of an insufficient understanding of the species’ variation, different names have been assigned to distinctive morphs. For example, populations whose individuals have dorsal coloration consisting of black spots surrounding yellow spots and forming ocelli were called ocellifera, while others with dorsum lacking spots have been variously named parambae and callistomma. Several authors, including Lynch and Duellman [22], Savage [42], and Cisneros-Heredia and McDiarmid [17], have described some of the interspecific variation characteristic of E. prosoblepon, but much remains to be done. For example, most descriptions and color photographs of E. prosoblepon depict the iris of this species as grey with fine dark reticulations, but Kubicki [24] reported individuals with a white iris with black reticulations. Unfortunately, most centrolenid descriptions have paid little attention to iris patterns; in fact, with the exception of the descriptions of Sachatamia ilex, Hyalinobatrachium ignioculus, and H. eccentricum (the last two taxa now under the synonymy of H. cappellei), iris coloration, if reported at all, usually is based on examination of photographs; thus, the inter and intraspecific variation in iris color is poorly known. Genetic data suggest that E. prosoblepon and E. callistomma might represent one evolving lineage (Figure 88). Morphological differences between E. callistomma and E. prosoblepon are in body size (callistomma being larger), dorsal coloration (without spots in callistomma; usually with spots in prosoblepon), and iris coloration (white with contrasting black reticulation in E. callistomma; less contrasting in E. prosoblepon). In terms of vocalization, E. callistomma has a call with more notes (three to four notes per call) than E. prosoblepon (two notes per call; Figure 91). At the moment, we consider that the morphological and acoustic differences justify recognition of E. callistomma as a valid species.

Specimens examined: Espadarana prosoblepon: Ecuador: Provincia de Azuay: 12.9 km W Luz María (2.6889 S, 79.474 W, 740 m), KU 217502, QCAZ 12603–04; Provincia de Bolívar: Balzapamba (1.7667 S, 79.1833 W, 800 m), KU 132555–56, 132462–65; Provincia de Carchi: near Maldonado (0.9 N, 78.1 W, 1410 m), KU 178156–57; Provincia de Cotopaxi: near La Mana (0.933 S, 79.2167 W, 300 m), QCAZ 8641; near Sigchos (0.7 S, 78.883 W), USNM 288441; km 8 of the Pucayacu-Sigchos road (1.0097 S, 79.23769 W), QCAZ 40674. Provincia de Esmeraldas: Reserva Biológica Bilsa (0.3447 S, 79.71 W, 500 m), KU 291165–75, QCAZ 22414–17; Viruela, QCAZ 10267; Reserva Biológica Bilsa (0.3447 S, 79.71 W, 500 m), KU 291165–75; Reserva Ecológica Cotacachi-Cayapas, Charco Vicente (0.792 N, 79.1978 W; 60 m), QCAZ 11364–65; 5 km W of Durango (1.0858 N, 78.74 W), QCAZ 13206, 13212, 13242. Provincia de El Oro: near Valle Hermoso (3.50194 S, 79.81722 W; 379 m), QCAZ 37249; Río Chillayacu (3.32834 S, 79.58102 W; 395 m), 16.8 km W of Piñas (3.667 S, 79.667 W, 600 m), USNM 286738–39. Provincia del Guayas: Chongon-Colonche hills near Guayaquil, (ca. -2.1 S, -80.15 W), USNM 288438; Provincia de Imbabura: near Lita (0.833 N, 78.4667 W, 520 m), KU 133482–83; 6 km E of Lita (0.79472 N, 78.4286 W), QCAZ 4318–19; Zona de amortiguamiento de Reserva Cotacachi Cayapas, near Rio Aguas Verdes (0.331010° N, 78.93152° W; 670 m), QCAZ 46009; km 5 in the Lita-Ibarra road (0.84773 N, 78.42175 W), QCAZ 39919. Provincia de Los Ríos: Estación Biológica Río Palenque, 56 km N of Quevedo (0.55 S, 79.3667 W, 220 m), KU 164616–22. Provincia de Santo Domingo de los Tsáchilas: Río Orito, on the Toachi-Chiriboga road (0.30561 S, 78.882 W, 1315 m), QCAZ 15356; Río Faisanes, on the Toachi-Chiriboga road (0.2608 S, 78.845 W; 1400 m), QCAZ 15357, 15360, 15362; La Florida (0.28361 S, 79.0189 W), QCAZ 20726–28, 20730–32; Otongachi, near La Unión del Toachi (0.3167 S, 78.95 W; 900 m), QCAZ 25094; 5 km NE of La Florida (0.25694 S, 79.0539 W), QCAZ 7184–893; 4 km NE of the Dos Ríos–Chiriboga road (0.305139 S, 78.884333 W; 1270 m), QCAZ 31982; Santo Domingo de los Colorados (0.25 S, 79.15 W, 660 m), KU 121054–55; 4 km NE of Dos Ríos (0.30278 S, 78.8678 W, 1140 m), KU 164623–34; 2 km E and 1 km S of Santo Domingo de los Colorados (0.24512 S, 79.15509 W, 600 m), KU 178158–66; La Palma (0.3167 S, 78.9167 W, 920 m), KU 178167. Provincia de Pichincha: Reserva Maquipucuna (0.12429 N, 78.62936 W; 1343 m), QCAZ 42179; 6 km NW (by air) of Pedro Vicente Maldonado (0.10421 N, 79.10279 W; 544 m), QCAZ 35430; La Concordia, Bosque Protector La Perla (0.01 N, 79.4 W, 190 m), QCAZ 12602, Reserva Mashpi, 18 km N of San Miguel de Los Bancos (0.15 S, 78.883 W, 1100 m), DFCH-USFQ 293–95.

Genus Hyalinobatrachium Ruiz-Carranza and Lynch, 1991 [6].
Etymology: The generic name Hyalinobatrachium comes from the Greek words hyalos (glass) and batrachion (frog), alluding to the translucent and delicate appearance of the species in this genus [6].
Hyalinobatrachium adespinosai Guayasamin, Vieira, Glor, Hutter 2019 [194] (Figure 98, Figure 99, Figure 100 and Figure 101).
Hyalinobatrachium adespinosai Guayasamin, Vieira, Glor, Hutter, 2019 [194]. Holotype: ZSFQ 1648.
Type locality: “San Jacinto River (1.3447 S, 78.1814 W; 1795 m asl), Tungurahua province, Ecuador”.

Common names: English: Adela’s Glassfrog. Spanish: Rana de Cristal de Adela.

Etymology: The specific epithet adespinosai honors Adela Espinosa, an Ecuadorian conservationist and board member of the Jocotoco Foundation (http://www.jocotoco.org/wb#/EN/LaFundacion). Adela has concentrated her work to the conservation of species and ecosystems. Hyalinobatrachium adespinosai is found only within the limits of a natural reserve owned by Adela and her husband, Antonio Páez [194].

Identification: Among glassfrogs, H. adespinosai (Figure 98) is diagnosable mainly by having a transparent pericardium. However, it is morphologically cryptic with four closely related taxa (H. anachoretus, H. pellucidum, H. esmeralda); these species display a similar size and color pattern (pale green dorsum with yellow dots and a transparent venter and pericardium; red heart visible ventrally). However, calls have diverged noticeably (Figure 99); the major difference is the structure of the call, with two species (H. adespinosai and H. anachoretus) having pulsed calls and the others having tonal vocalizations. The call of H. adespinosai sp. nov. is further differentiated from that of H. anachoretus by being longer, having more pulses per note, and being produced at a higher rate [194]. Genetic distances for the 16S marker between H. adespinosai and closely related species (H. pellucidum, H. esmeralda, H. yaku) are >2.5%. However, H. adespinosai and H. anachoretus only have a 1% genetic distance, although they are separated by a major biogeographic barrier (i.e., Marañon river valley) [194].

Diagnosis: The following characters are found in Hyalinobatrachium adespinosai: (1) Dentigerous process of the vomer lacking teeth; (2) snout truncated in dorsal and lateral profiles; (3) tympanum barely visible, hidden under skin, with coloration similar to that of surrounding skin; (4) dorsal skin shagreen, lacking tubercles; (5) ventral skin areolate; cloacal ornamentation absent, paired round subcloacal warts absent; (6) parietal peritoneum transparent; pericardium with thin layer of iridophores (in life, a red heart is mostly visible ventrally); liver, viscera, and testes covered by iridophores; (7) liver white, bulbous; (8) humeral spines absent; (9) hand webbing formula: I (2–3)—(2–2⁺) II (1–1⁺)—3^1/3 III (2–2⁺)—(2⁻–2) IV; (10) foot webbing moderate; webbing formula: I 1—(1^2/3–2⁻) II (1–1⁻)—(2⁻–2^1/3) III (1–1⁺)—(2⁺–2^1/3) IV 2⁺—(1⁺–1^1/3) V; (11) fingers and toes with thin lateral fringes; ulnar and tarsal folds present, but difficult to distinguish, with thin layer of iridophores that extends to ventrolateral edge of Finger IV and Toe V; (12) nuptial excrescence present as a small pad on Finger I (Type V), concealed prepollex; (13) when appressed, Finger I longer than II; (14) diameter of eye about two times wider than disc on Finger III; (15) coloration in life: Dorsal surfaces pale yellowish green with small pale yellow spots and minute grey to black melanophores; bones white; (16) coloration in preservative: Dorsal surfaces pale cream with minute melanophores; (17) iris coloration in life: White with pale yellow hue and minute lavender spots; (18) melanophores absent from most fingers and toes, but present on Finger IV and Toes IV and V; (19) males call from underside of leaves; advertisement call consisting of single note, distinctly pulsed (9–13 pulses per call), with duration of 0.382–0.430 s, and dominant frequency at 4645–5001 Hz; (20) fighting behavior unknown; (21) males attend egg clutches located on the underside of leaves overhanging streams; clutch size of 22 embryos (n = 1); (22) tadpoles unknown; (23) SVL in adult males 20.5–22.2 mm (n = 3), females unknown.

Figure 98. Hyalinobatrachium adespinosai in life, holotype, ZSFQ 1648. Photos by Jose Vieira/Tropical Herping.

Color in life (Figure 98): Dorsal surfaces apple green to yellowish green with diffuse yellow spots and minute grey to black melanophores. Melanophores absent from fingers and toes, except Finger IV and Toes IV and V. Ventrally, parietal peritoneum and pericardium transparent, with a red heart always visible, even when a very thin layer of iridophores is present on the pericardium of some individuals. Visceral peritoneum of gall bladder and urinary bladder transparent; hepatic and visceral peritonea white; ventral vein red. Iris pale yellowish white, with numerous minute lavender spots. Bones white [194].

Color in ethanol: Dorsal surfaces cream dotted with minute dark lavender melanophores; venter uniform cream; visceral peritoneum lacking iridophores; pericardium with a very thin layer of iridophores. Iris silvery white with minute lavender melanophores [194].

Biology and Ecology: All individuals were found on the underside of leaves of riverine vegetation along the San Jacinto River. The section of river was fast-flowing and had visible rapids. Although the population is locally abundant, individuals are difficult to observe because they are usually found at the canopy level (4–16 m above ground level). Males were calling in the months of July and August. One male (ZSFQ 1648) was apparently guarding an egg clutch containing 22 embryos; both the adult male and the egg clutch were on the same leaf most of the time, but the male also moved to nearby leaves [194].

Call (Figure 99): The description is based on recording from nine individuals. The call of Hyalinobatrachium adespinosai has a striking resemblance to the chirp of a cricket. Each call is composed by a single and high-pitched pulsed note and has a duration of 0.38–0.44 s (X = 0.38 ± 0.017). Time between calls varied from 2.0–11.0 s (X = 4.58 ± 2.3). The fundamental frequency, the same as the dominant frequency, is at 4645–5203 Hz (X = 4855 ± 152). There is no frequency modulation. The first harmonic is at 9336–9754 Hz and the second harmonic is at 14,159–14,444 Hz [194].

Figure 99. Call of Hyalinobatrachium adespinosai, holotype, ZSFQ 1648, recorded in field conditions at the type locality (San Jacinto River, 1795 m asl), Tungurahua province, Ecuador). Air temperature = 18 °C. Obtained from Guayasamin et al. [194].

Tadpole: Not described.

Distribution. Hyalinobatrachium adespinosai is only known from the type locality: San Jacinto River (1.3447 S, 78.1814 W; 1795 m asl), Tungurahua Province, Ecuador (Figure 100) [194].

Figure 100. Distribution of Hyalinobatrachium adespinosai in Ecuador (yellow spot).

Evolutionary relationships. Hyalinobatrachium adespinosai is sister to the Peruvian H. anachoretus [194]. Since the species was recently described, it is not included in the Hyalinobatrachium tree shown in Figure 101. The most closely related species to H. adespinosai share several morphological traits, including a red heart exposed ventrally (H. adespinosai + H. anachoretus + H. pellucidum + H. yaku) [194].

Figure 101. Evolutionary relationships among species in the genus Hyalinobatrachium, inferred using maximum likelihood and Bayesian criteria.

Conservation status: Hyalinobatrachium adespinosai has not been evaluated by the IUCN. However, Guayasamin et al. [194] suggested that it should be considered as Data Deficient.

Specimens examined. Hyalinobatrachium adespinosai: Ecuador: Provincia de Tungurahua: San Jacinto River (1.3447 S, 78.1814 W; 1795 m asl), ZSFQ 1647–48, 1650–52 (type series).

Hyalinobatrachium aureoguttatum (Barrera-Rodríguez and Ruiz-Carranza, 1989 [195]; Figure 101, Figure 102, Figure 103, Figure 104 and Figure 105).
Centrolenella aureoguttata Barrera-Rodriguez and Ruiz-Carranza, 1989 [195]. Holotype: ICN 17506.
Type locality: “Colombia, Departamento del Chocó, vertiente occidental de la cordillera Occidental, Municipio El Carmen de Atrato, Km 23 carretera El Carmen–Quibdó, 5°47′ latitud N y 76°20′ W, 1030 m.s.n.m.”.
Hyalinobatrachium aureoguttatum—Ruiz-Carranza and Lynch, 1991 [6].

Common names: English: Sun Glassfrog. Spanish: Rana de Cristal del Sol.

Etymology: The specific name aureoguttatum is derived from the Latin words aureus (gold) and guttatus (dappled, speckled, spotted), referring to the dorsal pattern of the species [195].

Identification: Hyalinobatrachium aureoguttatum is easily distinguished from most glassfrogs by having a completely transparent ventral parietal peritoneum, a white bulbous liver, white visceral peritoneum, and by the absence of humeral spines. It differs from all other Hyalinobatrachium by having, in life, large yellow spots on the dorsum (Figure 102). The most similar and related species is H. valerioi, from which is differentiated mainly by having large dorsal yellow spots produced by xanthophores and iridophores (absent in H. valerioi; see Figure 121).

Figure 102. Hyalinobatrachium aureoguttatum in life. (Left and center) Adult male, QCAZ 45365; photos by L. A. Coloma. (Right) Adult male, QCAZ 32068; photo by Martín Bustamante.

Diagnosis: (1) Vomers lacking teeth; (2) snout truncated in dorsal and lateral profiles (Figure 103); (3) tympanum of moderate size (tympanum diameter 30%–34% of eye diameter), with dorsolateral orientation and posterior inclination, supratympanic fold low, tympanic membrane clearly differentiated from surrounding skin; (4) dorsal surfaces smooth to shagreen, lacking spicules; (5) venter smooth; lacking pair of enlarged subcloacal warts; (6) ventral parietal peritoneum transparent (condition P0); pericardium polymorphic, with or lacking iridophores; white peritonea covering intestines, stomach, and testes; transparent peritoneum covering kidneys, and urinary and gall bladders (Figure 102, Condition V5–V6); (7) liver bulbous, covered by white peritoneum (condition H2); (8) humeral spines absent; (9) basal webbing between Fingers I and II, extensive webbing between outer finger: I (2–2^1/4)—(2–2^1/4) II (1⁻–1⁺)—(2–3⁺) III (1–2⁺)—(1–1⁺) IV; (10) foot webbing extensive: I (0⁺–1)—(1–1^2/3) II (0⁺–1)—(1–1^1/2) III (0⁻–1)—(1^1/2–2) IV (1^1/2–2⁺)—(1⁻–1) V; (11) ulnar and tarsal folds absent; (12) concealed prepollex; in males, nuptial pad Type III; (13) Finger I longer than Finger II (Finger II about 87%–95% length of Finger I); (14) disc of Finger III of moderate size, 35%–56% of eye diameter; (15) in life, dorsum greenish yellow with two to seven large yellow spots, and with or without brown flecks; ventral parietal peritoneum transparent, pericardium polymorphic (white or transparent), gastrointestinal peritoneum white; bones white; (16) in ethanol, dorsum cream with large white spots, and with or without brown flecks; (17) in life, iris white to yellow, with minute dark lavender flecks around the pupil or forming an horizontal stripe; (18) fingers and toes lacking melanophores, except for few melanophores on proximal portions of Toes IV and V; (19) males usually call from the underside of leaves; call undescribed; (20) fighting behavior unknown; (21) eggs deposited on the underside of leaves; prolonged parental care provided by males; maternal care absent; (22) oral apparatus of tadpoles with an emarginate disc; M-shaped upper jaw sheath; tooth row formula 2(2)/3; (23) minute body size; in males, SVL 20.4–24.0 mm (X = 21.8 ± 0.631, n = 36); in females, SVL 22.9–23.9 mm (X = 23.3, n = 3).

Figure 103. Hyalinobatrachium aureoguttatum, adult male, QCAZ 27429. (A) Head in lateral view. (B) Head in dorsal view. (C) Hand in ventral view. (D) Foot in ventral view. Illustrations by Juan M. Guayasamin

Color in life (Figure 102): Dorsum greenish yellow with two to seven large yellow spots (diameter 0.5–2.1 mm), and with or without black flecks; upper lip unpigmented; ventral parietal peritoneum transparent; pericardium polymorphic, transparent (red heart visible ventrally) or white (heart not visible ventrally); transparent peritonea covering the kidneys, and urinary and gall bladders; white peritonea covering the liver, intestines, stomach, and testes; bones white; iris white to yellow, with minute dark lavender flecks around the pupil or forming a horizontal stripe.

Color in ethanol: Dorsum cream with large white spots and with or without dark flecks; iris white with dark lavender pigment; white peritonea covering the liver, intestines, stomach, and testes; pericardium polymorphic (white, partially white, or transparent).

Biology and Ecology: According to Barrera-Rodríguez and Ruiz-Carranza [195], most individuals of Hyalinobatrachium aureoguttatum were found on the underside of leaves (Heliconia spp.) 100–700 cm above small streams during the night. Amplectant pairs and egg clutches were found during July and August 1987. Amplexus is axillary, and eggs are deposited on the underside of leaves. In Durango (Provincia de Esmeraldas, Ecuador), individuals were reproductively active (amplectant pairs and egg clutches) 24–26 May 2006. At Estero Piedras, the species was reproducing in August 2007; a clutch with 36 eggs was found on the underside of a leaf on a bush along a stream [167]. Valencia-Aguilar et al. [196] studied populations from the Pacific lowlands of Colombia. They found that males exhibit high fidelity to their territory; each male repeatedly uses the same leaf (Heliconia sp., Anthurium sp., Philodendron sp., Cyclanthus sp., Calathea sp., Musa sp.) for perching, calling, mating, and clutch attendance. Territoriality seems to be low, since several males where found in close proximity and fights or aggressive behaviors were not observed in intrusion events by co-specific males.

Female place green eggs on the underside of leaves; clutches 25–49 eggs [167,195,196,197]. Males provide parental care during the day and night until hatching (mean = 17.1 days ± 1.8). Males are polygynous and simultaneously attend up to five clutches [196].

Call: Most males call from the undersides of leaves, but some have been observed calling from leaf tops. The specific spectral and temporal characteristics of the call are unknown.

Figure 104. Hyalinobatrachium aureoguttatum, ontogenetic variation of tadpoles. (A) Gosner Stage 25, QCAZ 37752. (B) Gosner Stage 27, QCAZ 32072. Photos by L. A. Coloma. Figure modified from Terán-Valdez et al. [167].

Tadpole: A description of the tadpole can be found in Ibáñez et al. [197] and Terán-Valdez et al. [167] (Figure 80 and Figure 104).

Distribution (Figure 105): Hyalinobatrachium aureoguttatum is known from extreme southwestern Panama and the Pacific lowlands and western slopes of the Cordillera Occidental of Colombia and Ecuador at elevations below 1340 m [101,167,195,197,198,199]. In Ecuador, this species has been reported from the provinces of Esmeraldas and Imbabura at elevations below 600 m (Specimens Examined). In Ecuador, the species has a potential distribution of 4,481 km² within the Chocoan Tropical Forest and Western Foothill Forest ecoregions.

Conservation status: Listed globally as Near Threatened by the IUCN [200]. Given that Ecuadorian populations of the species are fragmented because of agriculture, pasture lands, and mining, we suggest placing the species in the Endangered category based on IUCN criteria B1, B2a, B2b(iii). The species is partially protected within the Mache-Chindul reserve.

Evolutionary relationships (Figure 101): Hyalinobatrachium aureoguttatum and H. valerioi are sister species.

Remarks: Some populations from Colombia usually have small (but visible) brown flecks on the dorsal surfaces of the head and body; Ecuadorian populations lack these flecks (Figure 102). Also, all Colombian specimens examined by us have a white pericardium, a character that is polymorphic in the Ecuadorian populations (white, partially white, or transparent pericardium). It is possible that these differences correspond to independent evolutionary lineages; however, we prefer to maintain these populations as one species until more data (e.g., acoustic, molecular) are available.

Figure 105. Distribution of Hyalinobatrachium aureoguttatum in Ecuador (yellow dots).

Specimens examined: Hyalinobatrachium aureoguttatum: Ecuador: Provincia de Esmeraldas: stream affluent of the Río Durango (1.05 N, 78.6167 W; 100–150 m), QCAZ 27429, 6302, 6303, 6441–42, 28802; 2 km E of San Francisco (1.0872 N, 78.6905 W; 60–80 m), on the San Francisco-Durango road, QCAZ 32101–02, 32105, 32129, 32132–33; Río Quingue, nearby Caimito (0.72096 S, 80.09117 W, 47 m), QCAZ 37306. Provincia de Imbabura: 6 km SE of Lita (0.79 N, 78.43 W; 600 m), QCAZ 4323. Colombia: Departamento del Chocó: Municipio El Carmen de Atrato, km 23 on road El Carmen–Quibdó (5°47′ N; 76°20′ W, 1030 m), ICN 17507, 17509–10, 17512; km 44 on road El Carmen–Quibdó, 630 m, ICN 17252–54, 17515–16; km 53 on road El Carmen–Quibdó, 420 m, ICN 17248, 17257, 17260, 17262, 17266–67, 17520–21, 17525, 17527–28, 17531–34, 17536–37.

Hyalinobatrachium chirripoi (Taylor, 1958 [201]; Figure 106, Figure 107 and Figure 108).
Cochranella chirripoi Taylor, 1958 [201]. Holotype: KU 36865.
Type locality: “Cocales Creek, Suretka, (Cantón de Talamanca,) Limón Province”, Costa Rica. Savage [183] commented on the type locality.
Centrolenella chirripoi—Savage, 1967 [202].
Hyalinobatrachium chirripoi—Ruiz-Carranza and Lynch 1991 [6].
Hyalinobatrachium cardiacalyptum McCranie and Wilson, 1997 [203]. Holotype: USNM 342161. Type locality: “Caño El Cajón (14°21′ N, 85°29′ W), at its junction with the Río Patuca, Departamento de Olancho, Honduras, elevation 200–225 m”. Placed in synonymy by Cisneros-Heredia and McDiarmid, 2007 [17].

Common names: English: Chirripó Glassfrog. Spanish: Rana de Cristal Chirripó.

Etymology: The specific name chirripoi is named for the Chirripó Indians, local inhabitants of the area where the species was first found [201].

Identification: Hyalinobatrachium chirripoi is easily distinguished from most glassfrogs by having, in life, a lime green dorsum with small yellow spots, a completely transparent ventral parietal peritoneum (red heart visible ventrally; Figure 106), and by lacking humeral spines. Additionally, it differs from most Hyalinobatrachium by having more webbing between Fingers II and III.

Figure 106. Hyalinobatrachium chirripoi in life. Reserva Canandé, 372 m, Esmeraldas province, Ecuador. Photos by Jaime Culebras.

Diagnosis: (1) Vomers lacking teeth; (2) snout truncated in dorsal view, truncated to slightly protruding in lateral profile; (3) tympanum almost completely concealed, small when visible (tympanum diameter 2.3%–2.8% of SVL); tympanic membrane not differentiated from surrounding skin; (4) dorsal surfaces shagreen, lacking spicules; (5) venter areolate; lacking pair of enlarged subcloacal warts; (6) ventral parietal peritoneum transparent (condition P0); pericardium transparent; white peritonea covering the intestines and stomach; transparent peritoneum covering urinary and gall bladders; kidneys covered by a mostly translucent peritoneum, except for some iridophores on their ventral portions (condition V6); (7) liver bulbous, covered by white peritoneum (condition H2); (8) humeral spines absent; (9) basal to moderate webbing between Fingers I and II, extensive webbing between outer finger: I (2⁺–2^1/3)—(2–2⁻) II 1—(2^2/3–3⁻) III (2⁻–2)—(1^1/3–1^1/2) IV; (10) foot webbing extensive: I (0⁺–1)—(1^1/2–1^2/3) II 1—(1^1/2–1^2/3) III (0⁺–1)—2⁻ IV (2⁻–2)—(1–1⁺) V; (11) ulnar and tarsal folds present, with minute iridophores; (12) concealed prepollex; in males, nuptial pad Type V; (13) Finger I longer than Finger II (Finger II about 85%–90% length of Finger I); (14) disc of Finger III of moderate size, 38%–47% of eye diameter; (15) in life, dorsum line green with small yellow spots, venter transparent (red heart visible; Figure 106); bones white; (16) in ethanol, dorsum cream with minute lavender spots and iridophores; (17) in life, iris yellowish white, with minute dark lavender flecks; (18) fingers and toes lacking melanophores, except for few on Toes IV and V; (19) males call from the upper and undersides of leaves; call undescribed; (20) fighting behavior unknown; (21) eggs deposited on the underside of leaves; maternal care absent; prolonged parental care provided by males; (22) elongate tadpole, with emarginate oral disc; tooth row formula 2/3; (23) small body size; in Ecuadorian males, SVL 25.0–25.5 mm (n = 2); Savage [148] reported that adult males have a SVL = 24–26 mm and that females are probably larger.

Color in life (Figure 106): Dorsum lime green with several small yellow spots; ventral parietal peritoneum transparent; pericardium transparent (red heart visible ventrally); transparent peritonea covering urinary bladder; white peritonea covering liver, intestines, and stomach; transparent peritoneum covering most of kidneys; bones white; iris yellowish white, with minute dark lavender flecks.

Color in ethanol: Dorsum cream with minute lavender spots and iridophores; ventral parietal pericardium translucent; white peritonea covering liver, intestines, and stomach; pericardium transparent; iris silvery white with dark lavender flecks.

Biology and Ecology: According to Kubicki [24,204], in southeastern Costa Rica, Hyalinobatrachium chirripoi deposits 65–80 greenish–white eggs in a single layer on the underside of leaves overhanging streams. Males were most often seen calling from below palm fronds or other smooth leaves between 1–4 m above the water. Males were also seen calling from the upper sides of vegetation, but much less frequently. One male was seen guarding eggs during the night, and another male was observed guarding eggs during daylight hours [24]. At Reserva Itapoa, Ecuador, in June 2014, a male was observed calling and guarding an egg clutch on the underside of a leaf. Maternal care is absent; prolonged parental care is provided by males [25].

Call (Figure 107): Most males call from the undersides of leaves, but some have been observed calling from the upper sides [24,204]. The call is a high-pitched insect-like buzz, very similar to that of H. colymbiphyllum [204]. We analyzed two notes from one individual (LBE-019). The typical advertisement call is composed by a single note. Note duration is 230–270 (mean = 250, SD = 28.3) ms. Notes are strongly pulsed and have 14–16 (mean = 15, SD = 1.4) amplitude peaks throughout the note, with pulses becoming difficult to distinguish near the end of the note. Pulses within a note have a rate of 59–61 (mean = 60, SD = 1) pulses per second. Notes have their peak amplitude in the first 50% of the note (relative peak time: Range = 0.0623–0.0638, mean = 0.063, SD = 0.001), where the peak amplitude occurs in the first several pulses. The dominant frequency of a note measured at peak amplitude is 4565 (mean = 4565, SD = 0) Hz and is contained within the fundamental frequency. The fundamental frequency has a lower limit of 1895–3273 (mean = 2584, SD = 974) Hz and a higher limit of 4651–5082 (mean = 4866, SD = 305) Hz.

Figure 107. Call of Hyalinobatrachium chirripoi recorded at Reserva Itapoa, 321 m, Esmeraldas province, Ecuador, LBE-C-019.

Egg masses and tadpoles: Greenish–white eggs (53–80) are deposited in a single layer on the underside of leaves; as development continues the embryos become pale tannish red [24,147,204]. Below, we present a summary of the tadpole description by Hoffmann [147]. Hatchlings (Gosner stages 25 and 26) have an unusual pattern on their dorsum: Two rows of brown-pigmented, spider-like dots of melanophores stretch longitudinally over the entire body and join in a semicircle at the tip of snout; another line of pigmented spots extends dorsally over the tail musculature. Because of the general lack of pigment and high transparency, the body coloration is partially determined by the red color of the gills and heart. As development continues (Gosner stages 25–41), the tadpoles become very elongated and are among the most elongated centrolenid larvae known (ratio of body length to body width of 2.41 ± 0.22); they have a completely transparent skin and the dorsolateral lines are mostly lost; the body has a pale rose shine and the tail is cream or, sometimes, yellowish. The oral disc is emarginate and, except for a moderate dorsal gap, is bordered by a single row of about 45 marginal papillae; the ventral ones being flatter than the lateral ones. The LTRF is 2(2)/3; the tooth rows are about of equal width (P-3 is slightly narrower) and extend across most of the oral disc. The A-2 gap is about as wide as the upper jaw, which is broadly arched; the lower jaw is V-shaped; the edges of both jaw sheaths are edged by many narrow serrations [147].

Distribution (Figure 108): Hyalinobatrachium chirripoi is known from lowland forest localities in Central America in southeastern Costa Rica and eastern Panama, and south into South America in western Colombia and Ecuador at elevations below 600 m ([24,148,204], this work). In Ecuador, this species has been observed in few localities within the Esmeraldas province (Río Quingue, Río Bogotá, Reserva Itapoa, Tesoro Escondido, Reserva Canandé), in the Chocó ecoregion, at elevations below 320 m.

Conservation status: Listed globally as Least Concern by the IUCN [205]. In Ecuador, the species has been recently found in five localities in the Chocó Ecoregion, an area under constant deforestation pressure. Although the Least Concern category seems accurate at the global level, in Ecuador, the species is threatened by habitat destruction because of agriculture, pasture lands, and mining. Thus, we suggest placing the species in the Endangered category at the local level, based on IUCN criteria B1, B2a, B2b(iii).

Figure 108. Distribution of Hyalinobatrachium chirripoi in Ecuador (yellow dots).

Evolutionary relationships (Figure 101): Molecular data support a sister relationship between Hyalinobatrachium chirripoi and H. colymbiphyllum.

Specimens examined: Hyalinobatrachium chirripoi: Ecuador: Provincia de Esmeraldas: Río Quingue (0.722° N, 80.08° W; 47 m), QCAZ 37309, 48271; Río Bogotá, DFCH-USFQ C1903; Reserva Itapoa (0.513° N, 79.134° W; 320 m), MZUTI 3609–10; Tesoro Escondido (0.542° N, 79.145° W; 225 m), MZUTI 3625; Reserva Canandé (0.526° N, 79.209° W; 310 m), MZUTI 4745. Costa Rica: Limón Province: Suretka, along Cocales Creek, KU 36862–64, 36866–70.

Hyalinobatrachium fleischmanni (Boettger, 1893 [206]; Figure 109, Figure 110 and Figure 111).
Hylella fleischmanni Boettger, 1893 [206]. Lectotype: SMF 3760, designated by Mertens (1967) [207].
Type locality: “San José, Costa Rica”.
Hylella chrysops Cope, 1894 [208]. Neotype: SMF 3760 (= holotype of Hylella fleischmanni), designated by Starrett and Savage (1973) [209]. Placed in synonymy by Boulenger (1895) [210].
Centrolenella fleischmanni—Noble (1924) [186].
Centrolenella viridissima Taylor, 1942 [211]. Holotype: EHT-HMS 27725 (now FMNH 100093). Type locality: “Agua de Obispo, Guerrero”, Mexico. Placed in synonymy by Starrett and Savage (1973) [209].
Cochranella fleischmanni—Taylor (1951) [15].
Cochranella decorata Taylor, 1958. Holotype: KU 36896. Type locality: “Hda. La Florencia, about 3 miles west of Turrialba, Cartago Province, Costa Rica”. Placed in synonymy by Starrett and Savage (1973) [209].
Cochranella millepunctata Taylor, 1958 [201]. Holotype: KU 36887. Type locality: “La Palma, San José Province, Costa Rica”. Placed in synonymy by Starrett and Savage (1973) [209].
Centrolenella fleischmanni—Goin (1964) [187].
Hyalinobatrachium fleischmanni—Ruiz-Carranza and Lynch (1991) [6].

Common names: English: Fleischmann’s Glassfrog (Liner, 1994). Spanish: Rana de Cristal de Fleischmann.

Etymology: The specific name fleischmanni is a patronym for Carl Fleischmann, a German collector–naturalist who donated the specimens used by Boettger to describe the species [84].

Identification: Hyalinobatrachium fleischmanni is easily recognizable by having, in life, a lime green dorsum with small yellowish spots, a completely transparent ventral parietal peritoneum, white heart and digestive tract (pericardium and gastrointestinal peritoneum covered by iridophores), lacking humeral spines, and having reduced webbing between inner fingers (Figure 110). In the Ecuadorian Pacific lowlands, similar species include H. aureoguttatum, which has large yellow spots on the dorsum; H. valerioi, distinguished by having a green dorsal reticulum; and H. chirripoi, which, in life, has a transparent pericardium (red heart visible through ventral skin). We note that H. fleischmanni represents a species complex that requires taxonomic revision [212].

Figure 109. Hyalinobatrachium fleischmanni in life. Ecuador, near Durango, 77 m, QCAZ 32107. Photos by Martín Bustamante.

Diagnosis: (1) Vomerine teeth absent; (2) snout subacuminate in dorsal aspect and rounded in profile; (3) tympanum not visible; supratympanic fold absent; (4) dorsal skin shagreen; (5) venter smooth; lacking pair of enlarged subcloacal warts; (6) ventral parietal peritoneum transparent (condition P0); white peritoneum covering heart, intestines and stomach; transparent peritoneum on urinary bladder (condition V5); (7) liver bulbous, hepatic peritoneum white (condition H2); (8) humeral spines absent; (9) webbing reduced between Fingers I, II and III, moderate between outer fingers (Figure 110); webbing formula I (2–2⁺)—2 II (1–1⁺)—(3) III (2)—(1⁺–2) IV (1–2) IV; (10) webbing between toes moderate; webbing formula on feet I (1–1^1/2)—(2) II (1)—(2–2⁺) III (1)—(2⁺–2⁻) IV (2–2⁺)—(1–1⁺) V; (11) ulnar and tarsal fold absent; (12) concealed prepollex; nuptial pad Type IV; (13) Finger II shorter than Finger I; (14) eye diameter larger than width of disc on Finger III (disc of Finger III width 33%–37% of eye diameter); (15) in life, lime green dorsum with yellow spots (Figure 109); venter transparent, pericardium usually white (but see Color in life); bones white; (16) in preservative, dorsum cream; (17) in life, iris yellowish to greyish white with some dark punctuations; (18) fingers and toes lacking melanophores; (19) males call from the underside of leaves; the call consists of a single “wheet” note, with duration of 150–300 ms, and a dominant frequency of 3800–4500 Hz at the beginning and 4800–5300 HZ at the end; (20) fighting behavior varies from vent to vent to amplexus-like; (21) egg clutches usually laid on the underside of leaves; maternal care absent; males provide prolonged parental care; (22) oral apparatus complete; oral disc with single row of marginal papillae laterally and ventrally, wide dorsal gap; jaw sheaths normal; 2(2)/3 labial tooth rows, A-2 with wide gap, tooth rows situated nearly lateral to mouth and upper jaw sheath; (23) minute body size; snout–vent length in adult males 19.3–26.8 mm (n = 13), and in adult females 22.4–31.1 mm (n = 7).

Color in life (Figure 109): Dorsum lime green with pale yellow or greenish spots. Venter transparent, pericardium usually white, visceral and hepatic peritonea white. Twomey et al. [19] report that within a single population of H. fleischmanni near San Gabriel Mixtepex (Oaxaca, Mexico), adults exhibited variation in the condition of the pericardium (from white to transparent).

Figure 110. Hyalinobatrachium fleischmanni, variation in hand webbing, ventral view. (A) KU 116447. (B) QCAZ 22301. Illustrations by Juan M. Guayasamin.

Color in ethanol: Cream dorsum with dark melanophores in the places where green coloration was in life. Venter translucent. Parietal peritoneum completely transparent, heart and all viscera covered by white lining.

Biology and ecology: The natural history of Hyalinobatrachium fleischmanni has been reviewed by several studies, including Greer and Wells [213], Villa [214], Jacobson [188], Hayes [189], Savage [148], Kubicki [24], and Delia et al. [215]. We summarize the essential information here but refer readers to those papers for more data. Individuals of Hyalinobatrachium fleischmanni have been found on vegetation up to 10 m above streams at night. Males are territorial and call from the lower (mainly) and upper surfaces of leaves. Females usually deposit eggs clutches on the lower surfaces of leaves, although some variation has been reported [215]; maternal care is absent [25]. Physical combat occurs when a male intrudes into an occupied territory. Initially the owner of the territory will call vigorously, but if that strategy fails, male–male combat starts usually with both males adopting an amplexus-like position. Both males may give quick calls. Although venter to venter combat was thought to be absent in Hyalinobatrachium, H. fleischmanni has been observed to adopt this combat position, with both males dangling upside down while holding vegetation with their hind limbs [215]. Males attend egg clutches at night, and may continue to advertise, sometimes ending with multiple clutches from different females. During the day, the male parent retreats to nearby vegetation. Only males are involved in parental care of the eggs. Delia et al. [25] observed that males of this species are attentive to individual embryo needs. Eggs are pale greenish white, and clutches are deposited on the underside of leaves. Each egg clutch has one layer of 10–50 eggs [189,216].

Call: The call consists of a single “wheet” note that rises at the end, with duration of 150–300 ms. The dominant frequency is 3800–4500 Hz at the beginning and 4800–5300 HZ at the end. Calls are emitted 4–19 per minute [148,188,209,213].

Tadpole: A description of the tadpole can be found in Starrett [146], Savage [148], and Hoffmann [147].

Distribution (Figure 111): Hyalinobatrachium fleischmanni is known from southern Mexico, across Mesoamerica, south to central Ecuador at elevations below 800 m [24,148,212,217,218]. In Ecuador, the species is known from localities in the Pacific lowlands (mostly below 300 m) in the provinces of Esmeraldas, Pichincha, Santo Domingo de los Tsáchilas, Los Ríos, and Guayas. In Ecuador, the potential distribution of the species is 39,738 km². The distribution of H. fleischmanni is an overestimation because, as currently recognized, it is a species complex [212].

Figure 111. Distribution of Hyalinobatrachium fleischmanni in Ecuador (yellow dots).

Conservation status: Globally, Hyalinobatrachium fleischmanni is considered as Least Concern by the IUCN because of its wide distribution and tolerance of habitat modifications [219]. However, recent studies suggest that this taxon represents a species complex that requires subdivision [212]. In Ecuador, about 55% of the potential distribution of the species is affected by human activities. Even though habitat destruction is considerable, H. fleischmanni is known to tolerate a degree of disturbance [24]. In Ecuador, we suggest maintaining the category of Least Concern until new taxonomic studies are available.

Evolutionary relationships (Figure 101): Our tree shows Hyalinobatrachium fleischmanni as the sister species of H. muiraquitan, which is endemic to the Brazilian Amazon basin. Although the Venezuelan H. tatayoi falls within the genetic variation of H. fleischmanni, recent studies suggest that H. fleischmanni represents a species complex that requires taxonomic revision [212].

Specimens examined: Hyalinobatrachium fleischmanni: Costa Rica: Alajuela, USNM 219249–61. San José, USNM 219263–80. Guanacaste, USNM 219282–303. Ecuador: Provincia de Los Ríos: Quevedo, USNM 60520; Río Palenque, USNM 286639–40; Patricia Pilar, USNM 286645; Hacienda Cerro Chico, USNM 286646. Provincia de Esmeraldas: 4 km W Durango, QCAZ 23549 km² in the San Francisco–Durango road, QCAZ 32073; Río Quingue, nearby Caimito (0.72096 S, 80.09117 W, 47 m), QCAZ 37308; Río Onzole, QCAZ 10433; Tesoro Escondido (0.542 N, 79.145 S, 225 m), MZUTI 3621–3625. Provincia de Santo Domingo de los Tsáchilas: Bosque Protector La Perla, QCAZ 12606. Honduras: Olancho: USNM 342162–342213. Nicaragua: Matagalpa: USNM 220013–18. Nueva Segovia: USNM 220019–36. México: Chiapas: USNM 115499.

Localities from the literatura: Ecuador: Provincia de Guayas: Cerro de Hayas (2.7299° S, 79.6297° W, 127 m), MZUA.AN.1693–1694; Provincia de Los Ríos: Macul (1.2279° S, 79.7531° W, 84m), MZUA.AN.660–661 [218].

Hyalinobatrachium iaspidiense (Ayarzagüena, 1992 [220]; Figure 112 and Figure 113).
Centrolenella iaspidiensis Ayarzagüena, 1992 [220]. Holotype: EBD 28803.
Type locality: “Quebrada Jaspe. San Ignacio de Yuruaní. Edo. Bolivar. Venezuela.”
Centrolene iaspidiensis—Duellman, 1993 [221]. Unintended combination.
Hyalinobatrachium iaspidiense—Myers and Donnelly, 1997 [104].
Hyalinobatrachium nouraguensis—Lescure and Marty, 2001 [222]. Holotype: MNHNP 1999.8604. Type locality: “Saut Arataye (environs du camp de base), Réserve des Nouragues (bassin de l’Approuague), Guyane française”. Placed in synonymy by Yánez-Muñoz, Pérez-Peña, and Cisneros-Heredia, 2009 [223].

Common names: English: Jaspe Glassfrog. Spanish: Rana de Cristal de Jaspe.

Etymology: The specific name iaspidiense is derived from the Greek word iaspis, meaning jasper, in reference to the type locality of the species, Quebrada de Jaspe.

Identification: Hyalinobatrachium iaspidiense is one of the most easily recognizable centrolenid. In life, it has a dorsum with large, lime green, irregular blotches and small, black spots (Figure 112). The only species with a similar dorsal coloration is H. mesai, a species known only from the southern slope of Sarisariñama-tepui, Venezuela [224]. It has been suggested, however, that H. mesai and H. iaspidiense represent the same species [223].

Figure 112. Hyalinobatrachium iaspidiense in life. (Left): Adult female (QCAZ 38438) from Ecuador, Yachana Reserve, 300–350 m; photo by Susan North. (Right): Adult male (SMNS 12247) from Guyana, Mabura Hill Forest Reserve, 60 m; photo by Raffael Ernst.

Diagnosis: (1) Vomerine teeth absent; (2) snout truncated in dorsal aspect and slightly protruding in profile; (3) tympanum not visible; supratympanic fold absent; (4) dorsal skin shagreen; (5) venter smooth; pair of enlarged subcloacal warts, but low and difficult to see; (6) ventral parietal peritoneum transparent (condition P0); pericardium transparent; white peritoneum covering intestines and stomach; transparent peritoneum on urinary bladder (condition V6); (7) liver bulbous, hepatic peritoneum white (condition H2); (8) humeral spines absent; (9) webbing absent between Fingers I, II, and III, moderate between outer fingers; webbing formula III (2–2⁺)—(1⁺–2⁻) IV; (10) webbing between toes moderate; webbing formula on foot I (1–1⁺)—(2⁺–2^1/3) II (1⁺–1^1/3)—(2⁺–2^1/4) III (1–1⁺)—(2–2^3/4) IV (2⁺–2^1/4)—(1⁺–1^1/4) V; (11) ulnar fold present, enameled; external tarsal fold present, enameled; internal tarsal fold short and low; (12) concealed prepollex; nuptial pad Type IV; (13) Finger II shorter than Finger I (Finger II about 93% of Finger I); (14) disc of Finger III width 39%–57% of eye diameter; (15) in life, dorsum translucent with a yellowish–green background coloration, large lime green blotches, and small black spots (Figure 112); bones white; (16) in preservative, dorsum cream with large, irregular white blotches and small, black spots; (17) in life, iris yellowish to greyish white; (18) fingers and toes lacking melanophores, except few present on Finger IV and Toes IV and V; (19) males call from the underside of leaves; the call consists of a single pulsed note, with a dominant frequency at 4440–4710 Hz; (20) fighting behavior unknown; (21) egg deposition site unknown; parental care unknown; (22) tadpole unknown; (23) minute body size; in Venezuelan specimens: SVL in females 22.7 mm (n = 1), in males 19.8–21.8 mm (n = 8) [106]; in Ecuador, male SVL = 19.9 mm (n = 1), and female SVL = 21.9 mm (n = 1).

Color in life (Figure 112): Dorsum translucent, with a yellowish–green background coloration, large lime-green blotches, and small black spots. Venter transparent, pericardium transparent (red heart visible), visceral and hepatic peritonea white [223,225].

Color in ethanol: Dorsum cream lavender, with large irregular white marks and small black spots. Ulnar and tarsal folds enameled; some of the warts below cloaca enameled. Venter transparent, pericardium transparent, visceral and hepatic peritonea white; peritonea around urinary bladder transparent; gall bladder white (description based on two adults, MECN 4033, QCAZ 38438).

Variation: Ulnar and tarsal folds less evident and with few iridophores (QCAZ 38438).

Biology and ecology: Found on vegetation 4–7 m above streams [106]. In Ecuador (Yachana Reserve), an adult female was found on a long blade of tall grassy shrub overhanging water, located in the middle of a stream (4 m wide). Stream habitat is variable, but the female was found in a shallow area (~20 cm deep) where the stream becomes a riffle and is more fast-flowing. Stream bottom here is composed of about 80% pebbles and small rocks and 20% sand near edges. The forest adjacent to the stream is near pristine and about 25–30 m high [225]. Parental care unknown.

Call: The call consists of a single note, with 16 pulses and a duration of 62.7–75.1 ms. Dominant frequency at 4440–4710 Hz. The total frequency range for the call is between 3710 and 5850 Hz, with a secondary frequency at 8600–9200 Hz. The call was emitted frequently (7–10 calls per minute) [106].

Tadpole: Not described.

Distribution (Figure 113): Hyalinobatrachium iaspidiense is known from the Guiana region of Venezuela, central Guyana, Surinam (Sipaliwini District), and French Guiana, as well as from the Amazon Basin of Ecuador, Peru, and Brazil at elevations below 1000 m [106,222,225,226,227,228,229]. In Ecuador, the species is known from few localities in the Amazonian raiforest.

Figure 113. Distribution of Hyalinobatrachium iaspidiense in Ecuador (yellow dots).

Conservation status: Hyalinobatrachium iaspidiense is considered as Data Deficient by the IUCN [230]. However, given the large distribution of the species and the absence of immediate threats, we suggest that it should be considered as Least Concern, both globally and in Ecuador.

Evolutionary relationships (Figure 101): H. iaspidiense is the sister species of H. tricolor.

Specimens examined: H. iaspidiense: Ecuador: Provincia Napo: Yachana Reserve (0°52′21.71″ S, 77°14′13.43″ W; 300–350 m), QCAZ 38438; Estación Biológica Jatun Sacha (1.066 S, 77.617 W, 405 m), QCAZ 53023. Provincia Orellana: Km 66 on the Pompella Sur–Iro road (0.8022 S, 76.398 W, 280 m), QCAZ 54947. Provincia Sucumbíos: Totoa Nai’qui (0.03442° S, 76.75278° W, ca. 280 m), MECN 4033.

Hyalinobatrachium munozorum (Lynch and Duellman, 1973 [22]; Figure 114, Figure 115 and Figure 116).
Centrolenella munozorum Lynch and Duellman, 1973 [22]. Holotype: KU 118054.
Type locality: “Santa Cecilia, 340 m, Provincia Napo (Sucumbíos), Ecuador”.
Hyalinobatrachium munozorum—Ruiz-Carranza and Lynch, 1991 [6].
Hyalinobatrachium ruedai Ruiz-Carranza and Lynch, 1998 [27]. Holotype: ICN 40409. Type locality: “(Colombia,) Departamento de Caquetá, municipio de Miraflores, Parque Nacional Natural de Chiribiquete, campamento base, 530 m”. New synonymy.

Common names: English: Muñoz’s Glassfrog. Spanish: Rana de Cristal de Muñoz.

Etymology: The specific name munozorum is a patronym for Ildefonso Muñoz B. and Blanca Muñoz, hosts of the extensive herpetological work carried out in Santa Cecilia by W. E. Duellman and his students [22].

Identification: Hyalinobatrachium munozorum can be distinguished from most glassfrogs by having a green dorsum with pale yellow spots, a completely transparent ventral parietal peritoneum, white liver and digestive tract, and by lacking humeral spines (Figure 114). Similar species with a completely transparent venter that inhabit the Amazon basin include H. bergeri, H. carlesvilai, H. iaspidiense, H. mondolffi, Chimerella mariaelenae, and Teratohyla amelie. Hyalinobatrachium munozorum differs from C. mariaelenae by having, in life, a green dorsum with pale yellow spots (dorsum green with dark grey spots in C. mariaelenae) and by lacking humeral spines (small humeral spines present in adult males of C. mariaelenae). Teratohyla amelie has a uniform green dorsum, lacking the pale-yellow spots visible in H. munozorum. Hyalinobatrachium iaspidiense, in life, has a dorsum with large, green, irregular marks and small, black spots. Morphological differentiation among H. munozorum, H. mondolfii, and H. carlesvilai is minimal, although molecular divergence is considerable [22,231,232]; the pupil of H. bergeri and H. carlesvilai is surrounded by a dark grey ring (absent in H. munozorum).

Figure 114. Hyalinobatrachium munozorum in life. Adult male from stream near Tena, 708 m, MZUTI 1616. Photos by Eduardo Toral.

Diagnosis: (1) Vomers lacking teeth; (2) snout varies from truncated to rounded in dorsal aspect and round to truncated in lateral view; (3) tympanum usually obscured by skin, small when evident, its diameter about 25% of eye diameter; supratympanic fold low; (4) dorsal skin shagreen; males lacking spicules; (5) ventral skin texture areolate; lacking pair of enlarged subcloacal warts; (6) ventral parietal peritoneum transparent, white pericardium, white peritonea covering intestines and stomach, transparent urinary bladder; (7) liver bulbous, covered by white peritoneum; (8) humeral spines absent; (9) webbing between inner fingers absent; webbing formula for outer fingers: III (1^1/2–2⁻)—(1–2⁻) IV; (10) feet about three-fourths webbed; webbing formula: I (0⁺–1⁻)—(1^1/3–2⁻) II (0⁺–1)—(1^1/2–2) III (0⁺–1^1/3)—(2⁻–2^1/3) IV (2⁻–2^1/3)—(0⁺–1) V; (11) ulnar and tarsal folds variable, from inconspicuous to pronounced; (12) concealed prepollex; in males, nuptial pad not evident; (13) Finger I slightly longer than Finger II (Finger II length 93.7%–97.6% of Finger I); (14) disc of Finger III of moderate size, its width 37.9%–56.2% of eye diameter; (15) in life, dorsum green with diffuse yellow spots, venter transparent; bones white; (16) in preservative, dorsum cream lavender with small unpigmented spots; (17) in life, iris pale to bright gold with dark punctuations; (18) dorsal surfaces of fingers and toes lacking melanophores; (19) call composed by a single note with a duration of 145–178 (mean = 170, SD = 9.9) ms; each call is mostly tonal, with a dominant frequency at 5719–5906 (mean = 5812, SD = 64) Hz; (20) fighting behavior unknown; (21) egg clutches laid on the underside of leaves; parental care unknown; (22) tadpoles unknown; (23) minute body size; in males, SVL 19.9–21.9 mm (X = 20.6 ± 0.691; n = 8); in females, SVL 20.9–23.6 mm (n = 3).

Color in life (Figure 114): Dorsal surfaces of head and body green with small, pale yellow spots and minute dark brown flecks. Parietal peritoneum transparent. White peritonea covering heart, liver, digestive tract, and testes; transparent urinary bladder. Iris pale gold with minute dark flecks. See Taxonomic Remarks.

Color in ethanol: Dorsal surfaces of head, body, and limbs cream with numerous minute lavender flecks. White visceral and hepatic peritonea. Layer of iridophores covering heart.

Variation: Specimens in the type series have a round snout in lateral view, and low ulnar and tarsal folds that are inconspicuous and unpigmented. Additional specimens from the type locality (Santa Cecilia) show the following variation: Snout truncated in lateral view, ulnar and tarsal folds conspicuous and white (KU 155493–96); conspicuous tarsal fold and low ulnar fold (KU 152488, 152489, 175215). Specimens identified as Hyalinobatrachium ruedai (a synonym of H. munozorum; see below) by Ruiz-Carranza and Lynch [27] and Cisneros-Heredia and McDiarmid [233] show the following variation: Snout truncated to rounded in dorsal and lateral views, ulnar and tarsal folds vary from inconspicuous to low and white. A female (KU 154749) has a truncated snout and conspicuous white ulnar and tarsal folds. The pericardium of all specimens from Ecuador and Colombia (including the type series) is white. In some individuals (KU 123225, 152488–89, 155494, 155496, 175504), the layer of iridophores is thinner. Cisneros-Heredia and McDiarmid [17] reported that one specimen showed a bicolored iris in life, with a light grey circumpupilary zone. These authors also reported variation in the iris coloration in preservative, from lavender–cream background with dense dark punctuations and lavender mid-line to uniform white.

Biology and ecology: The type series was found on leaves of bushes and trees at night: One over a pond, one away from water in primary forest, one on a palm frond 2 m above a stream, and one on an herbaceous leaf more than 2 m above a stream. The specimen from Lago Agrio was obtained from the foliage of a large tree that was felled during the clearing of primary forest [22]. A specimen from Tena was collected at night on the leaf of a bush at a rivulet in secondary forest [17]. Males called from riverine vegetation at a locality nearby Tena on August 2012. Parental care is unknown.

Call (Figure 115): We analyzed 18 notes from 1 individual (LBE-C-020) recorded at Río Bigal, Orellana province, Ecuador. The call is composed by a single note. Note duration is 145–178 (mean = 170, SD = 9.9) ms. Notes are mostly tonal, but they exhibit one to seven (mean = 4.5, SD = 1.6) low-amplitude peaks. Notes usually have their peak amplitude in the first 50% of the note (relative peak time: Range = 0.1699–0.661, mean = 0.448, SD = 0.123). The dominant frequency of a note measured at peak amplitude is 5719–5906 (mean = 5812, SD = 64) Hz and is contained within the fundamental frequency. The fundamental frequency has a lower limit of 5625–5812 (mean = 5708, SD = 63) Hz and a higher limit of 5812–6000 (mean = 5906, SD = 64) Hz.

Figure 115. Vocalization of Hyalinobatrachium munozorum recorded at Río Bigal, 930 m, Orellana province, Ecuador, by Morley Read (LBE-C-020).

Tadpole: Not described.

Distribution (Figure 116): Hyalinobatrachium munozorum is known from localities in the Amazonian lowlands and Andean foothills of Colombia, Ecuador, Peru, and Bolivia at elevations below 980 m ([17,22,27,134,232], this work). In Ecuador, this species has been recorded from localities below 300 m; although, there is one juvenile (identification unconfirmed), found at 920 m (Specimens examined). In Ecuador, the potential distribution of the species is 101,986 km².

Figure 116. Distribution of Hyalinobatrachium munozorum in Ecuador (yellow dots).

Conservation status: Globally, Hyalinobatrachium munozorum is listed as Least Concern by the IUCN [234]. Although H. munozorum is rare in collections, we presume that this is a consequence of inadequate sampling at the canopy level [166]. The distribution of the species is large and lacks immediate threats; thus, the Least Concern conservation status is justified.

Evolutionary relationships (Figure 101): H. mondolfii is the closest relative of H. munozorum.

Taxonomic Remarks: Lynch and Duellman [22] used the terms “heart visible” and “heart not visible” to illustrate the condition of the ventral parietal peritoneum; species that have a transparent (=clear) peritoneum were considered as having a “heart visible”, whereas species with a white parietal peritoneum had a “heart not visible”. Ruiz-Carranza and Lynch [27] discussed the character states of the pericardium; they applied the term “heart visible” for species that have a transparent parietal peritoneum as well as a transparent pericardium (red heart visible in life). In the same work, Ruiz-Carranza and Lynch [27] described H. ruedai, which was diagnosed by having a white pericardium; H. ruedai was compared with congenerics with a white heart, a list that did not include H. munozorum. We assume that Ruiz-Carranza and Lynch [27] interpreted the condition of “heart visible” for H. munozorum as equivalent to a visible red heart. As mentioned above, the type series of H. munozorum and H. ruedai have a white heart. The only other characteristic that could separate H. ruedai from H. munozorum is the hand webbing; however, the range observed in H. ruedai falls within the variation of H. munozorum. Therefore, herein we place Hyalinobatrachium ruedai Ruiz-Carranza and Lynch, 1998 [27], in the synonymy of Centrolenella munozorum Lynch and Duellman, 1973 [22].

Specimens examined: H. munozorum: Ecuador: Provincia Sucumbíos: Santa Cecilia (00°03′ N, 76°58′ W; 340 m), KU 118054 (holotype), 105251, 123225, 150620 (paratypes), 152488–89, 155493–96, 175504. Provincia Orellana: Río Yasuní (00°51′ S, 76°23′ W; ca. 250 m), KU 175215; Tiputini Biodiversity Station (00°37′ S, 76°10′ W; 190–270 m), DFCH-D105. Provincia Zamora Chinchipe: Shaime (4°20′ S, 78°40′ W; ca. 920 m), QCAZ 31056 (identification not certain). Provincia Napo: Tena (00°59′ S, 77°49′ W; 500 m), DFCH-USFQ 0735. Provincia Pastaza: Río Manderoyacu, EPN 6427; km 6 vía San Ramón-El Triunfo (1.355S, 77.86456), QCAZ 33261. Colombia: Departamento Caquetá: Municipio de Miraflores, Parque Nacional Natural Chiribiquete, 530 m, ICN 40409–11 (type series of H. ruedai), IND-AN 5448-52. Peru: Departamento de Huanuco: Finca Panguana, Río Llullapichis, 4–5 km upstream from Río Pachitea (9°36′ S, 74°55′ W; 200 m), KU 154749, 172167–69; Departamento de Cuzco: 40 km E Quince Mil on road to Puerto Maldonado, above Río Marcapata (13°09′ S, 71°25′ W), KU 197028–29.

Hyalinobatrachium pellucidum (Lynch and Duellman, 1973 [22]; Figure 117, Figure 118, Figure 119 and Figure 120).
Centrolenella pellucida Lynch and Duellman, 1973 [22]. Holotype: KU 143298.
Type locality: “Río Azuela, 1740 m, Quito–Lago Agrio road, Provincia Napo (Sucumbíos), Ecuador.”
Hyalinobatrachium pellucidum—Ruiz-Carranza and Lynch, 1991 [6].
Hyalinobatrachium lemur Duellman and Schulte, 1993. Holotype: KU 211768. Type locality: “west slope of Abra Tangarana, 7 km (by road) northeast of San Juan de Pacaysapa (06°12′ S, 76°44′ W, 1080 m), Provincia Lamas, Departamento San Martín, Perú”. Synonymy by Castroviejo-Fisher, Padial, Chaparro, Aguayo, and De la Riva, 2009 [231].

Common names: English: Andean Glassfrog. Spanish: Rana de Cristal Andina.

Etymology: The specific name pellucidum is derived from the Latin word pellucidus (clear, transparent) and refers to the transparent parietal peritoneum of this centrolenid frog [22].

Identification: Hyalinobatrachium pellucidum is distinguishable from most glassfrogs by having a completely transparent ventral parietal peritoneum, a white liver, and transparent pericardium (in life, red heart visible ventrally; Figure 117). On the Amazonian slopes of the Andes and Amazon basin, the only other species with a visible red heart in life is H. iaspidiense, which differs from H. pellucidum by having large lime green blotches and small black spots on the dorsum (both absent in H. pellucidum).

Figure 117. Hyalinobatrachium pellucidum in life. (Left and center): Male from Río Napinaza, Morona Santiago province, Ecuador, QCAZ 42000. (Right): Male from Miazi Alto, Zamora Chinchipe province, 1250 m, QCAZ 41648. Photos by Luis A. Coloma.

Diagnosis: (1) Vomerine teeth absent; (2) snout truncated in dorsal aspect and profile (Figure 118); (3) tympanum partially hidden under skin, its diameter about 30% of eye diameter; supratympanic fold low; (4) dorsal skin shagreen; (5) ventral skin texture areolate; lacking pair of enlarged subcloacal warts; (6) ventral parietal peritoneum transparent (condition P0); pericardium transparent; white peritoneum covering intestines and stomach; transparent peritoneum on urinary bladder (condition V6); (7) liver bulbous, hepatic peritoneum white (condition H2); (8) humeral spines absent; (9) webbing absent between Fingers I and II, extensive between outer fingers (Figure 118); webbing formula II 1^1/2–3⁺ III 2⁺—(2⁻–2) IV; (10) webbing between toes extensive; webbing formula on foot I 1—1^1/2 II 1⁺—2⁻ III 1⁺—2⁺ IV 2—1 V; (11) ulnar and tarsal folds present, usually white (but see Variation); (12) concealed prepollex; nuptial pad type unknown; (13) Finger I slightly longer than Finger II (Finger II about 95% of Finger I); (14) disc of Finger III width about 50% of eye diameter; (15) in life, dorsum pale green with diffuse yellow spots; venter transparent, exposing the red heart (Figure 117); color of bones white; (16) in preservative, dorsum creamy white with minute purple flecks visible under magnification; (17) iris pale silvery bronze in life; (18) dorsal surfaces of fingers and toes lacking melanophores; (19) each has a single, tonal note with a duration of 98–140 (mean = 128, SD = 8.7) ms; dominant frequency is at 5599–5857 (mean = 5690, SD = 58) Hz; (20) fighting behavior unknown; (21) egg clutches placed on the underside of leaves; maternal care absent; prolonged parental care provided by males; (22) tadpoles undescribed; (23) minute body size; in males, SVL 20.4–21.4 mm; in one female SVL 21.6 mm.

Figure 118. Hyalinobatrachium pellucidum, holotype, KU 143298. (A) Head in lateral view. (B) Head in dorsal view. (C) Hand in ventral view. Illustrations by Juan M. Guayasamin.

Color in life (Figure 117): Dorsum pale green with diffuse yellow spots; venter and hidden surfaces of limbs lacking pigment; fingers and toes yellow; parietal peritoneum clear; red heart visible; bones white; iris pale silvery bronze [22].

Color in ethanol: Dorsum creamy white with minute purple flecks and small white granules visible under magnification; digestive tract and liver covered by white peritonea; pericardium lacking iridophores; ulnar and tarsal folds white.

Variation: Castroviejo-Fisher et al. [231] reported the following variation: The holotype of H. pellucidum has marked and enameled ulnar, tarsal, and cloacal folds, while specimens previously identified as H. lemur (a synonym of H. pellucidum) only show weak and non-enameled folds. The variation is interpreted as being the product of preservation artifacts and intraspecific variation. Two additional specimens from the same locality and near the type locality of H. lemur (~45 Km straight line) show intermediate states regarding ulnar, tarsal, and cloacal folds. The specimen KU 217297 has an enameled but weak ulnar fold, a weak but non-enameled tarsal fold, and an enameled and weak cloacal fold; KU 217295 has an enameled and marked ulnar fold, an enameled but weak tarsal fold, and an enameled and marked cloacal fold. A specimen (MHNCP 4880) collected in Cusco, Peru, and assigned to H. pellucidum, has very weak and barely enameled ulnar, tarsal, and cloacal folds.

Biology and ecology: The holotype of Hyalinobatrachium pellucidum was found on the leaf of an herb over a small stream at night. The following species were in the same stream and in other small streams nearby: Nymphargus megacheirus, N. anomalus, N. siren, Centrolene pipilata, Hyloscirtus phyllognathus [22]. Short-term maternal care is absent; males provide prolonged parental care [25].

Call (Figure 119). We analyzed 36 notes from one individual (USNM 286708), recorded at the type locality of the species, Río Azuela, 1740 m, Napo province, Ecuador, by Roy McDiarmid. The advertisement call is relatively short which has a single note per call (i.e., call = note). Notes are moderate in duration and the note duration is 98–140 (mean = 128, SD = 8.7) ms. Notes are tonal and do not show any clear amplitude peaks. The dominant frequency is 5599–5857 (mean = 5690, SD = 58) Hz and is contained within the fundamental frequency. The fundamental frequency has a lower limit of 5163–5685 (mean = 5381, SD = 678) Hz and a higher limit of 5685–5943 (mean = 5778, SD = 56) Hz.

Figure 119. Call of Hyalinobatrachium pellucidum recorded at Río Azuela, 1740 m, Napo province, Ecuador, recorded by Roy McDiarmid (USNM 286708).

Tadpole: Not described.

Distribution (Figure 120): Hyalinobatrachium pellucidum is known from a few records in the lower montane rainforest on the Amazonian Andean slopes of Ecuador and Peru, at elevations between 1000–1740 m ([22,231], this work). In Ecuador, Hyalinobatrachium pellucidum inhabits the Eastern Foothill Forest and Eastern Montane Forest ecoregions, and it is known from localities between 1013 and 1740 m; these localities are Río Azuela, Río Reventador, km 6.6 on the Limón–Macas road, and Cordillera del Cóndor (Specimens Examined).

Figure 120. Distribution of Hyalinobatrachium pellucidum in Ecuador (yellow dots).

Conservation status: Globally, Hyalinobatrachium pellucidum is listed as Near Threatened by the IUCN [235]. In Ecuador, the species is severely fragmented because of agriculture, pasture lands, and mining. Thus, we suggest that, locally, it should be placed in the Vulnerable category, following IUCN criteria b1, B2a, B2biii.

Evolutionary relationships (Figure 101): Hyalinobatrachium pellucidum is sister to H. yaku.

Specimens examined: Hyalinobatrachium pellucidum: Ecuador: Provincia de Sucumbíos: Río Azuela (0.1167 S, 77.6167 W; 1740 m), Quito–Lago Agrio road; KU 164691 (holotype), USNM 286708–10; Río Reventador, USNM 286711–12. Provincia de Morona Santiago: km 6.6 on the Limón-Macas road (ca. 2.92816 S, 78.344 W; 1013 m), QCAZ 29438. Provincia de Zamora Chinchipe: Cordillera del Cóndor, Miazi Alto (4.25044 S, 78.61356 W; 1282 m), QCAZ 41560–61.

Hyalinobatrachium esmeralda: Colombia: Departamento de Boyacá, Municipio de Pajarito, Inspección Policía Corinto, finca ‘El Descanso’, quebrada ‘La Limonita’, 1600–1650 m, ICN 9592–94, 9596, 9602–03 (type series of H. esmeralda).

Hyalinobatrachium valerioi (Dunn, 1931 [236]; Figure 121, Figure 122 and Figure 123).
Centrolene valerioi Dunn, 1931 [236]. Holotype: MCZ 16003.
Type locality: “La Palma, Costa Rica, 4500 feet”.
Cochranella valerioi—Taylor, 1951.
Cochranella reticulata—Taylor, 1958 [201]. Holotype: KU 32922. Type locality: “near bridge across Río Reventazón at the Inter-American Institute of Agriculture, Turrialba, Cartago Province, Costa Rica”. Placed in synonymy by Starrett and Savage, 1973 [209].
Centrolenella valerioi—Starrett and Savage, 1973 [209].
Hyalinobatrachium valerioi—Ruiz-Carranza and Lynch, 1991 [6].

Common names: English: Reticulated glassfrog [237], Valerio’s glassfrog. Spanish: Rana de Cristal Reticulada, Rana de Cristal de Valerio.

Etymology: The specific name valerioi is a patronym for Manuel Valerio, who, with Emmett R. Dunn, discovered the species in the field.

Identification: Ecuadorian population tentatively assigned to Hyalinobatrachium valerioi can be distinguished from all other glassfrogs by its green dorsum with numerous large yellowish–green spots (Figure 121), transparent ventral parietal peritoneum, and most visceral peritonea covered by iridophores. The pericardium varies from white to mostly transparent (red in life). Among Ecuadorian centrolenid, only H. aureoguttatum is similar, but differs by having large yellow to golden dorsal spots (Figure 102), which are produced by the interaction of xanthophores (yellow) and iridophores (white); the spots in H. valerioi are structurally different since they lack iridophores.

Diagnosis: (1) Vomerine teeth absent; (2) snout truncated in dorsal view, truncated to protruding in lateral view; (3) tympanum with a dorsolateral orientation, of moderate size (tympanum diameter 30%–43% of eye diameter); tympanic membrane evident; only lower border of tympanic annulus visible; (4) dorsal skin finely shagreen; (5) ventral skin granular; no cloacal ornamentation; (6) parietal peritoneum transparent (condition P0), iridophores covering all visceral peritonea; pericardium variable, from white to mostly translucent (condition V5–V6); (7) liver bulbous, covered by white peritoneum (condition H2); (8) humeral spine absent; (9) webbing between Fingers I and II reduced, moderate between Fingers II and III, expanded between outer fingers, webbing formula: I (2⁺–2^1/3)—2 II (1–1⁺)—(3⁻–3^1/3) III (1^1/2–2^2/3)—(1⁺–2⁻) IV; (10) webbing on feet: I (1–1^1/2)—(2–2⁺) II (1–1^2/3)—(2–2^1/4) III (1⁻–1^1/3)—(2⁺–2^1/3) IV (2–2^1/2)—(1–1⁺) V; (11) ulnar and tarsal fold absent; (12) nuptial excrescences Type V in adult males; concealed prepollex; (13) first finger slightly longer than second; (14) eye diameter larger than width of disc on Finger III (disc of Finger III 32%–36% of eye diameter); (15) in life, dorsal surfaces yellowish green with green reticulum (or green with numerous yellowish–green spots); ventral parietal peritoneum transparent; bones white; (16) in preservative, dorsal surfaces cream with dark melanophores; (17) in life, iris golden with dark pigmentation scattered throughout, especially around the pupil; (18) melanophores absent from fingers and toes, except for few on Toes IV and V; (19) males usually call from underside of leaves, but sometimes they call from the upper leaf surface; call is a short seet, with a duration of 200–250 milliseconds, and a modulated frequency; (20) amplexus-like fighting behavior; (21) egg clutches laid on underside of leaves; nocturnal and diurnal parental care by males; maternal care absent; (22) tadpole with ventral mouth, spiracle posterior and mid-lateral; oral disc complete, moderate, with single row of marginal papillae laterally and ventrally, large dorsal gap; tooth row formula 2(2)/3, A-2 with large gap above mouth; (23) minute body size; SVL in adult males 18.1–24.4 mm (n = 23), in adult females 20.2–25.1 mm (n = 7).

Figure 121. Hyalinobatrachium valerioi in life. (Top row): Male from Reserva de Biodiversidad Mashpi, 1080 m, Ecuador; photos by Lucas Bustamante/Tropical Herping. (Bottom row): Male guarding eggs, Selva Verde Lodge, Costa Rica; photo by Carlos Martínez. Note differences in coloration.

Color in life (Figure 121): According to Kubicki [24], dorsum yellowish green with green reticulation. Transparent venter; pericardium varies from white to almost transparent; white liver and digestive tract. Iris golden with dark pigmentation scattered throughout, especially surrounding pupil; several individuals with dark pigmentation restricted to lateral regions of pupil. Bones white.

Color in ethanol: Cream dorsum with dark melanophores in the places that where green in life. Venter translucent cream. Parietal peritoneum completely transparent, all visceral peritonea covered by white lining. Pericardium polymorphic and white or translucent.

Biology and ecology: The following information was obtained from Savage [42], McDiarmid and Adler [238], McDiarmid [216], and Hayes [189]. A nocturnal frog that inhabits forested streams. Males are territorial and vigorously call from under leaves where females deposit their egg clutches. Physical combat occurs between males if another male intrudes into a male’s territory despite the calls of the owner; both males may squeak during the fight; once one of the males is pinned venter down, he is held for a while, then leaves. Pale greenish–white eggs (25–40 in number) are deposited in a single-layer on the underside of a leaf. Males continue to advertise and may end up with as many as seven clutches from different females. The males attend the eggs both during the day and at night. At night, the male continues to call and, occasionally exhibits hydric brooding behavior, in which he apparently empties his bladder over the eggs. Only males are involved in parental care of the eggs. McDiarmid [4] attributed the higher survivorship to hatching in this species than in the syntopic Hyalinobatrachium colymbiphyllum to the diurnal parental care. Vockenhuber et al. [239] experimentally verified the positive effect that parental care has on embryonic survivorship; also, they identified arthropod predation as the main cause for embryonic mortality.

Call (Figure 122): Males usually call from the underside of leaves, but sometimes they vocalize from the upper leaf surface. In Costa Rica, the call is a high-pitched seet lasting 200 to 250 milliseconds, with a dominant frequency initially of 7.0 kHz rising to 7.5 kHz and then dropping again to 7.2 kHz, repeated every 7 to 10 s [209]. Kubicki [24] reported call variation among Costa Rican populations. Below, we describe a call of an individual tentatively assigned to Hyalinobatrachium valerioi (USNM 201475) recorded near Santo Domingo de los Colorados by RWM; we analyzed 19 notes from one individual (Figure 122). Each note is composed by a single note. The note duration is 46–78 (mean = 72, SD = 7.7) ms. Notes are generally tonal and do not show any clear amplitude peaks throughout the note. The dominant frequency at peak amplitude is 5771–6632 (mean = 6197, SD = 253) Hz and is contained within the fundamental frequency. The fundamental frequency has a lower limit of 5599–5943 (mean = 5771, SD = 91) Hz and a higher limit of 6374–6718 (mean = 6632, SD = 76) Hz.

Figure 122. Call of a male (USNM 201475) tentatively assigned to Hyalinobatrachium valerioi, recorded near Santo Domingo de los Colorados, Ecuador, recorded by RWM.

Egg masses and tadpoles: The following information was obtained from Savage (2002) and incorporates data from McDiarmid and Adler [238], McDiarmid [216], and Hayes [189]. Green egg masses are deposited in a single-layer jelly mass on the underside of leaves; average clutch size is 35, with 40 eggs being the upper limit. Egg masses are attacked by diurnal wasps, which remove eggs one at a time and carry them away, presumably to a nest, until the egg mass is depleted [4]. Starrett [146] described the tadpole of H. valerioi as follows (as Cochranella reticulata): Tadpole similar to that of H. fleischmanni, but tail longer at hatching and color paler; tail of older tadpole two- and three-fifths times as long as body; dorsal part of body with brown blotches; tail musculature with brown blotches dorsally and laterally; pigment is in depressions between myotomes posteriorly; tail fins clear. Mouth ventral, nearly terminal; papillae surrounding all but anterior portion of the disc. LTRF 2(2)/3; A-1 complete, A-2 rows very short, occurring only lateral to the jaw sheaths; outermost posterior row not well developed at this stage (probably, in larger tadpoles, nearly as long as inner two); anterior jaw sheath is more heavily pigmented than the posterior one that is doubly arched and weakly pigmented medially, often appearing broken; posterior jaw sheath armed with small, equal length serrations. Hoffmann [147] recently provided a very detailed description of the tadpole of the species. He mentioned that tadpoles of H. valerioi are distinguished by their accented slenderness and by the noticeably longer tail, about three times longer than the body; additionally, the oral disc of H. valerioi is more ventrally positioned than in other glassfrog species [147].

Distribution (Figure 123): As currently recognized, Hyalinobatrachium valerioi occurs from Costa Rica to southwestern Ecuador [156]. In Ecuador, it occurs in the northern and southern Pacific lowlands, foothills, and slopes below 1500 m in the provinces of Azuay, Carchi, Esmeraldas, Los Ríos, Pichincha, and Santo Domingo de los Tsáchilas (Specimens examined), within the Western Foothill Forest and Western Montane Forest ecoregions. In Ecuador, it has a potential distribution of 49,705 km².

Figure 123. Distribution of populations tentatively assigned to Hyalinobatrachium valerioi in Ecuador (yellow dots).

Conservation status: Globally, Hyalinobatrachium valerioi is categorized as Least Concern by the IUCN [240]. The Ecuadorian populations currently assigned to H. valerioi (see Figure 123) might represent an undescribed species. The habitat of the species is severely fragmented, and the species has exhibited populations declines in several historical localities (e.g., La Florida, Río Palenque, Manta Real; Luis A. Coloma, pers. comm.; JMG, pers. obs.). During the last year (2015–2019), we have found reproducing populations at Reserva Los Cedros and Reserva Mashpi. Based on the current available information, we suggest that H. valerioi should be considered as Endangered in Ecuador (IUCN criteria A2c, A2e).

Evolutionary relationships (Figure 101): Hyalinobatrachium valerioi is the sister species to H. aureoguttatum.

Taxonomic remarks: The coloration of Hyalinobatrachium valerioi is variable across its distribution. Population may differ in having the pericardium completely transparent or covered with iridophores [24]. Also, at least in Costa Rica, there is notorious call variation [24]. At the moment, we are still uncertain about the taxonomic position of the populations from Ecuador; further studies are needed to assess if H. valerioi, as currently defined, is composed by several morphologically similar species.

Specimens examined: Hyalinobatrachium valerioi: Ecuador: Provincia de Azuay: 12.9 km W of Luz María, KU 217503–04. Provincia de Carchi: Maldonado, KU 178082–91. Provincia de Cañar: Manta Real (03°10′ S 80°26′ W), DHMECN 0134. Provincia de Imbabura: Reserva Los Cedros (0.3026° N, 78.781° W; 1360 m), MZUTI 3268. Provincia de Los Ríos: Río Palenque (0.55° S, 79.36667° W), USNM 286746–49, KU 147580, 164650. Provincia de Santo Domingo de los Tsáchilas: 5 km W of La Florida (0.25694° S, 79.0538° W; 860 m), QCAZ 12600, 27652; 4 km NE of Dos Ríos (0.25694° S, 79.0538° W), KU 164651–56; Río Faisanes (0.26083° S, 78.845° W). Provincia de Pichincha: Tandapi (0.41638° S, 78.7988° W), KU 178092; 1 km E of Pedro Vicente Maldonado (0.833° S, 79.0347° W; 670 m), QCAZ 12605; Reserva de Biodiversidad Mashpi (0.164° N, 78.867° W; 1080 m), MUTI 3921.

Localities from the literature: Hyalinobatrachium valerioi: Ecuador: Provincia de Esmeraldas: Bilsa Biological Station [160].

Hyalinobatrachium yaku Guayasamin, Cisneros-Heredia, Maynard, Lynch, Culebras, Hamilton 2017 [241] (Figure 124, Figure 125 and Figure 126).
Hyalinobatrachium yaku Guayasamin, Cisneros-Heredia, Maynard, Lynch, Culebras, Hamilton, 2017 [241]. Holotype: MZUTI 5001.
Type locality: “Stream affluent of the Kallana river (1.4696° S, 77.2784° W, 325 m), nearby the Kichwa community of Kallana, province of Pastaza, Ecuador”.

Common names: English: Yaku Glassfrog. Spanish: Rana de Cristal Yaku.

Etymology: The epithet yaku is the Kichwa word for water [241].

Identification: In life, Hyalinobatrachium yaku is distinguishable from all other glassfrogs by having middorsal dark green spots on the anterior half of the body and, ventrally, a completely transparent peritoneum and pericaridum (Figure 124).

Diagnosis: (1) Dentigerous process of the vomer lacking teeth; (2) snout truncated in dorsal and lateral views; (3) lower half of tympanic annulus visible; tympanic membrane clearly differentiated and with coloration similar to that of surrounding skin; (4) dorsal skin shagreen; (5) ventral skin areolate; cloacal area glandular, with tubercular and slightly enameled patch on each side of cloaca, lacking paired round subcloacal warts; (6) parietal peritoneum, pericardium, kidneys, and urinary bladder transparent (lacking iridophores); hepatic, gastrointestinal, and testicular peritonea covered by iridophores; (7) liver white, bulbous; (8) humeral spines absent; (9) basal webbing between Fingers I and II, moderate webbing between external fingers; hand webbing formula: I 2—2 II 0⁺—3⁺ III 2⁻—(1–2⁻) IV; (10) foot webbing moderate; webbing formula: I (1–1⁺)—(2–2⁻) II (0⁺–1)—(2⁺–2^1/3) III 1—2^1/3 IV 2^1/3—(1–1^1/3) V; (11) fingers and toes with thin lateral fringes; ulnar and tarsal folds present, but difficult to distinguish, with thin layer of iridophores that extends to ventrolateral edge of Finger IV and Toe V; (12) nuptial excrescence present as a small pad on Finger I (Type V), concealed prepollex; (13) when appressed, Finger I longer than Finger II; (14) diameter of eye about 2.1 times wider than disc on Finger III; (15) coloration in life: Dorsal surfaces apple green to yellowish green with small yellow spots and minute grey to black melanophores; posterior head and anterior half of the body with few small, well-defined dark green spots placed mid-dorsally; bones white; (16) coloration in preservative: Dorsal surfaces pale cream with minute lavender to black melanophores; (17) iris coloration in life: Silver to yellow, with minute dark spots concentrated around pupil; (18) melanophores present on Finger IV and Toes IV–V, absent on other fingers and toes; in life, hands and feet are cream with a light green hue, with tips of fingers and toes being yellowish green; (19) males call from the undersides of leaves; advertisement call consisting of a single tonal note; call duration note 0.3–0.4 s, dominant frequency 5219–5330 Hz, with no frequency modulation; (20, 21) clutch size unknown; maternal care unknown; prolonged parental care provided by males; (22) tadpoles unknown; (23) adults small; SVL in adult males 20.8–22.3 mm (n = 3), in adult female 21.1 mm (n = 1).

Figure 124. Hyalinobatrachium yaku in life. (Top row): adult male, MZUTI 5001, holotype, in dorsal and ventral view. (Bottom row): adult male, paratype, QCAZ 55628. Obtained from Guayasamin et al. [241].

Color in life (Figure 124): In adults, dorsum apple green to yellowish green with small yellow spots and minute dark melanophores; posterior head and anterior half of the body with few small, well-defined dark green spots placed mid-dorsally, the anterior-most spot generally being the largest. Hands and feet cream with a light green hue, with tips of fingers and toes being yellowish green; melanophores absent from fingers and toes, except Finger IV and Toes IV and V. Ventrally, parietal peritoneum and pericardium transparent (red heart fully visible); visceral peritoneum of gall bladder and urinary bladder transparent; hepatic and visceral peritonea white. Ventral vein red. Iris silver to yellow, with minute dark spots that encircle the pupil, giving the impression of diffuse rings. Bones white [241].

Color in ethanol: Dorsal surfaces cream dotted with minute dark lavender to black melanophores; venter uniform white; peritonea as in life. Iris white with lavender melanophores that become more numerous near the pupil. There are no traces of the characteristic middorsal dark-green spots in preserved specimens [241].

Variation: Juveniles have the same color pattern as adults; the number and extent of the middorsal green dots varies, but they are usually smaller and less pronounced posteriorly [241].

Biology and ecology: Only basic biological information of H. yaku is known. The data presented below are from Guayasamin et al. [241]. Males call from the underside of leaves of riverine vegetation. One male (holotype) was on the same leaf as two egg clutches, approximately 3 m above the stream. Observations suggest that prolonged parental care is provided by males. Syntopic species at the type locality are: Nymphargus mariae, Teratohyla midas, Agalychnis hulli, Callimedusa tomopterna, Boana calcarata, B. geographica, Osteocephalus fuscifacies, Pristimantis enigmaticus, and P. peruvianus. At the locality of Ahuano, the species was found along a small stream, tributary of the Arajuno River; the stream was slow flowing, very narrow (approximately 1 m wide), shallow (approximately 40 cm deep), and covered by secondary forest. At Ahuano, syntopic species were Teratohyla midas and H. ruedai. Individuals from San José de Payamino were found perched on leaves of small shrubs, ferns, and grasses (30–150 cm above ground) in disturbed secondary forest. Additionally, all individuals recorded at San José de Payamino were found >30 m from any stream. Syntopic species at San José de Payamino are reported by Maynard et al. [242].

Call (Figure 125): The information provided below is taken from Guayasamin et al. [241]. The advertisement call of Hyalinobatrachium yaku is a single and high-pitched tonal note, with no frequency nor amplitude modulation. The call lasts 0.27–0.4 s (0.3 ± 0.03) and has an average call rate of 9.0 calls/minute. Time between calls varied from 5.3–8.9 s (7.1 ± 1.1). The dominant frequency ranges from 5219–5330 Hz (5284 ± 35.0).

Figure 125. Call of Hyalinobatrachium yaku from Kallana, 325 m, Pastaza province, Ecuador; holotype, MZUTI 5001.

Tadpole: Not described.

Distribution (Figure 126): Hyalinobatrachium yaku is known from three localities on the Amazonian lowlands of Ecuador at elevations between 300–360 m. It is likely that H. yaku has a broader distribution, including areas in Amazonian Colombia and Peru [241].

Figure 126. Distribution of Hyalinobatrachium yaku in Ecuador (yellow dots).

Conservation status: According to Guayasamin et al. [241], available information is insufficient to suggest a conservation category, thus H. yaku is a Data Deficient species.

Evolutionary relationships (Figure 101): H. yaku and H. pellucidum are sister species.

Specimens examined: Hyalinobatrachium yaku: Ecuador: Provincia Pastaza: stream affluent of Kallana river (1.4696° S, 77.2784° W, 325 m), MZUTI 5001 (holotype), 5002 (paratypes). Provincia Orellana: Timburi-Cocha Research Station (0.4800° S, 77.2829° W, 300 m) near San José de Payamino, QCAZ 55628, 53352 (paratypes). Provincia Napo: Ahuano (1.0632° S, 77.5265° W, 360 m), ZSFQ 2322.

Genus Nymphargus Cisneros-Heredia and McDiarmid 2007 [17].
Etymology: The name Nymphargus is formed from the Greek nymphae in allusion to the nymphs, beautiful goddesses in Greek mythology that personify the creative and fostering activities of nature, living in mountains, valley, springs, and rivers; and, argus in allusion to the mythological Greek Argus, nephew of the nymph Io, a giant with a hundred eyes, whose eyes became the ocelli in the peacock’s tail. The name is masculine and alludes to the ocelli found on the dorsum of some of the species of the genus [17].
Nymphargus anomalus (Lynch and Duellman, 1973 [22]; Figure 127, Figure 128, Figure 129 and Figure 130).
Centrolenella anomala Lynch and Duellman, 1973 [22]. Holotype: KU 143299.
Type locality: “Río Azuela, 1740 m, Quito–Lago Agrio road, Provincia Napo, Ecuador”.
Cochranella anomala—Ruiz-Carranza and Lynch, 1991 [6], 57:21.
Nymphargus anomalus—Cisneros-Heredia and McDiarmid, 2007 [17].

Common names: English: Anomalous Glassfrog. Spanish: Rana de Cristal Anómala.

Etymology: The specific name anomalous refers to the unusual dorsal coloration of this species [22].

Identification: Nymphargus anomalus is unique by having, in life, a tan dorsum with black ocelli with orangish–tan centers (Figure 127 and Figure 128), instead of the usual green that characterizes most glassfrogs. Among centrolenids, only one species, N. ignotus, has a similar dorsal color pattern; however, N. ignotus is found on the Pacific slopes of the Andes, whereas N. anomalus is restricted to the Amazonian slopes of the Andes. Nymphargus anomalus further differs from other species having ocellated patterns (N. cochranae, N. ignotus, N. laurae) by having scattered black and lavender flecks between ocelli (Figure 127).

Diagnosis: (1) Vomers lacking teeth; (2) snout truncated in dorsal and lateral profiles (Figure 129); (3) tympanum relatively small, oriented almost vertically, with evident lateral and posterior inclinations, its diameter about 20% of eye diameter; tympanic annulus mostly visible, with supratympanic fold covering its dorsal margin; tympanic membrane differentiated and translucent, partially pigmented; (4) in males, dorsal skin shagreen with or without minute spiculae; in females, dorsal skin shagreen without spiculae; (5) venter areolate; pair of enlarged subcloacal warts (Figure 15); (6) anterior half of ventral parietal peritoneum white, posterior half transparent (condition P2); white pericardium; translucent peritoneum covering intestines (condition V1); (7) liver with four clearly defined lobes covered by transparent peritoneum (condition H0); (8) humeral spines absent; (9) webbing absent between Fingers I–III, basal or moderate between Fingers III and IV; webbing formula: III (2^1/3–3⁺)—(2⁺–2^2/3) IV (Figure 129); (10) webbing between toes moderate; webbing formula on foot I 2⁻—(2–2^1/4) II (1–1^1/4)—(2⁺–2^1/3) III (1–1⁺)—(2⁺–2^1/4) IV (2⁺–2^2/3)—(1⁺–1^1/2) V; (11) ulnar fold low; inner and outer tarsal folds present, but low and difficult to distinguish; (12) concealed prepollex; nuptial pad Type I; (13) Finger I slightly shorter than Finger II (Finger I length 91.5%–99.2% of Finger II); (14) disc width of Finger III about 50% of eye diameter; (15) in life, dorsum pale brown or tan with dark brown flecks and black ocelli with brownish–orange centers (Figure 127); color of bones white; (16) in preservative, dorsum cream to tan with black dots and lavender flecks and black and lavender ocelli enclosing whitish centers; (17) iris pale yellow with thin black reticulation; bright yellow circumpupilar ring; (18) dorsal surfaces of all fingers and toes usually with few melanophores, except Finger I that usually lacks melanophores; (19) males call from the upper surfaces of leaves; calls undescribed; (20) fighting behavior unknown; (21) eggs deposited on mossy branches over streams; parental care unknown; (22) tadpoles unknown; (23) small body size; SVL in males 21.2–24.8 mm (X = 23.1 mm; n = 16); SVL in three adult females: 25.7, 27.0, and 27.0 mm.

Figure 127. Nymphargus anomalus in life, ZSFQ 2123, from a tributary of Río San Jacinto, 1672 m, Tungurahua province. Photos by Jose Vieira/Tropical Herping.

Figure 128. Ontogenetic color change in Nymphargus anomalus, QCAZ 48107. Photos by Luis A. Coloma.

Figure 129. Nymphargus anomalus, adult male, KU 143299. (A) Head in lateral view. (B) Hand in ventral view. Illustrations by Juan M. Guayasamin.

Color in life (Figure 127 and Figure 128): Dorsum pale brown or tan with slight yellow to pink hue; dorsal surfaces of head, body, and limbs with small flecks and black ocelli that enclose orange spots. Anterior part of venter white, posterior half of venter translucent. Iris pale yellow with black reticulations; color of iris becomes whitish towards margins. Nymphargus anomalus exhibits ontogenetic variation in its dorsal coloration, being green in postmetamorphs and then gradually acquiring the tan to pale brown coloration that is characteristic of adults. The spotted pattern also changes ontogenetically; postmetamorphs have yellow spots that progressively change towards ocelli composed of black rings with orange centers.

Color in ethanol: Dorsum cream to tan with lavender flecks and black and lavender ocelli enclosing whitish centers. Eyelids bear some iridophore aggregations visible through the skin. White pericardium; hepatic and visceral peritonea are translucent. Peritonea covering urinary and gall bladders, kidneys, and testes without iridophores.

Biology and ecology: The holotype of Nymphargus anomalus was found on a mossy limb of a bush about 1.5 m above a cascading rivulet at night. The following species were found in the same stream and in other small streams nearby: Hyloscirtus phyllognathus, Centrolene pipilata, Nymphargus megacheirus, N. siren, and Hyalinobatrachium pellucidum [22]. At Río Yana Challuwa Yaku, males where found calling on leaves and branches during the night. Two egg clutches were on a mossy branch above the stream. Parental care is unknown.

Call: Not described.

Tadpole: Not described.

Distribution (Figure 130): Nymphargus anomalus is known only from four localities (Volcán Sumaco, Río Azuela, Río Yana Challuwa Yaku, nearby Río Jacinto) on the Amazonian slopes of the Ecuadorian Andes, at elevations between 1668–1795 m. The habitat of the species is within the Eastern Montane Forest region ([22], this work).

Conservation status: Nymphargus anomalus is listed as Critically Endangered by the IUCN [243]. Until recently, the species was known from a single individual collected on 23 October 1971 (Río Azuela). Surveys at the type locality have failed in find this species ([91], RWM and JMG, pers. obs.), but other populations have been discovered (Figure 130; Specimens examined). In Ecuador, because of its restricted distribution and habitat fragmentation, we suggest that it should be considered as Endangered (IUCN criteria B1, B2a, B2biii).

Figure 130. Distribution of Nymphargus anomalus in Ecuador (yellow dots).

Evolutionary relationships (Figure 101): Lynch [244] suggested that N. anomalus is the sister species of N. ignotus. The hypothesis by Lynch is based on the presence of two putative synapomorphies: Pale brown coloration and small ocelli on elevated warts, with the corresponding plesiomorphic conditions being green coloration and no ocelli. Based on mitochondrial genes and a taxon sampling that does not include N. ignotus, we found that the sister species of N. anomalus is N. megacheirus, which has a coloration that matches the plesiomorphic states.

Specimens examined: Nymphargus anomalus: Ecuador: Provincia de Napo: Río Azuela (0.11667 S, 77.6167 W, 1740 m), KU 143299. Parque Nacional Sumaco, Volcán Sumaco, near the Pavayacu refuge (0.61497 S; 77.59065 W; 1771 m), QCAZ 41312–13. Provincia de Pastaza: Reserva Comunitaria Ankaku, zona de amortiguamiento del Parque Nacional Llanganates, cabecera del Río Yana Challuwa Yaku (01.26764 S; 78.04797 W; 1668 m), males: QCAZ 45696–97, 45699, 45701, 45703–11, 45728–29; females: QCAZ 45698, 45700, 45702. Provincia de Tungurahua: stream tributary of the San Jacinto River (1.3447 S, 78.1814 W; 1795 m asl), ZSFQ 899.

Nymphargus balionotus (Duellman, 1981 [120]; Figure 131, Figure 132 and Figure 133).
Centrolenella balionota Duellman, 1981 [120]. Holotype: KU 164702.
Type locality: “3.5 km (by road) northeast of Mindo, 1540 m, Provincia de Pichincha, Ecuador (00°01′ S, 78°44′ W)”.
Cochranella balionota—Ruiz-Carranza and Lynch, 1991 [6].
Centrolene balionotum—Cisneros-Heredia and McDiarmid, 2006 [17].
“Cochranella” balionota—Guayasamin, Castroviejo-Fisher, Trueb, Ayarzagüena, Rada, Vilà, 2009 [1].
Nymphargus balionotus—Guayasamin, Cisneros-Heredia, Vieira, Kohn, Gavilanes, Lynch, Hamilton, Maynard, 2019 [21].

Common names: English: Mindo Glassfrog. Spanish: Rana de Cristal de Mindo.

Etymology: The specific epithet balionotus combines the Greek words balios (spotted, dappled) and notos (back), and is used in allusion to the dorsal color pattern of the species [120].

Identification: Nymphargus balionotus is unique by having, in life, a pale green dorsum with reddish–brown dorsolateral stripes, small reddish–brown spots, and larger yellow spots (Figure 131). Additionally, males are small (SVL < 23 mm) and possess a blade-like ventral crest on the humerus that can be confused with a humeral spine. See Remarks.

Figure 131. Nymphargus balionotus in life, ZSFQ 533. Ecuador, Reserva Río Manduriacu, 1254 m, Tungurahua province. Photos by Jose Vieira/Tropical Herping.

Diagnosis: (1) Vomers lacking teeth; (2) snout truncated in dorsal view, protruding lateral profile (Figure 132); (3) tympanum relatively small, oriented almost vertically, with evident lateral and posterior inclinations, its diameter 23%–30% of eye diameter; tympanic annulus mostly visible, with supratympanic fold covering its dorsal margin; tympanic membrane differentiated and translucent, not pigmented; (4) in males, dorsal skin smooth to shagreen, lacking spicules; females unknown; (5) venter slightly granular; pair of enlarged subcloacal warts; (6) anterior half of ventral parietal peritoneum with iridophores, posterior half transparent (condition P2); pericardium covered by iridophores; peritoneum covering digestive tract lacking iridophores (condition V1); (7) liver with four or five lobes, lacking iridophores (condition H0); (8) males with a bladelike ventral crest on the humerus that resembles a small humeral spine; (9) webbing absent between inner fingers; webbing formulae for outer fingers IV (2–2^1/2)—(2⁻–2) V (Figure 132); (10) webbing between toes moderate; webbing formula on foot: I (1^1/3–1^1/2)—(2–2⁺) II (1⁺–1^1/2)—(1–1^1/2) III (1^1/3–2)—(2⁻–3) IV (2⁻–3)—1^1/2 V; (11) ulnar fold low; inner and outer tarsal folds present, but low and difficult to distinguish; (12) concealed prepollex; nuptial pad in males variable, ranging from Type I to Type III; (13) Finger I slightly longer than Finger II (Finger II length 91.8%–97.4% Finger I); (14) disc of Finger III width about 36%–44% of eye diameter; (15) in life, dorsum pale green with reddish–brown dorsolateral stripes, small reddish–brown spots, and larger yellow spots (Figure 131); color of bones pale green; (16) in preservative, dorsum cream with lavender and white dorsolateral stripes, small dark lavender spots, and larger white spots; (17) iris greyish white with numerous minute brown spots; (18) dorsal surfaces of fingers and toes lacking melanophores; (19) males call from the upper sides of leaves overhanging streams; call undescribed; (20) fighting behavior unknown; (21) egg deposition site unknown; parental care unknown; (22) tadpoles unknown; (23) minute body size; in adult males, SVL 20.1–21.8 mm (X = 20.8 ± 0.534, n = 11); females unknown.

Figure 132. Nymphargus balionotus, adult male, KU 164708. (A) Head in lateral view. (B) Hand in ventral view. Illustrations Juan M. Guayasamin.

Color in life (Figure 131): Pale green dorsum with reddish–brown dorsolateral stripes, small reddish–brown spots, and elevated larger yellow spots. Head with reddish–brown interorbital bar and postorbital stripe that is continuous with dorsolateral stripe. Large, elevated yellow spot on anteromedial edge of each eyelid. Pale green bones ([120], this work).

Color in ethanol: Dorsal surfaces cream with lavender and white dorsolateral stripes, small dark lavender spots, and larger white spots. White lining covers about anterior half of ventral parietal peritoneum. White pericardium; digestive tract, liver and kidneys covered by translucent peritonea.

Biology and ecology: During the night, all individuals at the type locality were calling less than 1 m above the water from the upper surfaces of leaves of herbs and ferns overhanging a trickling stream. Other stream-breeding frogs at the type locality included Nymphargus grandisonae, N. griffithsi, and Hyloscirtus alytolylax [120]. Nymphargus balionotus is nocturnal, arboreal, epiphyllous, and usually found on vegetation along clear-water streams in pristine or in moderately disturbed foothill and cloud forests [87]. Parental care unknown.

Call: Not described.

Tadpole: Not described.

Distribution (Figure 133): Nymphargus balionotus occurs between 400 and 1540 m along the western slope of the Cordillera Occidental of Colombia, from El Tambito in the Departamento de Cauca, south to Ecuador [21,87,115,116,120,245]. In Ecuador, it is known from localities in the provinces of Carchi, Imbabura, Pichincha, and Cotopaxi at elevations between 1400 and 1540 m (Specimens Examined). It is found within the Western Foothill and Montane Forest ecoregions.

Figure 133. Distribution of Nymphargus balionotus in Ecuador (yellow dots).

Conservation status: Nymphargus balionotus is listed by the IUCN as Vulnerable at a global level [245]. The species was abundant at the type locality (3.5 km NE of Mindo), where 13 individuals were observed in two nights (7–8 April 1975) [120]. A single reproductive population is known in Ecuador, Reserva Río Manduriacu, which is threatened by mining activities [21]. Thus, in Ecuador, we suggest the category of Critically Endangered, following IUCN criteria B2a + B2b(iii).

Evolutionary relationships: Lynch and Ruiz-Carranza [246] suggested that Nymphargus balionotus is the nearest relative of Nymphargus armatus and N. griffithsi, based on the following characters: (i) Snout short and truncate, (ii) slender habitus, (iii) similar webbing and dentition, and (iv) a large bladelike ventral crest present on the humerus. Based on mitochondrial data, a recent study found that N. balionotus is sister to N. manduriacu [21].

Remarks: Ecuadorian individuals of Nymphargus balionotus have a small, pointed humeral spine. The Colombian populations have a variable crista ventralis that, in some cases, takes the form of a pointed humeral spine. The polymorphism in this characteristic has created confusion in the generic placement of the species [6,21,139]. It also has been suggested that at least two species are currently confused under the name Nymphargus balionotus (M. Rada, pers. com.)

Specimens examined: Nymphargus balionotus: Ecuador: Provincia de Carchi: Cabeceras del Río Baboso (00°53″ N, 78°27″ W; 1400 m), DH-MECN 0865. Provincia de Pichincha: 3.5 km NE Mindo (0.0322 S, 78.761 W, 1540 m), KU 164702 (holotype), 164701, 164703–11. Provincia de Imbabura: Reserva Río Manduriacu, 1240–1254 m, ZSFQ 0531–33.

Photographic record: Ecuador: Provincia de Cotopaxi: Río Lomapi, NE of La Maná, Reserva Ecológica Illinizas (0.8270275 S, 79.0843359 W, ca. 1300 m); photo by Martín Bustamante.

Nymphargus buenaventura Cisneros-Heredia and Yánez-Muñoz, 2007 [115] (Figure 134 and Figure 135).
Cochranella buenaventura Cisneros-Heredia and Yánez-Muñoz, 2007 [115]. Holotype: DHMECN 3563.
Type locality: “Reserva Buenaventura (03°38′ S, 79°45′ W, 1200 m elevation), canton Piñas, Provincia de El Oro, República del Ecuador”.
Nymphargus buenaventura—Cisneros-Heredia and McDiarmid, 2007 [17].

Common names: English: Buenaventura Glassfrog. Spanish: Rana de Cristal de Buenaventura.

Etymology: The specific epithet refers to the type locality, Reserva Buenaventura [115].

Identification: Nymphargus buenaventura can be distinguished from most centrolenids by its green dorsum with small pale yellow to cream spots, reduced webbing between fingers, absence of humeral spine, and absence of iridophores on the digestive visceral peritonea, but with iridophores covering the renal capsules. The last character is shared with at least three other species, N. cariticommatus, N. griffithsi, and N. wileyi. Nymphargus cariticommatus occurs only on the Amazonian Andean slopes of Ecuador, whereas N. buenaventura is restricted to the Pacific Andean slopes. Nymphargus griffithsi differs by having dark dorsal spots. Nymphargus wileyi differs by having a uniform green dorsum and inhabiting the Amazonian slopes of the Ecuadorian Andes. Another species that could be confused with N. buenaventura is N. lasgralarias, which lacks the yellow spots characteristic of the former.

Figure 134. Nymphargus buenaventura in life from Reserva Buenaventura, 1200 m, El Oro province, Ecuador. Photo by Mario Yánez-Muñoz.

Diagnosis: Nymphargus buenaventura has the following traits: (1) Vomerine teeth absent; (2) snout truncated in dorsal view and profile; nostrils elevated, slightly concave internarial area; (3) lower half of tympanic annulus evident, oriented dorsolaterally with dorsoventral inclination; supratympanic fold absent or very weak; (4) dorsal skin slightly shagreen with scattered flat tubercles corresponding to light spots; (5) ventral skin areolate; pair of large, flat subcloacal warts; subcloacal skin granular and enameled; (6) upper half of parietal peritoneum covered by iridophores (condition P2), all other peritonea transparent, except for renal capsules and pericardium covered by iridophores (condition V1); (7) liver tetralobed, uncovered by iridophores (condition H0); (8) humeral spine absent; (9) webbing absent between Fingers I, II, and III; basal between Fingers III and IV; webbing formulae: III 2^2/3—2^1/2 IV; (10) webbing on feet moderate; webbing formulae: I 2—2⁺ II 1⁺—2^1/2 III 1—2^1/2 IV 2^1/2—1^2/3 V; (11) outer ventral edges of forearms and tarsi with low folds; (12) unpigmented nuptial pad Type I, concealed prepollex; (13) first finger shorter than second, (14) eye diameter larger than width of disc on Finger III; (15) color in life: Dorsum green with scattered pale yellow spots; bones green; (16) color in preservative: Dorsal surfaces pale lavender with scattered cream spots; (17) in life, iris yellowish silver with thin dark maroon reticulations; (18) yellow–green hands with bright yellow discs, melanophores present on outer fingers and outer toes; (19) males call from upper sides of leaves along streams; call undescribed; (20) fighting behavior unknown; (21) egg clutches placed on upper sides of leaves along streams; parental care unknown; (22) tadpoles unknown; (23) small body size; SVL of adult males 20.9–22.4 mm (n = 4); SVL in one adult females 23.5 mm.

Color in life (Figure 134): Pale green dorsum with pale yellow to cream spots on all dorsal surfaces, including arms and legs. Hands and feet yellowish green with pale yellow discs. Venter cream. Iris yellowish silver with fine maroon reticulations.

Color in ethanol: Lavender dorsum with cream spots. Anterior two-thirds of parietal peritoneum white, posterior section translucent; pericardium white; most visceral peritonea clear except for white peritoneum covering the renal capsules.

Biology and ecology: Very little information is known on the natural history of Nympharugus buenaventura. It has been found on leaves of shrubs and low trees, 1–3 m above streams. A male was found near an egg clutch with 38 embryos [115]. Parental care is unknown.

Call: Not described.

Tadpole: Not described.

Distribution (Figure 135): Nymphargus buenaventura is known from three localities in El Oro province (Reserva Buenaventura, Marcabelí, Cascadas de Manuel) and one in Azuay province (Luz María), southwestern Ecuador, within the Deciduous Forest ecoregion. The localities are: Reserva Buenaventura (03°38′43″ S, 79°45′48″ W, 1200 m), Luz María (02°41′02″ S, 79°25′01″ W, 770 m), Marcabelí (03°44′48″ S, 79°53′36″ W, 667 m), and Cascadas de Manuel (03°12′22″ S, 79°43′34″ W, 800 m) [115,247].

Figure 135. Distribution of Nymphargus buenaventura in Ecuador (yellow dots).

Conservation status: Nymphargus buenaventura is classified as Data Deficient by the IUCN [248]. The species seems to be endemic to southwestern Ecuador, an area with severe deforestation. Thus, we suggest placing the species in the Endangered conservation category, following IUCN criteria B1, B2a, B2biii. The species is partially protected by a private reserve (Reserva Buenaventura, managed by Fundación Jocotoco).

Evolutionary relationships (Figure 136): Nymphargus buenaventura has not yet been included in any molecular phylogeny. However, because of its close morphological resemblance to N. griffithsi, N. wileyi, and N. cariticommatus, it is considered as part of the Nymphargus clade (sensu Guayasamin et al. [1]).

Specimens examined: Nymphargus buenaventura: Ecuador: Provincia de El Oro: Reserva Buenaventura (03°38′ S, 79°45′ W, 1200 m), DHMECN 3563 (holotype), 2524, 3561, 3562 (paratypes).

Figure 136. Evolutionary relationships of glassfrogs in the genus Nymphargus, inferred using maximum likelihood and Bayesian criteria.

Nymphargus cariticommatus (Wild, 1994 [137]; Figure 137, Figure 138 and Figure 139).
Cochranella cariticommata Wild, 1994 [137]. Holotype: KU 202806.
Type locality: “11.2 km WSW Plan de Milagro (03°02′ S, 78°35′ W, 2350 m), Provincia Morona-Santiago, Ecuador”.
Nymphargus cariticommatus—Cisneros-Heredia and McDiarmid, 2007 [17].

Common names: English: Unadorned Glassfrog. Spanish: Rana de Cristal Escueta.

Etymology: The specific epithet, cariticommatus, is an adjective formed from the Latin word caritus, meaning without, joined with Kommos, a Greek work meaning decoration or embellishment. The name refers to the species’ plain appearance and lack of distinctive features [137].

Identification: Nymphargus cariticommatus is likely to be confused with Nymphargus wileyi, N. griffithsi, and N. buenaventura. However, in life, N. cariticommatus has a green dorsum with small yellow dots (Figure 137; uniform green dorsum in N. wileyi), an esophageal peritonea covered by iridophores (translucent in N. griffithsi and N. buenaventura, and with a thin layer of iridophores in N. wileyi; but see Remarks), and slightly less hand webbing on the outer fingers than N. buenaventura. Additionally, N. cariticommatus is found on the Amazonian versant of the Andes, whereas N. griffithsi and N. buenaventura inhabit the Pacific versant.

Figure 137. Nymphargus cariticommatus in life from Ecuador, Zamora Chinchipe province, Reserva Tapichalaca, 2200 m. Photos by Mario Yánez (left) and Marco Reyes (right).

Diagnosis: (1) Vomers lacking teeth; (2) snout round in dorsal profile, and truncated in lateral profile (Figure 138); (3) tympanum evident and of moderate size (tympanum diameter 26.7%–27.1% of eye diameter), dorsal border of tympanic annulus covered by supratympanic fold, tympanic membrane differentiated from surrounding skin; (4) dorsal surfaces shagreen, with minute spicules in males; (5) pair of enlarged subcloacal warts; (6) anterior two-thirds of ventral parietal peritoneum white, posterior third transparent (condition P2); silvery white pericardium; translucent peritonea covering the intestines, stomach, testes, gall bladder, and urinary bladder; esophagus white; kidneys white with unpigmented spots (condition V1); (7) liver tetralobed, three ventral lobes partially covering one smaller lobe; hepatic peritoneum transparent (condition H0); (8) humeral spines absent; (9) no webbing between inner fingers; webbing formula as follows: IV 2^3/4—2^2/3 V in one male, IV 3⁺—2^3/4 V in one female; (10) foot webbing moderate: I 2—2^1/4 II (1–1^1/2)—(2^1/3–2^2/3) III (1^1/3–1^1/2)—(2^1/2–2^2/3) IV (2^2/3–2^3/4)—2 V (Figure 138); (11) low ulnar fold in males, not visible in females; low inner tarsal fold present in both sexes; (12) concealed prepollex; nuptial pad Type I in males; (13) Finger II slightly longer than Finger I (Finger I about 90%–94% length of Finger II); (14) disc of Finger III of moderate size, 50%–56% of eye diameter; (15) in life, dorsum green with small yellow spots (Figure 137); (16) in ethanol, dorsum lavender with small white spots; (17) iris silvery white with thin black reticulation, yellow ring around pupil; (18) dorsal surfaces of fingers and toes lacking melanophores; (19) males call from the upper side of leaves; call described as soft ‘zeet, zeet, zeet’, but specific characteristics unknown; (20) fighting behavior unknown; (21) eggs deposited on the upper side of leaves; parental care unknown; (22) tadpoles unknown; (23) small body size; one adult male, SVL 23.3 mm; one adult female, SVL 25.5 mm.

Figure 138. Nymphargus cariticommatus. Drawings of the holotype, adult male, KU 202806. Modified from Wild [137].

Color in life (Figure 137): Dorsal surfaces of head, body, and limbs green with small yellow spots. Dorsal surfaces of fingers and toes yellow. Iris silvery white with black reticulations.

Color in ethanol: Dorsum of body and limbs lavender with few, small, scattered white spots (fewer in females). Cheek with pale white flecks; margin of lip unpigmented; tympanum with few lavender flecks in males, lacking in females. Hands and feet lacking pigment [137]. Liver covered by transparent peritoneum; white parietal peritoneum covering anterior two-thirds of venter; silvery white pericardium; transparent peritonea covering the intestines, stomach, testes, gall bladder, and urinary bladder; peritoneum around kidneys with a thin silvery white lining; iris silvery white, with purple reticulations. In the original description, Wild [137] stressed the white esophagus in N. cariticommatus, similar to that seen in Teratohyla midas. In the preserved type series, only a thin layer of iridophores is evident.

Biology and ecology: Nymphargus cariticommatus was found on the upper sides of leaves and ferns within 2 m of cascading streams [137]. Parental care is unknown.

Call: Not described.

Tadpole: Not described.

Distribution (Figure 139): Nymphargus cariticommatus is endemic to the Amazonian slopes of the southern Andes of Ecuador at elevations between 2200 and 2700 m ([115,116,137,139], this work). The species has been recorded from localities in the provinces of Zamora Chinchipe and Morona Santiago. The habitat of the species is within the Eastern Montane Forest ecoregion.

Figure 139. Distribution of Nymphargus cariticommatus in Ecuador (yellow dots).

Conservation status: Globally, Nymphargus cariticommatus is listed as Data Deficient by the IUCN [249]. The last reports of the species come from surveys at the Reserva Biológica Tapichalaca on 2004–2005 ([115], Yánez-Muñoz and Meza-Ramos, pers. com.). In Ecuador, the species is threatened by habitat destruction and fragmentation because of agriculture, pasture lands, and mining. We suggest considering the species as Endangered (IUCN criteria B1, B2a, B2biii).

Evolutionary relationships (Figure 136): Based on mitochondrial data, Nymphargus cariticommatus is sister to N. sucre.

Taxonomic Remarks: As mentioned in the diagnosis, subtle morphological differences separate Nymphargus cariticommatus from N. wileyi. We have examined all the available material of the two species (see Specimens Examined), including the type series, and observed that none of the specimens of N. wileyi have dorsal spots, whereas all individuals of N. cariticommatus have them. Also, in the preserved material, the esophageal peritoneum of N. cariticommatus is more pronounced and extended than in N. wileyi, in which the iridophores are restricted to the anterior portion of the esophageal peritoneum. Considering the data at hand, we maintain the species status of N. wileyi.

Specimens examined: Nymphargus cariticommatus: Ecuador: Provincia de Morona Santiago: 11.2 km WSW Plan de Milagro (03°07′ S, 78°30′ W; 2350 m), KU 202806 (holotype), KU 202805; El Cruzado (3.050 S, 78.517 W; 2194 m), USNM 288435–36. Provincia de Zamora Chinchipe: Reserva Biológica Tapichalaca (4.49208 S, 79.128389 W; 2200 m), DH-MECN 1974, 2429; 18.1 km E of Loja on the old road to Zamora, 2700 m, QCAZ 33977; Shucos, on the old road from Loja to Zamora, MRy 544.

Nymphargus cochranae (Goin 1961 [97]; Figure 140, Figure 141, Figure 142, Figure 143 and Figure 144).
Cochranella cochranae Goin, 1961 [97]. Holotype: BM 1912.11.1.68.
Type locality: “El Topo, Río Pastaza, Eastern Ecuador, 4200 feet.”
Centrolenella cochranae—Goin, 1964 [187].
Cochranella cochranae—Ruiz-Carranza and Lynch, 1991 [6].
Nymphargus cochranae—Cisneros-Heredia and McDiarmid, 2007 [17].

Common names: English: Cochran’s Glassfrog. Spanish: Rana de Cristal de Cochran.

Etymology: The specific epithet honors Doris M. Cochran (1898–1968), herpetologist and curator of the Smithsonian Institution [97].

Identification: Nymphargus cochranae can be distinguished from all other glassfrogs by its green dorsum with small dark blue to black ocelli enclosing orange dots (Figure 140), relatively large size (adult males, SVL 24.0–26.2 mm; adult females, SVL 27.8–30.3 mm), absence of webbing between fingers (Figure 141), and lacking humeral spines. Nymphargus cochranae further differs from other species with dorsal ocellated patterns (N. anomalus, N. lindae sp. nov., N. ignotus, N. laurae, N. ocellatus) by having relatively smaller ocelli (Figure 142). Additionally, N. ignotus is found on the Pacific slopes of the Andes, whereas N. cochranae is restricted to the Amazonian slopes of the Andes. N. anomalus further differs by having scattered black and lavender flecks between ocelli (absent in N. cochranae); N. ocellatus has a green dorsal coloration with large dark rings that, in life, have greenish white centers; N. laurae, known from a single adult male is smaller (SVL = 19.7 mm), has pointed papillae on Toes I–IV (absent in N. cochranae), and larger ocelli. Nymphargus cochranae is most similar to its sister species, N. lindae sp. nov., in female body size (N. cochranae, SVL = 27.8–30.3 mm; N. lindae sp. nov., SVL 27.2–27.8 mm) and the relative size of ocelli (minute in N. cochranae and clearly larger in N. lindae sp. nov.), which is the most conspicuous difference between the two species. The only other species similar to N. cochranae is N. megacheirus, the dorsal dots of which coincide with spicules and could be confused with ocelli. However, N. megacheirus is larger (SVL of males = 26.8–31.5 mm; SVL of females = 31.2–32.9 mm) and the males have conspicuous spicules and spiculated warts on the dorsum (minute spicules in males of N. cochranae).

Figure 140. Nymphargus cochranae in life. Ecuador, Volcán Sumaco, 1500 m, QCAZ 31113. Photos by Luis A. Coloma.

Diagnosis: (1) Vomers usually with teeth, but see Variation; (2) snout truncated in dorsal profile, and truncated to slightly protruding in lateral profile (Figure 141); (3) tympanum relatively small, its diameter 19.5%–27.8% eye diameter, dorsal border of tympanic annulus covered by supratympanic fold, tympanic membrane pigmented as surrounding skin; (4) dorsal skin of males and females shagreen, minute spicules usually present in males; (5) pair of enlarged subcloacal warts; (6) anterior two-thirds of ventral parietal peritoneum white, posterior third transparent (condition P2); white pericardium; translucent peritonea covering intestines, stomach, testes, kidneys, gall bladder, and urinary bladder (condition V1); (7) liver tetralobed, two large ventral lobes partially covering two smaller lobes; hepatic peritoneum transparent (condition H0); (8) humeral spines absent; (9) hand webbing absent; (10) foot about two-thirds webbed: I (2–2⁺)—(2^1/3–2^1/2) II (1^1/2–2⁻)—(3⁻–3) III (1^3/4–2)—(3⁻–3⁺) IV (3⁻–3⁺)—(2–2⁺) V; (11) ulnar and tarsal folds low; (12) concealed prepollex; nuptial pad Type I in males; (13) Finger I about same length as Finger II (Finger II 96.3%–101.8% of length of Finger I); (14) disc of Finger III small, 19.4%–28.3% of eye diameter; (15) in life, dorsum green with small dark blue to black ocelli enclosing orange dots (Figure 140); bones green; (16) in ethanol, dorsal surfaces lavender with small black ocelli enclosing white dots; (17) iris creamy white with thin dark grey reticulation, pupil surrounded by pale brownish–yellow coloration; (18) dorsal surfaces of hands and feet lacking melanophores, except for proximal portion of Finger IV and Toe V; (19) males call from the upper side of leaves or branches overhanging streams; single, high-pitched note; spectral and temporal characteristics of call unknown; (20) fighting behavior unknown; (21) eggs deposition site unknown; parental care unknown; (22) tadpoles unknown; (23) small to medium body size; SVL of adult males 24.0–26.2 mm (X = 25.0 ± 0.812, n = 6); SVL of adult females 27.8–30.3 mm (X = 28.9 ± 1.014, n = 7).

Color in life (Figure 140): Green dorsum with small dark blue to black ocelli enclosing bright orange spots. Ventral surfaces mostly white, except for posterior third, which is translucent. Bones green. Iris greyish white with a thin black reticulation; area immediately around pupil pale brownish yellow.

Color in ethanol (Figure 143): Dorsal surfaces lavender with small black ocelli enclosing white spots. Pericardium and anterior two-thirds of ventral parietal peritoneum white (presence of iridophores). Liver, intestines, stomach, testes, kidneys, gall bladder, and urinary bladder without white lining.

Figure 141. Nymphargus cochranae. Head in lateral and ventral views, adult female, QCAZ 22197. Hand in ventral view, adult male, QCAZ 31113. Illustrations by Juan M. Guayasamin.

Figure 142. Dorsal patterns of ocellated glassfrogs, excluding Nymphargus laurae and N. lindae sp. nov. (A) N. anomalus, KU 143299. (B) N. cochranae, KU 121035. (C) N. ocellatus, LSU 25990. (D) N. ignotus, ICN 14750. Figure modified from Lynch and Duellman [22].

Figure 143. Nymphargus cochranae, holotype in preservative, BMNH 1912.11.1.68. Ecuador, El Topo, Río Pastaza. Photos by Martín Bustamante.

Variation: Goin [97] described Nymphargus cochranae as lacking teeth on the dentigerous process of the vomer, but also noted the presence of teeth in six other specimens [187]. Most specimens (n = 13) examined by us present teeth (USNM 286634 lacks teeth) and we assume that this is the usual character state in the species.

Biology and ecology: Nymphargus cochranae is active during the night. In the reproductive season, males call at night from the upper surfaces of leaves or branches near streams. Parental care is unknown.

Call: Described by Lynch and Duellman [22] as a single, high-pitched note. The specific spectral and temporal characteristics of the call are unknown.

Tadpole: Not described.

Distribution (Figure 144): Nymphargus cochranae occurs in Ecuador and Colombia on the Amazonian slopes of the Andes at elevations between 1170 m and 1960 m. The species has been reported from localities in the provinces of Napo, Orellana, Pastaza, Tungurahua, Sucumbíos, and Zamora Chinchipe (Specimens Examined). There is an unconfirmed record from the Ecuadorian Amazonia (Garza 1, at 300 m; QCAZ 1216). The potential distribution of the species is 21,723 km², within the Eastern Foothill Forest and Eastern Montane Forest regions. The only record of the species in Colombia comes from the Serranía de Churumbelos [250].

Figure 144. Distribution of Nymphargus cochranae in Ecuador (yellow dots).

Conservation status: Globally, Nymphargus cochranae is considered as Vulnerable by the IUCN [251]. In Ecuador, the species remains abundant at several localities, including the Sumaco and Cayambe Coca National Parks. However, part of the species’ range is fragmented; thus, we suggest considering N. cochranae as Near Threatened.

Evolutionary relationships (Figure 136): Nymphargus cochranae is sister to N. lindae sp. nov. However, one sequenced individual (QCAZ 31340) is inferred as sister to a clade formed by N. griffithsi + N. lasgralarias.

Taxonomic Remarks: Nymphargus cochranae might be a species complex (see Evolutionary relationships). Also, see taxonomic remarks of N. laurae and N. lindae sp. nov.

Specimens examined: Nymphargus cochranae: Ecuador: Provincia de Orellana: km 13 on Loreto–Coca road (0.5836 S, 77.234 W), QCAZ 22196–97; Provincia Napo: Cascada San Rafael (0.1127 S, 77.596 W, 1280–1372 m), QCAZ 002, USNM 286632–37, 284304–10; 14.7 km (by road) NE of Río Salado (0.1289 S, 77.608 W, 1310 m), USNM 286638; 14 km by road SW of Reventador (ca. 0.11278 S, 77.596 W; 1400–1500 m), USNM 284304–06; Pacto Sumaco (0.726 S, 77.566 W; 1400 m), QCAZ 31113; Río Salado, 1 km upstream from Rio Coca (0.1916 S, 77.6997 W, 1420 m), KU 164517–18. Provincia de Pastaza: Abitagua, 8 km NW Mera (1.41667 S, 78.1667 W, 1300 m), KU 121033–35; Provincia de Tungurahua: Río El Topo (1.4166 S, 78.1667 W, 1220 m), BMNH 1912.11.1.68; 11 km E Río Negro (1.433 S, 78.13 W, 1170 m), KU 146605; Provincia de Sucumbíos: S slope Cordillera del Due above Río Coca (0.0833 N, 77.66 W, 1150 m), KU 123216–18. Provincia de Zamora Chinchipe: Estación Científica San Francisco (3.967 S, 79.066 W, 1960 m), QCAZ 31340–41.

Localities from the literature: Nymphargus cochranae: Ecuador: Provincia de Zamora Chinchipe: Alto Nangaritza (4.25026 S, 78.61746 W; 1256–1430 m) [252]; Contrafuerte de Tzunantza (4.046 S, 78.922 W) [253]; Alto Machinaza 1 (3.8976 S, 78.482 W) [254], Alto Machinaza 2 (3.7708 S, 78.554 W) [254].

Nymphargus colomai new species Guayasamin and Hutter (Figure 145).
http://zoobank.org/urn:lsid:zoobank.org:act:46978D55-22DF-409E-870C-F3C2203A5E6E

Common names: English: Coloma’s Glassfrog. Spanish: Rana de Cristal de Coloma.

Etymology: The specific name is a patronym for Luis A. Coloma in recognition of his pioneer and continual efforts in studying and protecting amphibians, as well as mentoring numerous students, including JMG. Luis Coloma is the Director of the Jambatu Center for Research and Conservation of Amphibians (Centro Jambatu de Investigación y Conservación de Anfibios; see http://www.anfibiosecuador.ec/). The Jambatu Center houses dozens of critically endangered amphibians, in an outstanding effort to conserve frogs and toads. Luis received the Sabin Award for Amphibian Conservation in 2007 (http://www.amphibians.org/grants/sabin-award/).

Holotype: QCAZ 41590, adult male, collected from Miazi Alto (4.25044° S, 78.61356° W; 1260 m), Cordillera del Cóndor, Provincia de Zamora Chinchipe, Ecuador, by Juan M. Guayasamin and Elicio Tapia on 8 April 2009.

Paratypes: QCAZ 41591–92, adult males, and QCAZ 41641, juvenile, collected by Juan M. Guayasamin, Elicio Tapia, Silvia Aldás, and Holger Braun on 8–12 April 2009, at the same locality as holotype.

Generic placement: The new species is placed in the clade Nymphargus (sensu Guayasamin et al. [1]) based on morphological and molecular data. All species in Nymphargus share an absence of webbing between Fingers I–III and an absence or reduced webbing between Fingers III and IV; additionally, males lack humeral spines (except N. grandisonae). Nymphargus colomai sp. nov. has the aforementioned traits. Molecular analyses of mitochondrial genes unambiguously place the new species in the clade Nymphargus (Figure 16).

Identification: Nymphargus colomai sp. nov. is unique by having a white iris with a contrasting black horizontal stripe and a dorsum that varies from dull yellowish green to tangerine yellow, brown, or grey olive, always with numerous small yellow to orange spots (Figure 145). Additionally, N. colomai sp. nov. lacks humeral spines and hand webbing. Among glassfrogs that inhabit the Amazonian slopes of the Andes, only N. cariticommatus, N. chancas, N. humboldti sp. nov., N. siren, and N. sucre could be confused with N. colomai sp. nov. However, N. cariticommatus, N. chancas, N. siren, and N. humboldti sp. nov. have a green dorsum with few and well-defined dorsal spots, whereas N. colomai sp. nov. has a non-green dorsum (see above) with numerous and diffuse yellow dorsal spots. Moreover, N. siren is smaller than N. colomai sp. nov. (N. siren, SVL < 23.3 mm; N. colomai sp. nov. SVL = 24.7–25.7 mm); and N. cariticommatus has white esophagus and renal capsules (cream in N. colomai sp. nov.) Both N. colomai sp. nov. and the Peruvian N. chancas share the clearly defined black horizontal stripe on a silver background, but the two species are easily differentiated by their dorsal color patterns (in N. colomai sp. nov.: Dull yellowish green to tangerine yellow, brown, or grey olive, with numerous small yellow to orange spots; in N. chancas: Yellowish–green dorsum with small yellow dots; compare Figure 145 and Figure 146) and genetics (Figure 136).

Figure 145. Nymphargus colomai sp. nov. in life from stream nearby Miazi Alto, Zamora Chinchipe province, Ecuador. (Top left): QCAZ 41590, holotype; other frogs are part of the type series. Photos by Holger Braun and Juan M. Guayasamin.

Figure 146. Nymphargus chancas in life from 5.9 km SE from the type locality of Abra Tangarana, San Martín, 6°19′15.65” S 76°41′44.16” W, 973 m, CORBIDI 10471. Photos by Jesse Delia (upper row) and Evan Twomey (lower row).

Diagnosis: (1) Dentigerous process of the vomer lacking teeth or with few teeth (up to 3); (2) snout truncated to slightly protruding in lateral profile; truncated in dorsal view; (3) tympanum oriented almost vertically, with slight lateral and posterior inclinations, its diameter about 23%–27% of eye diameter; upper half of tympanic annulus obscured by supratympanic fold and warts; tympanic membrane pigmented as surrounding skin; (4) dorsal skin shagreen, with numerous spiculated warts and spicules in males; females unknown; (5) venter areolate; pair of enlarged subcloacal warts; (6) white parietal peritoneum covering about anterior half of venter (condition P2); white pericardium; translucent peritonea on kidneys, intestines, stomach, gall and urinary bladders (condition V1); (7) liver lobate, covered by translucent peritoneum (condition H0); (8) humeral spines absent; (9) webbing absent between Fingers I, II, and III, absent or basal between Fingers III and IV; webbing formula III (2^3/4–3)—(2^1/2–2^2/3) IV; (10) feet about two-thirds webbed; webbing formula: I 2⁻—2^1/3 II (1⁺–1^1/3)—2^1/2 III 1^1/2—(2^2/3–2^3/4) IV 3⁻—(1^1/2 – 2⁻) V; (11) ulnar and tarsal folds present, low; (12) concealed prepollex; in males, nuptial pad Type I; (13) Finger II slightly longer than Finger I; (14) disc of Finger III width about 46%–50% of eye diameter; (15) in life, dorsum varies from dull yellowish green to tangerine yellow, brown, or grey olive with numerous small yellow to orange spots; bones green; (16) in preservative, dorsum grey to greyish lavender with numerous unpigmented spots; (17) in life, iris silvery white with slight yellow hue and a clearly marked horizontal black stripe; (18) melanophores on dorsal surfaces of Fingers III and IV and Toes IV and V; (19) males call from the upper surfaces of leaves; call unknown; (20) fighting behavior unknown; (21) egg deposition site unknown; parental care unknown; (22) tadpole unknown; (23) small body size; in males, SVL 25.0–25.7 mm (n = 3); females unknown.

Description of holotype: Adult male, SVL 25.0 mm. Head slightly wider than long (head length 98% of head width); snout truncated in dorsal and lateral profiles; canthus rostralis indistinct, slightly concave; loreal region slightly concave; lips slightly flared; nostril protuberant, closer to tip of snout than to eye, directed frontolaterally; internarial area barely depressed. Eyes large, directed anterolaterally at an angle ~50°; transverse diameter of disc of Finger III 48% eye diameter. Supratympanic fold low, obscuring upper portion of tympanic annulus; tympanum oriented mostly vertically, but with slight posterolateral inclination; tympanic membrane translucent, pigmented as surrounding skin. Dentigerous process of vomer low, situated transversely between choanae, with one or two teeth; choanae large, longitudinally rectangular; tongue ovoid, with ventral posterior fifth not attached to floor of mouth and posterior margin slightly notched; vocal slits extending posterolaterally from about the lateral margin of tongue (at about half the length of tongue) to angle of jaws. Humeral spine absent. Low ulnar folds evident on external and internal ventrolateral margins of arm; relative lengths of fingers: III > IV > II > I; webbing absent between Fingers I–III, basal between Fingers III and IV, webbing formula III 3–2^2/3 IV; discs expanded, nearly elliptical; disc pads nearly triangular shaped; subarticular tubercles small, round, simple; low supernumerary tubercles present; palmar tubercle elliptical, simple; nuptial pad large (Type I), ovoid, granular, extending from ventrolateral base to dorsal surface of Finger I, covering proximal half of Finger I. Length of tibia 57% SVL; low inner tarsal fold evident; outer tarsal fold absent; foot two-thirds webbed; webbing formula of foot: I 2⁻—2^1/3 II 1^1/4—2^1/2 III 1^1/2—2^3/4 IV 3⁻—2⁻ V; discs on toes round to elliptical, lacking papillae; disc on Toe IV slightly narrower that disc on Finger III; disc pads triangular; inner metatarsal tubercle large, ovoid; outer metatarsal tubercle small and inconspicuous; subarticular tubercles small, round; supernumerary tubercles low. Skin on dorsal surfaces of head, body, and lateral surface of head and flanks shagreen with numerous spiculated warts and minute spicules; lower flanks show glandular cells; throat smooth; belly and lower flanks areolate; cloacal opening directed posteriorly at upper level of thighs; cloacal warts present, unpigmented. Ventral surface of thighs with pair of enlarged tubercles.

Coloration of holotype in life (Figure 145): The holotype has a grey olive dorsal coloration, with numerous yellow to orange spots. The venter is white anteriorly and translucent posteriorly. The iris is silvery white, with a clearly marked black horizontal stripe. Dorsally, at the posterior end of the head, there is a circular, concave area with dark grey collocation.

Color variation in life (Figure 145): Dorsum can be dull yellowish green, tangerine yellow, brown, or grey olive, always with numerous small and diffuse yellow to orange spots. Anterior half of venter white, posterior portion transparent. Iris might have a slight yellow hue.

Coloration of holotype in ethanol: Dorsal surfaces of head, body, and limbs greyish brown, with small, unpigmented spots. Middorsal dark lavender spot at scapular level. Anterior half of ventral parietal peritoneum white, posterior half cream. White pericardium; translucent peritonea covering digestive tract, liver, kidneys, and gall and urinary bladders.

Color variation in ethanol: Dorsal surfaces of head, body, and limbs vary from greyish brown to lavender with small, unpigmented spots.

Measurements of holotype (mm): Nymphargus colomai sp. nov., adult male, QCAZ 41590: SVL = 25.0, head length = 8.1, head width = 8.3, snout length = 3.3, IOD = 2.6, upper eyelid width = 2.5, eye diameter = 3.3, tympanum diameter = 0.9, tympanum–eye distance = 1.7, femur = 13.1, tibia = 14.2, foot length = 11.5, hand length = 5.7, disc of Finger III width = 1.6, disc of Toe IV width = 1.5.

Biology and ecology: Nymphargus colomai sp. nov. is active during the night. All individuals were found on vegetation 100–170 cm above fast-flowing streams. Males call from the upper surfaces of leaves. Parental care is unknown.

Call: Not described.

Tadpole: Not described.

Distribution (Figure 147): Nymphargus colomai sp. nov. is known only from its type locality at the Cordillera del Cóndor, Zamora Chinchipe province, Ecuador.

Figure 147. Distribution of Nymphargus colomai sp. nov. in Ecuador (yellow dot).

Conservation status: Following UICN criteria, we consider Nymphargus colomai sp. nov. as Endangered, following IUCN criteria B2a, B2(iii). The main threats for this species are habitat destruction (i.e., cattle, agriculture) and contamination associated with mining.

Evolutionary relationships (Figure 136): N. colomai sp. nov. is inferred as sister to N. mariae.

Taxonomic comments: Nymphargus colomai sp. nov. was previously confused with N. chancas [252], which we restrict to Peru. Preserved specimens are morphologically similar, but in life, several differences are evident (Figure 145 and Figure 146). The two species are not closely related (Figure 136).

Nymphargus garciae (Ruiz-Carranza and Lynch, 1995 [255]; Figure 148 and Figure 149).
Cochranella garciae Ruiz-Carranza and Lynch, 1995 [255]. Holotype: ICN 11752.
Type locality: “Departamento de Cauca, municipio de Inzá, Km 64–73 carretera Popayán a Inzá, vertiente oriental Cordillera Central, 2°34′ latitud N, 76°4′ W de Greenwich, 2590–2660 m [Colombia]”.
Nymphargus garciae—Cisneros-Heredia and McDiarmid, 2007 [17].

Common names: English: García’s Glassfrog. Spanish: Rana de Cristal de García.

Etymology: The specific epithet is a patronym for Dr. Evaristo García, author of the pioneer book Los ofidios venenosos del Cauca, published in 1896 [255].

Identification: Nymphargus garciae can be differentiated from other glassfrogs by its moderate body length (SVL 25.9–28.4 mm in adult females, SVL 25.1–29.9 mm in adult males) ([255], this work), green dorsum usually with dark green to blue spots, and absence of humeral spines. In Ecuador, N. garciae could be confused with Centrolene buckleyi and N. megacheirus. However, C. buckleyi has basal webbing between Fingers III and IV (absent in N. garciae), lacks dark spots on the dorsum (usually present in N. garciae), and males with prominent humeral spines (spines absent in N. garciae). Nymphargus megacheirus has a similar dorsal coloration and hand-webbing pattern as N. garciae, but the two species have allopatric distributions, N. garciae inhabiting higher elevations (1900–2700 m) than N. megacheirus (1300–1740 m). Additionally, N. megacheirus is slightly larger than N. garciae (in N. megacheirus, SVL 26.8–31.5 mm in adult males, 31.2–32.9 mm in adult females). Two species endemic to Colombia, N. nephelophilus and N. oreonympha, are extremely similar to N. garciae. Nymphargus oreonympha is reported to have basal webbing between Fingers III and IV (webbing absent in N. garciae), whereas N. nephelophilus has a smooth skin (shagreen in N. garciae) and lacks ulnar and tarsal folds (present in N. garciae). See Remarks.

Figure 148. Nymphargus garciae in life from Ecuador, Napo province, trail from Oyacachi to El Chaco, 3012 m, MZUTI 764. Photos by Eduardo Toral.

Diagnosis: (1) Vomerine teeth absent; (2) snout rounded in dorsal aspect, truncated to slightly sloping in lateral profile; (3) tympanum oriented almost vertically, with slight posterior inclination, small in size (tympanum diameter 22.9%–30.6% of eye diameter); tympanic annulus visible except for dorsal border covered by supratympanic fold; tympanic membrane partially pigmented but differentiated from surrounding skin; (4) dorsal surfaces shagreen, with spicules evident in males; (5) pair of enlarged subcloacal warts (Figure 15); (6) anterior half of parietal peritoneum white (condition P02; white pericardium and renal capsules, translucent visceral peritoneum (condition V1); (7) tetralobed liver covered by transparent peritoneum (condition H0); (8) humeral spines absent; (9) no webbing between fingers, although junction of lateral fringes resemble basal webbing; (10) webbing formula on foot: I (2⁻–2⁺)—(2⁺–2^2/3) II (1–1^3/4)—(2^1/2–3⁻) III (1^1/4–1^3/4)—(2^1/3–3⁻) IV (2^1/3–3⁺)—(1^3/4–2⁺) V; (11) ulnar and tarsal folds present, enameled; (12) nuptial pad Type I, concealed prepollex; (13) Finger II slightly longer than Finger I (FI/FII = 0.87–0.95); (14) disc of Finger III relatively large (3DW/ED = 0.56–0.77); (15) in life, dorsum green with or without green to blue spots; white upper lip and ulnar and tarsal folds; bones green; (16) in preservative, dorsum lavender with or without dark lavender spots; (17) iris greyish white with thin black reticulation; (18) fingers lacking melanophores dorsally; melanophores restricted to proximal dorsal areas of Toes IV and V; (19) males call from upper side of leaves; call unknown; (20) fighting behavior unknown; (21) eggs deposition site unknown; parental care unknown; (22) tadpoles unknown; (23) medium body size; in Ecuador, males SVL 24.9–29.9 mm (X = 26.7 ± 1.723; n = 7); in Colombia, males SVL 25.1–27.7 mm (X = 26.3 ± 0.745; n = 15), females SVL 25.9–28.4 mm (n = 3).

Color in life (Figure 148): Dorsum dark green with cream tubercles and dark green to blue spots; hands and feet yellowish green. Glandular pericloacal region white. Pericardium and anterior half of ventral parietal peritoneum white; translucent visceral peritonea. Based on observations on preserved specimens, we assume that part of the type series of Nymphargus garciae (ICN 11743, 11759) was uniform green in life, corresponding to the uniform lavender coloration in ethanol. Some of the photographs of Ecuadorian individuals of N. garciae show a uniform green dorsal coloration.

Color in ethanol: Dorsal surfaces lavender with or without small dark spots; spicules resemble minute white spots in males. White upper lip; tympanic membrane pigmented with purple specks. Cloacal region with white warts. Dorsally, fingers and toes mostly unpigmented, but some pigmentation visible on proximal half of Toes IV and V. White parietal peritoneum covering anterior half of venter; silvery–white pericardium; transparent peritoneum covering liver and urinary bladder; gastrointestinal peritoneum translucent; renal capsules white with minute unpigmented spots, as those in N. wileyi ([20]: Figure 12). Internal traits based on a dissected male (ICN 11759).

Biology and ecology: Adults of Nymphargus garciae were found on vegetation and rocks along streams; one gravid female (ICN 7495) had numerous eggs, which had a dark brown animal and a cream vegetal pole [255]. Parental care is unknown.

Call: Not described.

Tadpole: Not described.

Distribution (Figure 149): Nymphargus garciae is known from localities on the eastern slope of the Cordillera Central at elevations of 1900–2700 m in Colombia [101,255], and from the Cordillera Oriental of the Ecuadorian Andes at 2550–3012 m. In Ecuador, this species is found in the Eastern Montane Forest ecoregion.

Figure 149. Distribution of Nymphargus garciae

Article Menu

Glassfrogs of Ecuador: Diversity, Evolution, and Conservation

Abstract

1. Introduction

2. Methods

2.1. Taxonomy and Species Concept

2.2. Characters and Terminology

2.3. Evolutionary Relationships

2.4. Biogeographic Regions of Ecuador

2.5. Potential Distribution

2.6. Registration of New Nomenclatural Acts

3. Results

3.1. Phylogenetic Systematics

3.2. Potential Distribution

4. Species Accounts