New insights into the diversity of Oreobates frogs (Anura: Craugastoridae): description of a new species from the Peruvian Yungas and comments on O. quixensis and O. saxatilis

423 New insights into the diversity of Oreobates frogs © 2025 Deutsche Gesellschaft für Herpetologie und Terrarienkunde e.V. (DGHT), Germany Open access at https://www.salamandra-journal.com 30 October 2025 ISSN 0036–3375 SALAMANDRA 61(4): 423–441 SALAMANDRA German Journal of Herpetology New insights into the diversity of Oreobates frogs (Anura: Craugastoridae): description of a new species from the Peruvian Yungas and comments on O. quixensis and O. saxatilis Ernesto Castillo-Urbina1,2, Miguel Vences3, Mathias Dezetter4, Frank Glaw6, Christine Burgos2, Lucia Aliaga2, Grecia Torres-Ccasani1,2, Sydney F. Hope4,5, Alejandro Mendoza1,2 & Jörn Köhler7 1 Universidad Nacional Mayor de San Marcos, Museo de Historia Natural (MUSM), Departamento de Herpetología, Av. Arenales 1256, Lima 11, Peru 2 Asociación Ararankha: Ecología y Conservación, Lima, Peru 3 Zoological Institute, Technische Universität Braunschweig, Mendelssohnstr. 4, 38106 Braunschweig, Germany 4 Association Nature Conserv’Action, Montpellier 34430, France 5 Department of Psychology, Hunter College, City University of New York, 695 Park Avenue, New York, NY 10065, USA 6 Zoologische Staatssammlung München (ZSM-SNSB), Münchhausenstr. 21, 81247 München, Germany 7 Hessisches Landesmuseum Darmstadt, Friedensplatz 1, 64283 Darmstadt, Germany Corresponding author: Jörn Köhler, ORCID 0000-0002-5250-2542, e-mail: [email protected] Manuscript received: 27 May 2025 Accepted: 1 August 2025 by Stefan Lötters Abstract. We investigate the taxonomic status of specimens of the pristimantine frog genus Oreobates recently collected in montane rainforest within the Biocorredor Bosques de Vaquero ‘Shunku Sacha’, San Martín Department, Peru. A phylogenetic analysis of these frogs based on DNA sequences of the mitochondrial 16S rRNA gene revealed a lineage closely related to Oreobates colanensis. However, that lineage is divergent from O. colanensis by 10.1% uncorrected pairwise distance in the 16S gene fragment, and from other species of Oreobates included in the analysis by more than 13.8%. Moreover, specimens in that lineage are distinguished from those of all recognized nominal species of Oreobates by a unique combination of morphological characters, including skin on the dorsum granular, absence of dorsolateral folds, presence of nuptial pads in males, absence of vocal slits and vocal sacs in males, and the absence of basal webbing on toes. Consequently, we describe this lineage as a new species, Oreobates shunkusacha sp. n. We also report on additional vouchers of Oreobates collected in the lowland Amazonian rainforest of Panguana, Huánuco Department, central Peru. Based on morphology, mitochondrial (16S) and nuclear genes (POMC, RAG-1), we demonstrate that Oreobates specimens from this locality belong to two distinct species-level lineages, referred to as O. quixensis and O. saxatilis, thus providing conclusive evidence for sympatric occurrence of these two nominal taxa that were previously considered to have parapatric ranges. Furthermore, we provide and discuss data suggesting that a misidentification of previously sequenced specimens, including the paratopotype of O. saxatilis, is likely responsible for the paraphyly of O. quixensis and O. saxatilis in previously published mitochondrial trees. Upon correction of these putative misidentifications, the morphologically diagnosable species O.quixensis and O. saxatilis are monophyletic and occur in close sympatry at certain localities in the upper Amazon basin. Key words. Amphibia, Pristimantinae, molecular genetics, morphology, Peru, sympatry, systematics, taxonomy, misidentification, conservation. Introduction The genus Oreobates Jiménez de la Espada, 1872 comprises a group of Neotropical frogs with assumed direct development which are distributed from southern Colombia to northern Argentina, with most species occurring along the eastern slopes of the Andes, reaching elevations of up to 3850 m a.s.l. (Padial et al. 2012, Montero-Mendieta et al. 2021). These frogs inhabit a variety of ecosystems, including humid Amazonian lowland, semi-deciduous forests, montane humid and dry forests, as well as high elevation Puna grasslands (Padial et al. 2008, 2012). Currently, Oreobates includes 26 described species (Frost 2025) but, despite recent taxonomic progress, species diversity re- 424 Ernesto Castillo-Urbina et al. mains incompletely known. Candidate species have been identified by molecular data, including populations from Bolivia and Peru (Padial et al. 2012, Köhler & Padial 2016), and await formal taxonomic study. Given the suggested high rate of speciation of the genus particularly in the Yungas montane forests, which remain underexplored and partly difficult to access, and the comparatively low number of specimens available for certain species in scientific collections, a comprehensive understanding of the genus’ diversity, distribution and morphological variation is still lacking (Padial et al. 2012, Pansonato et al. 2020, Venegas et al. 2021). In addition, phylogenetic studies confirm that Oreobates, as currently defined, is monophyletic, but there are partly ambiguous results with respect to the most basal phylogenetic relationships within the genus (Padial et al. 2012, 2014, Montero-Mendieta et al. 2021, Venegas et al. 2021). During fieldwork in Peru, we collected specimens of Oreobates. Among these is one putative new species from the Biocorredor Bosques de Vaquero ’Shunku Sacha’ in San Martín Department, Peru, which includes the three Conservation Concessions (CC) Sacha Runa, Yaku Kawsanapa, and Cordillera de Vaquero. It is an ecological corridor within the northern sector of the buffer zone of the Cordillera Azul National Park, connecting it to the southern margins of the Cordillera Escalera Regional Conservation Area. The Cordillera Azul is a mid-elevation mountain ridge (< 2350 m a.s.l.) limited by the Huallaga river basin to the west, and by the Ucayali river basin to the east. The physiography of this ridge, with basimontane forest surrounding areas of Yungas mountain forest, appears to have led to the diversification of unique and endemic frog species (e.g., Myers & Daly 1979, Flores & McDiarmid 1989, Lötters et al. 1997, 2002, Lötters 2003, Brown & Twomey 2009). Recent taxonomic studies have revealed new species or re-discoveries of anurans that are currently considered to be endemic to the Cordillera Azul (Cusi et al. 2017, Castillo-Urbina et al. 2021, 2023, Köhler et al. 2022) but over all this mountain range remains little explored. The same is true for the Cordillera Escalera (< 2220 m a.s.l.), with most knowledge on amphibians from this area being limited to the southernmost portion of this mountain ridge (e.g., Duellman & Schulte 1993, Twomey et al. 2014, Cusi et al. 2020). We also collected Oreobates specimens in the Amazonian lowlands of central Peru. Specimens from this region are recovered as one lowland clade encompassing the species O. quixensis and O. saxatilis. Even though O. quixensis and O. saxatilis are reciprocally diagnosable morphologically, they were recovered as paraphyletic based on mitochondrial DNA phylogenetic analyses, which constitutes a taxonomic challenge (Padial et al. 2012). Recent molecular population structure analyses showed the data for samples of O. quixensis and O. saxatilis were mostly consistent with the presence of two clusters, but the clusters did not fully correspond to respective species identifications (Montero-Mendieta et al. 2021). Potential explanations included incomplete lineage sorting, misidentifications of voucher specimens and ongoing genetic flow between the two species (Padial et al. 2012, Montero-Mendieta et al. 2021). In this study, we investigate the taxonomic status of the Oreobates population discovered in the montane forest of the CCs Sacha Runa and Yaku Kawsanapa in the San Martín Department, provide evidence for its specific distinctness from all known species of Oreobates, and describe it as a new species. In addition, we provide new data for the lowland species O. quixensis and O. saxatilis and discuss the implications for their phylogenetic relationships and distribution. Materials and methods Fieldwork and voucher specimens Fieldwork was conducted in October 2022 and in March 2025 in different areas of the Biocorredor Bosques de Vaquero ‘Shunku Sacha’, San Martín Department, Peru, including the Conservation Concession (CC) Sacha Runa and the CC Yaku Kawsanapa, in collaboration with the community-based associations managing the two CCs, namely AESARUSA, and ABAFYK (see Shanee et al. 2015, 2020 for overviews about CCs in San Martín Department), and during multiple expeditions between 2008 and 2019 at the Area de Conservación Privada Panguana, Huánuco Department, central Peru. Specimens were observed and collected during opportunistic surveys at night and day. Geographical coordinates were recorded using handheld GPS receivers set to WGS84 datum. In March 2025, we also opportunistically recorded the field-active body temperature of individuals, as well as substrate and air temperatures at the exact location where each individual was found, using an infrared thermometer (Fluke 62 Max). Collected specimens were euthanized with an overdose of 5% lidocaine or benzocaine gel applied on the ventral surfaces of individuals (McDiarmid 1994). Tissue samples were taken prior to fixation and stored in 99% ethanol, while specimens were fixed using 96% ethanol or formalin and subsequently stored in 70% ethanol. Specimens were deposited in the herpetological collection of the Museo de Historia Natural, Universidad Nacional Mayor de San Marcos (MUSM), Lima, Peru, and Zoologische Staatssammlung München (ZSM), Germany. All additional museum abbreviations used follow those of Frost (2025). FGZC refers to Frank Glaw field numbers. Morphology Morphological measurements (in millimeters) were taken to the nearest 0.1 mm by the first author using a digital caliper, with the aid of a stereoscopic microscope. For proper comparison, the definition of morphological character states follows those established by Duellman & Lehr (2009) and the diagnostic and descriptive schemes follow those of Padial et al. (2012). The measurements taken and 425 New insights into the diversity of Oreobates frogs used throughout the text are as follows: SVL (snout–vent length), HL (head length; straight-line distance from the posterior corner of the mouth to the tip of the snout), HW (head width; measured at the level of the jaw angle), TD (tympanum diameter; measured horizontally), IND (internarial distance), IOD (interorbital distance; between the anterior margins of the orbits), ED (horizontal eye diameter), EW (upper eyelid width), END (eye–nostril distance; from the anterior margin of the orbit to the center of the nostril), HaL (hand length; from the proximal edge of the inner metacarpal tubercle to the tip of the third finger), TL (tibia length; from the femur–tibia articulation to the tibia–heel proximal articulation), THL (thigh length; from the middle of the cloacal slit to the proximal part of the femur–tibia articulation), and FL (foot length; distance from the proximal margin of the inner metatarsal tubercle to the tip of toe IV). Color in life is described based on digital photographs. Molecular genetics First, our molecular genetic analysis primarily aimed at the identification of lineage divergence among focal samples of Oreobates using the mitochondrial 16S rRNA (16S) gene. For representative taxon sampling, we used BLAST searches (Altschul et al. 1990) of newly generated 16S sequences of the Oreobates samples from Shunku Sacha against the GenBank nucleotide archive and downloaded a selection of sequences of representatives of all nominal species (and few candidate species) of Oreobates (sensu Venegas et al. 2021), with O. zongoensis being the only nominal species for which 16S sequence data are not available. Two species of Lynchius (L. simmonsi, L. waynehollomonae), the sister group of Oreobates (Pyron & Wiens 2011, Motta et al. 2016), were added to the sampling as outgroup taxa. Second, as our first analysis unexpectedly indicated the presence of two species-level lineages at Panguana, we performed the same analysis again by now adding all 16S sequences available from GenBank for the two nominal species O. quixensis and O. saxatilis, plus sequences from all Oreobates specimens collected by us at Panguana. To assess potential admixture between the two distinct Oreobates lineages occurring at Panguana, we furthermore sequenced the nuclear-encoded genes POMC and RAG-1 of these samples. DNA was extracted using a standard salt extraction protocol. Fragments of the mitochondrial gene for 16S rRNA (16S) and the nuclear-encoded gene for pro-opiomelanocortin (POMC) and the recombination-activating gene 1 (RAG-1) were amplified in polymerase chain reactions (PCR) with the following primers and cycling conditions: 16S: primers 16SAr-L (5’-CGCCTGTTTATCAAAAACAT-3’) and 16SBr-H (5’-CCGGTCTGAACTCAGATCACGT-3’) (Palumbi et al. 1991), cycling conditions 94 °C (60 sec), 33 cycles of 95 °C (45 sec), 55°C (45 sec), 72 °C (90 sec), and final elongation at 72°C (600sec); POMC: primers POMC-DRVF1 (5’-ATATGTCATGASCCAYTTYCGCTGGAA-3’) and POMC-DRVR1 (5’-GGCRTTYTTGAAWAGAGTCATTAGWGG-3’) from Vieites et al. (2007), cycling conditions 95 °C (120 sec), 40 cycles of 95 °C (45 sec), 47 °C (45 sec), 72 °C (60 sec), and final elongation at 72 °C (600 sec); RAG-1: two pairs of primers were successively used in a nested PCR approach, first PCR with primer pair Rag1-Mart-F1 (5’-AGCTGGAGYCARTAYCAYAARATG-3’) and Rag-1Mart-R6 (5’-GTGTAGAGCCARTGRTGYTT-3’), modified from Martin (1999), second PCR with primers Rag-1-AmpF2 (5’-ACNGGNMGICARATCTTYCARCC-3’) and Rag1-UC-R (5’-TTGGACTGCCTGGCATTCAT-3’) from Chiari et al. (2004) and Rakotoarison et al. (2015), both PCRs with cycling protocol 94 °C (240 sec), 45 cycles of 94 °C (45 sec), 45 °C (40 sec), 72 °C (120 sec), and final elongation at 72 °C (600 sec). PCR products were resolved on automated DNA sequencers by LGC Genomics (Berlin, Germany). All new DNA sequences were submitted to GenBank (accession numbers PX131115–PX131127 and PX125881–PX125892). 16S sequences were combined with those from other studies downloaded from GenBank. A full table with all sequences used, their accession numbers and metadata is available from the Zenodo repository (https://doi.org/10.5281/zenodo.16876682). To estimate evolutionary relationships within Oreobates, we used a Maximum Likelihood (ML) phylogenetic approach on the 16S alignment. Sequences were exported to AliView v1.26 (Larsson 2014), edited, and aligned using MAFFT v7.310 (Katoh & Standley 2013) with the L-INS-i parameter as the iterative refinement method (Katoh et al. 2005), resulting in an alignment of 588 base pairs. We employed the W-IQ-TREE web server (Trifinopoulos et al. 2016) to infer a molecular phylogeny under ML as the optimality criterion. The best evolutionary model was determined using ModelFinder under the Bayesian Information Criterion (BIC) implemented in IQ-TREE, identifying TIM2+F+I+G4 as the best-fitting substitution model for the dataset (-m TEST command; Kalyaanamoorthy et al. 2017). To evaluate branch support, ultrafast bootstrap values were obtained from 2000 pseudoreplicates and 10,000 iterations as the maximum number to stop (-bb 2000 and -nm 10,000 commands in IQ-TREE). Additionally, the Shimodaira-Hasegawa approximate likelihood ratio test (SH-aLRT) was performed with 1000 replicates (-alrt 1000 command; Shimodaira & Hasegawa 1999, Guindon et al. 2010). Genetic divergences were quantified as uncorrected pairwise distances (p-distances) using pairwise alignments in MEGA v11 (Tamura et al. 2021) with default settings (pairwise deletion). For the Panguana samples, the alignments of the nuclear-encoded POMC and RAG-1 gene fragments (430 and 1328bp, respectively) were analyzed independently from the mitochondrial sequences to understand concordance (or absence thereof) in their differentiation. We used a genealogy visualization approach to graphically represent the relationship among alleles (haplotypes). Haplotypes were estimated with the PHASE algorithm (Stephens et al. 2001), and haplotype genealogies estimated with the Fitch tree algorithm (Salzburger et al. 2011, Matschiner 2016) in Hapsolutely (part of iTaxoTools) (Vences et al. 2021, 2024). 426 Ernesto Castillo-Urbina et al. Threat status We assessed the Red List status of the species according to the IUCN Red List criteria (IUCN 2001). Given the habitats and elevations where we observed Oreobates from CCs Sacha Runa and Yaku Kawsanapa, we estimated its geographic range by calculating the area within the Biocorredor Bosques de Vaquero mountain ranges at elevations higher than 1350 m a.s.l. using QGIS software version 3.42.1. Nomenclatural acts The electronic edition of this article conforms to the requirements of the amended International Code of Zoological Nomenclature, and hence the new name contained herein is available under that Code from the electronic edition of this article. This published work and the nomenclatural acts it contains have been registered in ZooBank, the online registration system for the ICZN. The LSID (Life Science Identifier) for this publication is: urn:lsid:zoobank. org:pub:7C33D393-DC77-4C6C-8189-218DDF20C180. The electronic edition of this work was published in a journal with an ISSN, and has been archived and is available from the following digital repositories: zenodo.org, salamandrajournal.com. Results Identity of Oreobates populations from Shunku Sacha Our Maximum Likelihood tree (Fig. 1) recovered Oreobates monophyletic with high support (bootstrap/ SHaLRT values = 100/100). The tree topology is generally in agreement with those published previously (Padial et al. 2012, Köhler & Padial 2016, Pansonato et al. 2020, Vene gas et al. 2021), except for some ambiguous relationships receiving no support (O. amarakaeri, O. lundbergi) and most basal nodes receiving only low to moderate support. The three samples of the population from Shunku Sacha are recovered monophyletic and sister to O. colanensis with high support (99/99). Uncorrected pairwise distances in the 16S gene fragment of the samples from Shunku Sacha to O. colanensis amount to 10.1%, and range between 13.8–21.4% to all other known species of Oreobates included in the analysis. Morphological examination of the Shunku Sacha specimens and comparison with museum vouchers revealed that they exhibit a combination of external characters that distinguish them from all currently recognized nominal species of Oreobates (see below). Consequently, with these three independent lines of evidence (phylogenetic relationships, high mitochondrial divergence and morphological distinctness), we regard that population a distinct evolutionary lineage and describe it as a new species. Taxonomy Oreobates shunkusacha sp. n. ZooBank LSID: urn:lsid:zoobank.org:act: FF57198D-C042-4097-9ED9-9415826B9314 Holotype: MUSM 41865, adult female (Figs 2–3), from Conservation Concession Sacha Runa (-6.713915, -76.109471, 1496 m a.s.l.), Sauce District, San Martín Province, San Martín Department, Peru, collected on 28 October 2022 (21:34 h) by L. Aliaga, C. Burgos, S. F. Hope, and M. Dezetter. Paratypes: MUSM 41863–41864, two adult males, from CC Sacha Runa (-6.715414, -76.103853 and -6.714223, -76.10752, at 1596 and 1540 m a.s.l., respectively), collected on 28 October 2022 (between 19:31 to 20:47 h) by L. Aliaga, C. Burgos, S. F. Hope, and M. Dezetter; MUSM 42182, one adult female, and MUSM 42183–42184, two adult males, from CC Sacha Runa (-6.71545, -76.10317, 1600 m a.s.l.), MUSM 42185, one adult female, from CC Sacha Runa (-6.71488, -76.10691, 1552 m a.s.l.), collected between 19 and 22 March 2025 (between 19:50 to 22:16 h) by C. Burgos, E. Castillo-Urbina, M. Fernandez, M. Valle, T.Vasquez, and M. Dezetter. All localities in Sauce District, San Martín Province, San Martín Department, Peru. Referred specimens: MUSM 42186, 42188–42189, three adult males, from CC Yaku Kawsanapa (-6.64827, -76.16221, 1414 m a.s.l.) and MUSM 42187, one adult male, from CC Yaku Kawsanapa (-6.64734, -76.16152, 1351 m a.s.l.) Chazuta District, San Martín Province, San Martín Department, Peru, collected on 12 March 2025 (between 19:10 to 20:20h) by C. Burgos, E. Castillo-Urbina, M. Fernandez, M.Valle, T. Vasquez, and M. Dezetter. Definition: A moderately robust species of Oreobates (SVL of adult males 24.4–26.4 mm, n = 8; adult females SVL 37.9–41.8 mm, n = 3), defined by the following combination of characters: (1) skin of dorsum granular, granules small, round to subconical, homogeneous in size; a thin vertebral fold present; dorsolateral folds absent; venter smooth; posterior surfaces of thighs areolate, skin in groin smooth; discoidal fold present; postrictal tubercles present, rounded; (2) tympanic membrane distinct, large, about 40–54% of eye diameter; anterior part of tympanic annulus distinct; supratympanic fold present, well-defined, concealing the upper part of the tympanic annulus; (3) head slightly longer than wide to nearly as long as wide (HW/HL = 0.88–0.99); snout long, rounded in dorsal view and in lateral profile; canthus rostralis sinuous in dorsal view, rounded in cross-section; (4) cranial crests absent; upper eyelid bearing small, scattered, subconical tubercles, being slightly larger than dorsal tubercles; (5) dentigerous process of vomers large, prominent, oval in shape, situated posteromedial to choanae (posterior margin at level of choanae), their width about 1.6 times the diameter of choanae, bearing 5 to 6 vomerine teeth; (6) males without 427 New insights into the diversity of Oreobates frogs vocal slits and vocal sacs; (7) hands with long and slender fingers, first finger slightly longer than second; subarticular tubercles large and prominent, conical; supernumerary tubercles prominent, round, smaller than subarticular tubercles; fingertips round, barely enlarged, lacking circumferential grooves; lateral fringes on fingers present, weak; nuptial pads present on males, dorsally and medially on Finger I; (8) antebrachial and ulnar tubercles present, low and minute, 1–3 ulnar tubercles; (9) heel and outer side of tarsus bearing very low subconical tubercles, being slightly larger than tubercles on dorsal surfaces of thighs; (10) inner metatarsal tubercle ovate, prominent; outer metatarsal tubercle smaller than the inner metatarsal tubercle, subconical, prominent; subarticular tubercles conical, prominent; supernumerary tubercles smaller than subarticular tubercles, prominent, round; (11) toes long and slender, lateral fringes present and weak, webbing absent; toe V and III equal in length, reaching to the middle of second Figure 1. Maximum Likelihood phylogenetic tree of samples of Oreobates, inferred from an alignment of the mitochondrial 16S rRNA gene. Numbers at nodes are rounded bootstrap values in percent (2000 pseudoreplicates; not shown if < 50%), followed by SH-aLRT values (1000 replicates; not shown if < 50) as calculated with IQ-TREE. The taxon name is followed by sample locality and GenBank accession number, or voucher number for newly produced sequences (terminals in bold font). Two samples of Lynchius were used to root the tree (not shown for better graphical presentation). Inset photo depicts the holotype of O. shunkusacha sp. n. (MUSM 41865) in life. 428 Ernesto Castillo-Urbina et al. subarticular tubercle of Toe IV; tips of toes slightly enlarged, rounded, lacking circumferential groove or ungual flap on toe I, tips of toes II, III, IV, and V bearing distinct ungual flaps; foot length 49.8–57.8% of SVL; (12) axillary glands absent. Diagnosis: Oreobates shunkusacha differs from all other known species of Oreobates by the unique combination of skin on dorsum granular with granules being small, round to subconical and homogeneous in size, absence of dorsolateral folds, presence of nuptial pads in males, absence of vocal slits and vocal sacs in males, and the absence of basal webbing on toes. In external morphology, O. shunkusacha superficially resembles O. lehri, O. zongoensis, and O. machiguenga in having a granular dorsal skin texture, with granules being homogeneous in size and in the absence of dorsolateral folds. It mainly differs from O. lehri and O. zongoensis (characters in parentheses) in having nuptial pads in males (absent), and males lacking vocal slits (present). Furthermore, it differs from O. lehri by having the first finger slightly longer than second (finger I shorter than finger II), supernumerary tubercles on feet present, distinct and subconical (absent), and the absence of axillary glands (present). Additionally, it can be distinguished from O.zongo ensis by supernumerary tubercles on plantar surfaces being distinct and subconical (indistinct), tips on toes slightly enlarged, rounded (slightly enlarged, truncated), lateral fringes on toes present and weak (absent), dorsal color pattern in life with pale chevron-shaped marking (uniformly dark brown, markings absent), ventral surfaces with some dark mottling (uniformly pinkish-brown), and a golden iris in life (bright copper-red). The new species can be distinguished from O. machiguenga in having a disFigure 2. Adult female holotype of Oreobates shunkusacha sp. n. (MUSM 41865) in life (SVL 37.9 mm): (a) lateral, (b) dorsal and (c)ventral views. Not to scale. 429 New insights into the diversity of Oreobates frogs tinct supratympanic fold (absent), canthus rostralis sinuous in dorsal view (convex), dentigerous processes of vomers oval in shape (triangular), snout long (short), and webbing on toes absent (present, basal between toe III and toe IV). The new species may occur in geographical proximity to other species of Oreobates (i.e., O. saxatilis, O. quixensis, O. lundbergi). It differs mainly from all these by having skin of dorsum granular, with granules being small and homogeneous in size (finely tuberculate with distinctly larger scattered tubercles in O. saxatilis, tuberculate with many large subconical tubercles in O. quixensis, and smooth with small scattered tubercles in O. lundbergi). Furthermore, O.shunkusacha differs from O. saxatilis and O. quixensis by having nuptial pads in males (absent). Oreobates shunkusacha is most closely related to O.colanensis and mainly differs from it by the absence of dorsolateral folds (present, distinct), the presence of nuptial pads in males (absent), the absence of contrasting color pattern on the hidden surfaces of body, as axilla, groins, anterior and posterior surfaces of thighs, and shank (present, white or cream blotches on a dark brown background), and belly pale with faint brown marks to dark brown mottling (belly dark brown suffused with red with numerous white flecks). Holotype description: Adult female (Figs 2–3). Head slightly wider than body, head longer than wide (HW/ HL= 0.93); snout long, rounded in dorsal view and in lateral profile (Fig. 3c); nostrils slightly protuberant, oriented laterally; canthus rostralis slightly sinuous in dorsal view, rounded in cross-section; loreal region slightly concave, sloping gradually to the lips; lips not flared; upper eyelid with numerous, small, subconical granules; cranial crests absent. Supratympanic fold short, extending to anterior level of insertion of arm and to the inferior level of the end of the tympanum, concealing the upper part of the annulus posteriorly; tympanic membrane distinct, anterior part of tympanic annulus distinct; tympanic membrane nearly round, transparent, its diameter slightly less than half of eye diameter; postrictal tubercles present, round. Choanae not concealed by palatal shelf of the maxillary arch when roof of mouth is viewed from below; choanae medium in size, round, separated by a distance equal to five times the diameter of choana; dentigerous processes of the vomers prominent, oblique, oval, situated posteromedial to choanae (posterior margin at level of choanae), their width about 1.6 times the diameter of choanae, bearing 5–6 vomerine teeth. Skin on dorsum and flanks granular, with small, low, round granules, homogeneous in size, covering the entire dorsum; ventral surfaces smooth; thighs coarsely areolate posteriorly; occipital fold V-shaped; dorsum bearing one thin vertebral fold, hardly recognizable, form the tip of the snout to the pelvic region; dorsolateral fold absent; discoidal fold present, weak; thoracic fold present, distinct; skin in groin areolate. Antebrachial tubercle and three ulnar tubercles present, low and minute; palmar tubercle divided in two subunits, inner irregular in form larger than outer oval; thenar tubercle prominent, triangular in outline; supernumerary tubercles prominent; subarticular tubercles rounded, larger than supernumerary tubercles; finger tips narrowly rounded; pads absent, circumferential grooves and ungual flaps absent; fingers lacking lateral fringes and webbing; relative length of fingers: I > II < III > IV (Fig. 3e). Toes long and slender, foot length 52% of SVL; heel and outer side of tarsus bearing very low conical tubercles, tarsus lacking folds; inner metatarsal tubercle oval, prominent, larger than outer metatarsal tubercle; outer metatarsal tubercle conical, prominent; plantar supernumerary tubercles present, distinct, subconical; subarticular tubercles prominent, conical; toes with weak lateral fringes proximally; toe webbing absent; toe tips rounded, barely expanded, without circumferential groove or ungual flap on toe I; tips of toes II, III, IV, and V each bearing distinct ungual flap; relative length of toes: I < II < III > V < IV (Fig. 3d). Toe V and Toe III reaching to the middle of second subarticular tubercle of Toe IV. Measurements (in mm): SVL 37.9; TL 19.7; FL 19.3; HL15.2; HW 14.1; ED 4.2; TY 1.8; IOD 3.3; EW 3.5; IND3.3; E–N 4.8. Figure 3. Preserved female holotype of Oreobates shunkusacha sp. n. (MUSM 41865): (a) dorsal and (b) ventral views; (c) lateral view of the head; (d) plantar surface of right foot; and (e) palmar surface of right hand. 430 Ernesto Castillo-Urbina et al. In life (Fig. 2), dorsal surfaces dark-brown with a narrow pale chevron that extents onto the flanks; snout paler brown compared to dorsum; canthal stripe absent; two pale labial bars, irregular, narrow in width above and becoming wider below; lower jaw dark brown with some white speckles; supratympanic fold and anterior part of the annulus blackish; arms reddish brown with diffusely defined dark brown transverse bars; dorsal surfaces of hands brown with fine white markings; legs pale brown with diffusely defined dark-brown transverse bars, more clearly defined medially; dorsal surfaces of feet with well-defined dark-brown transverse bars. Ventrally, throat greyish with a pinkish tint, with few scattered white marks at the margin; brachial and antebrachial surface with a pale patch; chest, belly and ventral surface of forelimbs cream with dark brown mottling, palms and soles dark brown. Iris greenish-golden with black reticulation and a fine darkbrown circumpupillary line. In 70% ethanol (Fig. 3), color pattern is the same as in life with dorsal surfaces brown, but top of head turned paler than the background color. Markings on head are darker than the dorsum; pale chevron-shaped marking on dorsum and labial bars more distinct than in life; flanks dark brown; forearms brownish cream (lacking the reddish color in life), legs pale brown with dark brown cross-bars with some pale edges. Ventral surfaces whitish-cream with gray mottling. Variation: The studied females are larger in size compared to males. Body proportions are rather similar in all individuals. For variation in measurement and proportions see Table 1. All specimens have the skin on dorsum granular and with granules homogeneous in size, with the females having only few scattered slightly enlarged subconical granules, whereas in the males these slightly enlarged subconical granules are more numerous, especially on the posterior dorsum. The discoidal fold is weakly expressed in the females, whereas in the males the discoidal fold is distinct. The fine mid-dorsal fold starts at the anterior part of the snout, or in the scapular region, as in MUSM 41863 and 41864. The inner subunit of the palmar tubercle is irregular in shape in the holotype, but oval in all other specimens. Variation is also evident with respect to color pattern. Dorsal coloration varies from the presence of a pale chevron, irregular pale markings, as seen in MUSM 42182, or a lack of such pattern, as in MUSM 42187. Paratypes MUSM 41864 and 42183 exhibit faint pale dorsolateral stripes that begin at the posterior corner of the eye and extend to the lumbar region. In specimen MUSM 42189, the dorsolateral stripe is broader with blurred edges, and its pale coloration extends across the lumbar region. Female MUSM 42185 exhibits some large white blotches on the dorsal surface of the left hindlimb. Male MUSM 42186 has a thin light-brown vertebral line extending from the scapular region to the cloaca. Ventral coloration on belly ranges from cream with dark brown mottling to less contrasting mottling, as in MUSM 41864 and 42189, or pinkish cream with only faint mottling, as in MUSM 41863 and 42187 (Fig. 4). Distribution and natural history: Oreobates shunkusacha is only known from two localities, the CC Sacha Runa, located in Sauce District, and the CC Yaku Kawsanapa, located in Chazuta District, San Martín Department, Peru, at elevations of 1351–1600 m a.s.l., in the Biocorredor Bosques de Vaquero ‘Shunku Sacha‘, within the buffer zone of Parque Nacional Cordillera Azul (Fig. 5). CCs Sacha Runa and Yaku Kawsanapa comprise primary basimontane Yunga Table 1. Morphological measurements (in mm) and proportions of specimens of Oreobates shunkusacha sp. n. (see text for abbreviations). ** = holotype; * = paratype. MUSM 41865** 42182* 42185* 41864* 41863* 42183* 42184* 42186 42187 42188 42189 sex female female female male male male male male male male male SVL 37.9 41.8 38.8 24.7 26.3 26.4 24.6 24.7 24.4 24.5 25.2 TL 19.7 21.2 20.5 13.8 14.3 13.5 13.3 13.4 12.7 13.5 13.8 FL 19.3 21.3 21.0 13.7 14.1 13.6 13.7 13.3 12.1 14.1 12.6 HL 15.2 17.1 15.8 10.4 10.6 11.2 10.9 10.2 10.0 10.4 10.6 HW 14.1 16.0 15.6 9.1 9.3 10.3 9.7 10.1 8.7 9.6 10.0 ED 4.2 5.2 5.8 3.3 3.5 3.5 3.8 3.7 3.7 3.8 3.9 IOD 3.3 4.2 3.6 2.4 2.5 2.6 2.2 2.5 2.3 2.6 2.4 EW 3.5 3.7 3.1 2.4 2.5 2.2 2.3 2.2 2.1 2.4 2.2 IND 3.3 4.1 3.5 2.6 2.5 2.8 2.6 2.6 2.3 2.4 2.3 E-N 4.8 5.3 4.8 3.0 3.0 3.2 3.1 3.1 3.0 3.1 3.2 TY 1.8 2.3 2.3 1.6 1.6 1.9 1.7 1.7 1.5 1.8 1.6 TL/SVL 0.52 0.51 0.53 0.56 0.54 0.51 0.54 0.54 0.52 0.55 0.55 HL/SVL 0.40 0.41 0.41 0.42 0.40 0.42 0.44 0.41 0.41 0.42 0.42 HW/HL 0.93 0.94 0.98 0.88 0.88 0.91 0.89 0.99 0.87 0.92 0.94 TY/ED 0.43 0.44 0.40 0.48 0.46 0.54 0.45 0.45 0.41 0.47 0.41 431 New insights into the diversity of Oreobates frogs forests that are bordered by patches of secondary forest, with deforested areas and agricultural plots nearby (Fig.6). The individuals were found active during the night between 19:00 and 22:40 h, and were observed either on leaf litter or on moss, both on steep slopes and on plane relief within primary basimontane forest. Individuals were also observed a few meters from the mountain lake ‘Cocha La Encantada’, on the leaf litter within dense vegetation covered by moss and ferns (Fig. 6b). In March 2025, the mean field body temperature (± standard deviation) of active individuals was 18.3 ± 0.3 °C, while the mean substrate and air temperatures were 18.0 ± 0.5 °C and 21.3 ± 2.7 °C, respectively. Microhabitats consisted of dense forest undergrowth on rocky and sandy soil, covered by a thick layer of leaf litter and roots, near small surface and underground streams (Fig. 6c). These forest habitats are temporarily flooded during periods of heavy rain. Some individuals were observed retreating under tree roots and into complex hollow microhabitats. Gravid females were observed in March, with cream to yellow eggs visible through the skin on the posterior venter and flanks. In contrast, the female holotype collected in October was not gravid. Other anurans recorded in close sympatry with O. shunkusacha are Dendropsophus cf. aperomeus, Scinax cf. pedromedinae, Pristimantis sp. (lacrimosus group), P. sp. (danae group), Chiasmocleis cf. tridactyla, and Rhinella sp. (festae group). Oreobates saxatilis and O. quixensis occur at nearby sites at lower elevations (verified by preliminary 16S barcoding results). Calls remain unknown. Conservation status: The buffer zone of the Cordillera Azul National Park is facing high deforestation rates associated with agricultural expansion and is the National Park buffer zone in San Martín that experienced the highest fragmentation rate between 2017 and 2021, with the number of forest patches increasing nearly fivefold (Chávez & Puerta 2024). Within the Biocorredor Bosques de Vaquero, CC Sacha Runa and CC Yaku Kawsanapa are facing significant deforestation threats associated with the expansion of agricultural land for coffee crops toward higher elevations (Fig.6d, Supplementary Fig. S1). Even though these locally run, landscape-level Conservation Concession initiatives can provide effective solutions to deforestation, they currently lack access to support and economic resources (Shanee et al. 2015). All individuals were observed in undisturbed primary basimontane forest. Within the Biocorredor Bosques de Vaquero mountain range, we estimated a geographic range of 30.83 km² of basimontane Yunga Figure 4. Photographs showing variation among specimens of Oreobates shunkusacha sp. n. in life in (a) lateral, (b) dorsal, and (c)ventral views (* = mirrored images). Not to scale. 438 Ernesto Castillo-Urbina et al. O. saxatilis in its respective type series appears to be the most parsimonious conclusion to explain paraphyly in phylogenetic studies. The same is tentatively assumed for the second O. saxatilis sequence clustering with O. quixensis and originating from specimen MUBI 9200 (San Roque de Cumbasa, San Martín, Peru). Unfortunately, attempts to find this specimen in the collection to examine its morphological characters have failed thus far (J. C. Chaparro, pers. comm.). Apart from misidentification of specimens, confusion or contamination of samples in the laboratory during DNA extraction could also explain phylogenetic results, but we consider the latter scenario less likely. Upon correction of these putative misidentified specimens used for sequencing, the resulting phylogeny would be very clear in revealing two monophyletic clusters, one corresponding to O. quixensis and the other corresponding to O. saxatilis, with close sympatry of both species proven for the locality Panguana (Fig. 7). As is obvious from the mapping of localities included in the genetic sampling (Fig.5), as well as preliminary barcoding results of samples from lower elevations in the Biocorredor Bosques de Vaquero ‘Shunku Sacha’ (see above), individuals of both clades occur in close geographical proximity in the San Martín Department, Peru, and probably occur sympatrically at additional localities, including the type locality of O. saxatilis, if our assumptions are correct. Such distribution patterns of divergent but closely related sister species exhibiting overlapping ranges and occurring in sympatry at certain localities in lowland Amazonia is not unusual and has recently been documented for another sister pair of pristimantine frogs, Pristimantis asimus and P. reichlei (Köhler et al. 2024). Together with former studies, our investigation exemplifies that we are still at the beginning regarding the understanding of the diversity of Oreobates frogs. Additional fieldwork, broader sampling, and more comprehensive genetic data are needed to enhance the knowledge of Oreobates systematics, their morphological variation, their biolo gy and potential threats. Acknowledgements Many thanks to C. Aguilar-Puntriano (MUSM) who generously supported the work of our team in multiple ways. For sharing data, photographs of specimens and miscellaneous support, we thank J. C. Chaparro (MUBI) and J. M. Padial (Universidad Granada). J. Aparicio† (CBF), W. Böhme (ZFMK), B. T. Clark (BMNH), W. E. Duellman† and L. Trueb (KU) kindly provided access to the collection under their care. A. P. Motta (KU) kindly took the photographs of the O. saxatilis paratype and provided valuable comments on the manuscript. We thank H. G. Chanchari, Y. Trigoso García, F. Reategui Ramirez, E. R. García, V. Arce Sangama, A. del Aguila Ushiñahua, and M. Arce Chistama, members of the Asociación Ecológica Sacha Runa Sauce (AESARUSA), and C. Zumba Tananta, C. Gatica Rengifo, B. Shapiama Apagueño, J. P. Shapiama Zumba, S. Alvarez Garcia, P. Shapiama Zumba, members of the Asociación Agro Bio Agroforestal Yaku Kawsanapa (ABAFYK), who actively participated in fieldwork and collaborated in this project. We are grateful and acknowledge their important work in voluntarily conserving and managing the forests of the CC Sacha Runa and the CC Yaku Kawsanapa, as we recognize their valuable contributions to this research, and the importance of their local and traditional knowledge, which they willingly integrated through the participatory science methods we jointly applied. Thanks to R. Vilca Lucana, J. Fachin Ruiz, and the entire AMPA Perú team for their valuable contribution to fieldwork organization, and C. Aubert from Nature Conserv’Action for his key contribution to the funding and implementation of our field research. Thanks to K. Bravo Leyva for supervising logistics and planning together with D. E. Rado Tipte, who also provided drone photography and maps, and estimated the geographic range of O. shunkusacha. We thank D. A. Barrera Moscoso and C. A. Ramirez Peralta for their assistance with administrative and permit application procedures. We are also grateful to A. Wong, O. Machuca, D. Olivera (members of Ararankha: Ecología y Conservación), and J. C. Cusi for their support with sample and specimen processing, and to L. Flechtner, J. Horz, G. Keunecke, and L. Rothe for their support with laboratory work. For help during fieldwork in 2025, we thank M. Fernandez, M. Valle, T. and Vasquez. Fieldwork in the Biocorredor Bosques de Vaquero ’Shunku Sacha’ was conducted under the research and collection permits of SERFOR (RDG N° D000136-2022-MIDAGRI-SERFORDGGSPFFS-DGSPFS, RDG N° D000079-2024MIDAGRI-SERFOR-DGGSPFFS-DGSPFS), and that at ACP Panguana by INRENA (RDG N° 124-2008-INRENA-IFFS-DCB), and SERFOR (RDG N° 007-2014-SERFOR-DGGSPFFS, RDG N° 0406-2017-SERFOR-DGGSPFFS). Fieldwork in 2022 was funded by Nature Conserv’Action, and by Amazónicos por la Amazonía – AMPA Perú through the project “Ecoturismo científico y comunitario en el biocorredor de concesiones para conservación Cordillera Vaquero – Yaku Kawsanapa – Sacha Runa. Zona de amortiguamiento del Parque Nacional Cordillera Azul y el Área de Conservación Regional Cordillera Escalera, Región San Martín”, funded by the Spanish cooperation agency AECID Perú. Fieldwork in 2025 was led by Nature Conserv’Action and Ararankha: Ecología y Conservación, through the project “Co-exist: equitable partnerships and communities in action for the biodiversity of the Biocorridor Bosques de Vaquero in the Andean Amazon”; supported by the Support Programme for biodiversity Civil Society Organisations active in developing countries (ProBioDev) managed by the IUCN French Committee, and funded by the French development agency (AFD), Audemars Piguet Foundation for Trees, Fondation de France, Fondation Manthano, Fondation Egis, and Maisons du Monde Foundation; and also supported by funding from the fonds de dotation Le Poids du Vivant. References Altschul, S. F., W. Gish, W. Miller, E. W. Myers & D. J. Lipman (1990): Basiclocal alignment search tool. – Journal of Molecular Biology, 215: 403–410. Araujo-Vieira, K., A. C. C. Lourenço, J. V. A. Lacerda, M. L. Lyra, B. L. Blotto, S. R. Ron, D. Baldo, M. O. Pereyra, A. M. Suárez-Mayorga, D. Baêta, R. Barbosa Ferreira, C. L. Barrio-Amorós, C. Borteiro, R. A. Brandão, C. A. Brasileiro, M. A. Donnelly, M. J. M. Dubeux, J. Köhler, F. Kolenc, F. Sá Fortes Leite, N. M. Maciel, I. Nunes, V. G. D. Orrico, P. Peloso, T. L. Pezzuti, S. Reichle, F. J. M. Rojas-Runjaic, H. R. Da Silva, M. J. Sturaro, J. A. Langone, P. C. A. Garcia, M. Trefaut Rodrigues, D. R. Frost, W. C. Wheeler, T. Grant, J. P. Pombal Jr., C. F. B. Haddad 439 New insights into the diversity of Oreobates frogs & J. Faivovich (2023): Treefrog diversity in the Neotropics: Phylogenetic relationships of Scinaxini (Anura: Hylidae: Hylinae). – South American Journal of Herpetology, 26 (special issue): 1–143. Brown, J. L. & E. Twomey (2009): Complicated histories: three new species of poison frogs of the genus Ameerega (Anura: Dendrobatidae) from north-central Peru. – Zootaxa, 2049: 1–38. Castillo-Urbina, E., F. Glaw, C. Aguilar-Puntriano, M. Vences & J. Köhler (2021): Genetic and morphological evidence reveal another new toad of the Rhinella festae species group (Anura: Bufonidae) from the Cordillera Azul in central Peru. – Salamandra, 57: 181–195. Castillo-Urbina, E., M. Vences, C. Aguilar-Puntriano, F. Glaw & J. Köhler (2023): Contributing to the taxonomic inventory of green-colored rain frogs: A new species of the Pristimantis lacrimosus group (Anura: Strabomantidae) from the southern Cordillera Azul, central Peru. – Vertebrate Zoolo gy, 73: 1047–1061. Chávez, G. & R. Puerta (2024): Patrones de fragmentación en las zonas de amortiguamiento de las Áreas Naturales Protegidas en la región San Martín, Perú. – The Biologist, 22: 253–263. Chiari, Y., M. Vences, D. R. Vieites, F. Rabemananjara, P. Bora, O. Ramilijaona Ravoahangimalala & A. Meyer (2004): New evidence for parallel evolution of colour patterns in Malagasy poison frogs (Mantella). – Molecular Ecology, 13: 3763–3774. Cusi, J. C., L. A. G. Gagliardi-Urrutia, I. C. Brcko, D. B. Wake & R. von May (2020): Taxonomic status of the Neotropical salamanders Bolitoglossa altamazonica and Bolitoglossa peruviana (Amphibia: Caudata: Plethodontidae), with the description of a new species from northern Peru. – Zootaxa, 4834: 365–406. Cusi, J. C., J. Moravec, E. Lehr & V. Gvoždík (2017): A new species of semiarboreal toad of the Rhinella festae group (Anura, Bufonidae) from the Cordillera Azul National Park, Peru. – ZooKeys, 673: 21–47. Duellman, W. E. (1990): A new species of leptodactylid frog, genus Ischnocnema, from Peru. – Occasional Papers of the Musuem of Natural History, University of Kansas, 138: 1–7. Duellman, W. E. & E. Lehr (2009): Terrestrial-breeding frogs (Strabomantidae) in Peru. – Natur und Tier Verlag, Münster. Duellman, W. E. & R. Schulte (1993): New species of centrolenid frogs from northern Peru. – Occasional Papers of the Musuem of Natural History, University of Kansas, 155: 1–33. Flores, G. & R. W. McDiarmid (1989): Two new species of South American Centrolenella (Anura: Centrolenidae) related to C. mariae. – Herpetologica, 45: 401–411. Fouquet, A., A. Gilles, M. Vences, C. Marty, M. Blanc & N. J. Gemmell (2007): Underestimation of species richness in neotropical frogs revealed by mtDNA analyses. – PLoS One, 2: e1109. Frost, D. R. (2025): Amphibian Species of the World: an Online Reference. Version 6.2. – Electronic Database accessible at https://amphibiansoftheworld.amnh.org/index.php, accessed 23 June 2025. – American Museum of Natural History, New York, USA Guindon, S., J.-F. Dufayard, V. Lefort, M. Anisimova, W. Hordijk & O. Gascuel (2010): New algorithms and methods to estimate maximum-likelihood phylogenies: Assessing the performance of PhyML 3.0. – Systematic Biology, 59: 307–321. Kalyaanamoorthy, S., B. Q. Minh, T. K. F. Wong, A. Von Haeseler & L. S. Jermiin (2017): ModelFinder: Fast model selection for accurate phylogenetic estimates. – Nature Methods, 14: 587–589. Katoh, K. & D. M. Standley (2013): MAFFT multiple sequence alignment software version 7: Improvements in performance and usability. – Molecular Biology and Evolution, 30: 772– 780. Katoh, K., K. I. Kuma, H. Toh & T. Miyata (2005): MAFFT version 5: Improvement in accuracy of multiple sequence alignment. – Nucleic Acids Research, 33: 511–518. Köhler, J. & J. M. Padial (2016): Description and phylogenetic position of a new (singleton) species of Oreobates Jiménez de la Espada, 1872 (Anura: Craugastoridae) from the Yungas of Cochabamba, Bolivia. – Annals of Carnegie Museum, 84: 23–38. Köhler, J., E. Castillo-Urbina, C. Aguilar-Puntriano, M. Vences & F. Glaw (2022): Rediscovery, redescription and identity of Pristimantis nebulosus (Henle, 1992), and description of a new terrestrial-breeding frog from montane rainforests of central Peru (Anura, Strabomantidae). – Zoosystematics and Evolution, 98: 213–232. Köhler, J., F. Glaw, C. Aguilar-Puntriano, S. CastroviejoFisher, J. C. Chaparro, I. De la Riva, G. Gagliardi-Urrutia, R. Gutiérrez, M. Vences & J. M. Padial (2024): Similar looking sisters: A new sibling species in the Pristimantis danae group from the southwestern Amazon basin (Anura: Strabomantidae). – Zoosystematics and Evolution, 100: 565– 582. Larsson, A. (2014): AliView: A fast and lightweight alignment viewer and editor for large datasets. – Bioinformatics, 30: 3276–3278. Lötters, S. (2003): On the systematics of the harlequin frogs (Amphibia: Bufonidae: Atelopus) from Amazonia. III: A new, remarkably dimorphic species from the Cordillera Azul, Peru. – Salamandra, 39: 169–180. Lötters, S., P. Debold, K. Henle, F. Glaw & M. Kneller (1997): Ein neuer Pfeilgiftfrosch aus der Epipedobates pictusGruppe vom Osthang der Cordillera Azul in Perú. – Herpetofauna, 110: 25–34. Lötters, S., W. Haas, S. Schick & W. Böhme (2002): On the systematics of the harlequin frogs (Amphibia: Bufonidae: Atelopus) from Amazonia. I: Description of a new species from the Cordillera Azul, Peru. – Salamandra, 38: 95–104. Martin, A. P. (1999): Substitution rates of organelle and nuclear genes in sharks: implicating metabolic rate (again). – Molecular Biology and Evolution, 16: 996–1002. Matschiner, M. (2016): Fitchi: Haplotype genealogy graphs based on the Fitch algorithm. – Bioinformatics, 32: 1250–1252. McDiarmid, R. W. (1994): Preparing amphibians as scientific specimens. – pp. 289–296 in: Heyer, W. R., M. A. Donnelly, R. W. McDiarmid, L.-A. C. Hayek & M. S. Foster (eds): Measuring and Monitoring Biological Diversity: Standard Methods for Amphibians. – Smithsonian Institution Press, Washington. Montero-Mendieta, S., I. De la Riva, I. Irisarri, J. A. Leonard, M. T. Webster & C. Vilà (2021): Phylogenomics and evolutionary history of Oreobates (Anura: Craugastoridae) Neotropical frogs along elevational gradients. – Molecular Phylogenetics and Evolution, 161(107167): 1–13. 440 Ernesto Castillo-Urbina et al. Morales, V. R. & J. Icochea (2000): Review of the type material of Eleutherodactylus mendax and a new record of Eleutherodactylus bromeliaceus from Peru. – Journal of Herpetology, 34: 158–160. Motta, A. P., J. C. Chaparro, J. P. Pombal, Jr., J. M. Guayasamin, I. De la Riva & J. M. Padial (2016) Molecular phylogenetics and taxonomy of the Andean genus Lynchius Hedges, Duellman, and Heinicke, 2008 (Anura: Craugastoridae). – Herpetological Monographs, 30: 119–142. Myers, C. W. & J. W. Daly (1979): A name for the poison frog of Cordillera Azul, eastern Peru, with notes on its biology and skin toxins (Dendrobatidae). – American Museum Novitates, 2674: 1–24. Padial, J. M., J. C. Chaparro, S. Castroviejo-Fisher, J. M. Guayasamin, E. Lehr, A. J. Delgado C., M. Vaira, M. Teixeira Jr., C. R. Aguayo-Vedia & I. De la Riva (2012): A revision of species diversity in the Neotropical genus Oreobates (Anura: Strabomantidae), with the description of three new species from the Amazonian slopes of the Andes. – American Museum Novitates, 3752: 1–55. Padial, J. M., J. C. Chaparro & I. De la Riva (2008): Systematics of Oreobates and the Eleutherodactylus discoidalis species group (Amphibia, Anura), based on two mitochondrial DNA genes and external morphology. – Zoological Journal of the Linnean Society, 152: 737–773. Padial, J. M., T. Grant & D. R. Frost (2014): Molecular systematics of terraranas (Anura: Brachycephaloidea) with an assessment of the effects of alignment and optimality criteria. – Zootaxa, 3825: 1–132. Palumbi, S. R., A. Martin, S. Romano, W. O. McMillan, L. Stice & G. Grabowski (1991): The Simple Fool’s Guide to PCR. Version 2.0. – Privately published, Univ. Hawaii. Pansonato, A., A. Motta, P. Cacciali, C. F. B. Haddad, C. Strüssmann & M. Jansen (2020): On the identity of species of Oreobates (Anura: Craugastoridae) from central South America, with the description of a new species from Bolivia. – Journal of Herpetology, 54: 393–412. Pyron, R. A. & J. J. Wiens (2011): A large-scale phylogeny of Amphibia including over 2800 species, and a revised classification of extant frogs, salamanders, and caecilians. – Molecular Phylogenetics and Evolution, 61: 543–583. Rakotoarison, A., A. Crottini, J. Müller, M.-O. Rödel, F. Glaw & M. Vences (2015): Revision and phylogeny of narrow-mouthed treefrogs (Cophyla) from northern Madagascar: integration of molecular, osteological, and bioacoustic data reveals three new species. – Zootaxa, 3937: 61–89. Salzburger, W., G. B. Ewing & A. von Haeseler (2011): The performance of phylogenetic algorithms in estimating haplotype genealogies with migration. – Molecular Ecology, 20: 1952–1963. Shanee, N., S. Shanee & R. H. Horwich (2015): Effectiveness of locally run conservation initiatives in north-east Peru. – Oryx, 49: 239–247. Shanee, S., N. Shanee, W. Loc & M. J. Espejo-Uribe (2020): The development and growth of non-governmental conservation in Peru: Privately and communally protected areas. – Human Ecology, 48: 681–693. Shimodaira H. & M. Hasegawa (1999): Multiple comparisons of log-likelihoods with applications to phylogenetic inference. – Molecular Biology and Evolution, 16: 1114–1116. Stephens, M., N. J. Smith & P. Donnelly (2001): A new statistical method for haplotype reconstruction from population data. – American Journal of Human Genetics, 68: 978–989. Tamura K., G. Stecher & S. Kumar (2021): MEGA11: Molecular Evolutionary Genetics Analysis Version 11. – Molecular Bio logy and Evolution, 38: 3022–3027. Trifinopoulos J., L. T. Nguyen, A. von Haeseler & B. Q. Minh (2016): W-IQ-TREE: A fast online phylogenetic tool for maximum likelihood analysis. – Nucleic Acids Research, 44(W1): W232–W235. Twomey, E., J. Delia & S. Castroviejo-Fisher (2014): A review of northern Peruvian glassfrogs (Centrolenidae), with the description of four new remarkable species. – Zootaxa, 3851: 1–87. Vaz-Silva, W., N. M. Maciel, S. P. de Andrade & R. C. Amaro (2018): A new cryptic species of Oreobates (Anura: Craugastoridae) from the seasonally dry tropical forest of central Brazil. – Zootaxa, 4441: 89–108. Vences, M., A. Miralles, S. Brouillet, J. Ducasse, A. Fedosov, V. Kharchev, I. Kostadinov, S. Kumari, S. Patmanidis, M. D. Scherz, N. Puillandre & S. S. Renner (2021): iTaxoTools 0.1: Kickstarting a specimen-based software toolkit for taxonomists. – Megataxa, 6: 77–92. Vences, M., S. Patmanidis, J.-C. Schmidt, M. Matschiner, A. Miralles & S. S. Renner (2024): Hapsolutely: a user-friendly tool integrating haplotype phasing, network construction and haploweb calculation. – Bioinformatics Advances, 4: vbae083. Venegas, P. J., L. A. García-Ayachi, J. Ormeño, S. Bullard, A. Catenazzi & A. P. Motta (2021): Two new species of terrestrial-breeding frogs (Anura: Brachycephaloidea) from Cordillera de Colán, Peru. – Neotropical Biodiversity, 7: 279–296. Vieites, D. R., M. S. Min, D. B. Wake (2007): Rapid diversification and dispersal during periods of global warming by pletho dontid salamanders. – Proceedings of the National Academy of Sciences of the USA, 104: 19903–19907. Supplementary data The following data are available online: Supplementary Figure S1. Map of forest cover loss within the Biocorredor Bosques de Vaquero ‘Shunku Sacha’. Appendix Additional specimens examined Oreobates choristolemma: BOLIVIA: La Paz: Serranía de Bella Vista, CBF 5611 (holotype). Oreobates cruralis: BOLIVIA: Cochabamba: El Palmar, 1300m a.s.l., Parque Nacional Carrasco, ZFMK 72570; between Paractito and El Palmar, ZFMK 66964 ZFMK 66971–66972, ZFMK 72541– 72543; Los Guácharos (Chapare, 500 m a.s.l.), ZFMK 72532; La Paz: La Paz (locality in error), BMNH 1947.2.15.70 (holotype); road from Caranavi to Palos Blancos, ZFMK 80599; Santa Cruz: road to Bellavista near road to Samaipata, ZFMK 71997; La Hoyada, 1800 m a.s.l., Parque Nacional Amboró, ZFMK 72644; south of Cuevas, 1100 m a.s.l., ZFMK 72644. Oreobates discoidalis: ARGENTINA: Tucumán: Horco Molle, “13 km W of Tucumán,” Sierra de San Javier, ca. 1200 m a.s.l., BMNH 1947.2.15.63–65 (syntypes). 441 New insights into the diversity of Oreobates frogs Oreobates granulosus: PERU: Puno: Santo Domingo, Carabaya, 6000 ft (1800 m a.s.l. aprox.), BMNH 1947.2.15.72 (holotype). Oreobates ibischi: BOLIVIA: Santa Cruz: km 68.5 on Santa Cruz de la Sierra-Samaipata road, 750 m a.s.l., CBF 3341 (holotype); El Fuerte de Samaipata, 1900 m a.s.l., ZFMK 60402 (paratype). Oreobates lundbergi: PERU: Pasco: road Puagmaray to Oxapampa km 77, 2550 m a.s.l., MUSM 19321 (paratype), Paucartambo, MUSM 18424 (paratype). Oreobates pereger: PERU: Ayacucho: Yuraccyacu, Tambo– Valle del Apurímac trail, 2650 m a.s.l., MUSM 13980 (paratype; formerly LSUMZ 26120). Oreobates quixensis: ECUADOR: Napo: San José de Moti, MNCN 1708 (lectotype; photos only); PERU: Huánuco: Panguana, MUSM 40340, ZSM 176/2017. Oreobates sanctaecrucis: BOLIVIA: Cochabamba: Karahuasi, 2200 m a.s.l., ZFMK 72647; Santa Cruz: El Chapé, Parque Nacional Amboró, 2060 m a.s.l., MNK-A 1198 (holotype). Oreobates sanderi: BOLIVIA: La Paz: Colonia Eduardo Avaroa, ca. 30 km north of Caranavi on the road from Caranavi to Yucumo, ZFMK 80600–80601 (paratypes). Oreobates saxatilis: PERU: San Martín: Pongo de Shilcayo, about 4 km NNW of Tarapoto, 470 m, KU 212556 (holotype), KU 212327 (paratype; photos only); 27 km NE of Tarapoto, MUSM 8431–8432 (paratypes); Huánuco: Panguana, ZSM 1973– 1978/2008, ZSM 778–779/2010, ZSM 175/2017. Oreobates yanucu: BOLIVIA: Cochabamba: old Chapare road, 1500 m a.s.l., ZFMK 72569 (holotype). Oreobates zongoensis: BOLIVIA: La Paz: Valle de Zongo, 1250m a.s.l., CBF 2503 (holotype).