Fifty years after: A taxonomic revision of the amphibian species from the Ecuadorian biodiversity hotspot Abra de Zamora, with description of two new Pristimantis species

described, such as Parque Nacional Yasunı´ in western Amazon [5] or Parque Nacional Sangay [6] and the upper basin of the Pastaza River [7] in Tropical Andes. One such a biodiverse hot- spot, with a comparatively much smaller extension, is Abra de Zamora. The name “Abra de Zamora” was used by William E. Duellman and John D. Lynch in their ’70s publications [8, 9], to designate the “ridge (or crest) between Loja and Zamora Chinchipe provinces in the Cordillera Oriental in southern Ecuador” or just the area “13–15 kilometers east (by road) of Loja City, Ecuador”. The term Abra is used in orography to refer to the depression in a line of mountain peaks, which generally, for logistical reasons, is used to build the roads that cross the mountain range [10]; this is also the case of the road connecting two Ecuadorian cities from the tropical Andean region, Loja and Zamora. However, this name was not used by the local residents, who refer to the area between the two provinces closer to Loja as “El Tiro” (the Tiro Pass), and to the lower parts, close to El Tambo sector as “Los Trigales” (the Wheat Fields). Duellman and Lynch conducted their pioneer expeditions in search of amphibians on the old road that connected Loja with Zamora at that time (or Vı´a Antigua Loja-Zamora); this road was used only until the mid ’80s, when a new road (the E50 highway) was constructed on the other side of the valley, and is still in use today (Figs 1 and 2A). Between 1938 and 2010, eleven species of frogs were described from Abra de Zamora. The

first species, Lynchius flavomaculatus, was described in 1938 by Hampton Wildman Parker [11], with specimens collected by the Ecuadorian naturalist, Clodoveo Carrio´n Mora. In 1974, William E. Duellman described Gastrotheca psychrophila [8], based on specimens collected in 1968 and 1971. John D. Lynch described six Pristimantis species from Abra de Zamora in 1979 [9], with specimens collected in 1968, 1971 and 1975: P. balionotus, P. colodactylus, P. crypto- melas, P. percultus, P. versicolor and P. vidua. Linda Trueb described Telmatobius cirrhacelis in 1979, using specimens collected in 1975 [12]. Paratypes collected from Abra de Zamora were used for the description of an additional two species: P. andinognomus [13], with specimens collected in 2003, and Atelopus podocarpus [14], with specimens collected in 1968 by Lynch and 1975 by Duellman. Due to its location (Huancabamba depression, Andes), Abra de Zamora has been a center of endemism and probably a center of amphibian diversification [15]. From this list of eleven species originally described from this area, six species are strictly endemic to the area. An addi- tional five species are currently undescribed, and similarly have not yet been encountered outside the area. This information emphasizes the importance of the region as a center of endemism, and suggests its role as a center of amphibian diversification, particularly for some groups of Terrarana. The advancement of molecular techniques allows precise species identifi-

cation, which is especially important in areas of diversification and for cryptic species [16]. Unfortunately, many of these species were not encountered after their original description, and DNA samples were available only for a couple of them (G. psychrophila, P. andinognomus, P. cryptomelas and P. versicolor) which made their phylogenetic relationships impossible to assess. After almost 50 years of the important work of Duellman and Lynch we evaluated the presence and conservation status of the amphibian populations from this important hotspot. Through an intense fieldwork encompassing Abra de Zamora and several other potentially similar habitats in southern Ecuador, our study targets the following aspects: i. redescribe the species which were described from Abra de Zamora, including their genetic delimitation and advertisement call; ii. present the updated amphibian species list of Abra de Zamora with the description of two additional Pristimantis species; iii. search for their presence in similar mon- tane habitats in surrounding areas from southern Ecuador, in an attempt to more precisely evaluate their distribution; and iv. update recommendations regarding their conservation status. PLOS ONE Taxonomic revision of the amphibian species from Abra de Zamora PLOS ONE | https://doi.org/10.1371/journal.pone.0238306 September 10, 2020 2 / 59 Marsupial Frog (Gastrotheca psychrophila) in southern Ecuador". DS benefited from a research grant from Universidad Te´cnica Particular de Loja (Convocatoria Estancias Cortas de Investigacio´n 2019).

Competing interests: The authors have declared that no competing interests exist.