Phylogenetic Analysis Whithin the Pristimantis Unistrigatus (Anura, Craugastoridae) Group Based on Morphological Characters

Home , Craugastoridae, Craugastor raniformis, Pristimantis bogotensis, Pristimantis frater, Pristimantis lynchi, Pristimantis pugnax

Zoología-Sistemática http://www.icn.unal.edu.co/ Caldasia 36(1):107-124.Hoyos et2014 al. PHYLOGENETIC ANALYSIS WHITHIN THE PRISTIMANTIS UNISTRIGATUS (ANURA, CRAUGASTORIDAE) GROUP BASED ON MORPHOLOGICAL CHARACTERS Análisis filogenético en el grupoPristimantis unistrigatus (Anura, Craugastoridae) basado en caracteres morfológicos

Julio Mario Hoyos Catalina Mantilla Pontificia Universidad Javeriana, Departamento de Biología, Unidad de Ecología y Sistemática (UNESIS), Apartado 56710, Bogotá Colombia. [email protected], [email protected]

Diana Galindo Universidad de los Andes, Departamento de Biología, Bogotá, Colombia. dmgalindo48@ uniandes.edu.co

Lina salgar Pontificia Universidad Javeriana, Departamento de Biología, Unidad de Ecología y Sistemática (UNESIS), A.A. 56710, Bogotá, Colombia. [email protected]

ABSTRACT We present a phylogenetic analysis within the Pristimantis unistrigatus group (Anura, Craugastoridae) of Colombia. Characters from the superficial muscles of the hands and feet as well as external characters were taken for analysis. Most of the muscle characters were observed directly, and some were taken from the literature. Similarly, the external ones were taken mostly from the original descriptions and others from the literature as well. Two matrices were constructed, as the species belonging to this group have changed in recent years with respect to the initially proposed when the group was defined. The results lead us to conclude that the group is not monophyletic, although there are some relationships that are worth to survey because they are kept in the very last cladograms obtained for both proposals. It is suggested that these last relationships should be explored in particular, and the overall group in general, increasing the number of characters and taxa that belong to P. unistrigatus. An open question we left is whether actually is worth to keep these informal taxonomic hierarchy called group within the genera of anurans.

Key words. Neotropical frogs, Cordillera Oriental, superficial muscles, Pristimantis.

RESUMEN Se presenta un análisis filogenético del grupo Prsitimantis unistrigatus (Anura, Craugastoridae) de Colombia. Para esto se tomaron caracteres de la musculatura superficial de manos y pies y caracteres externos. La mayoría de los caracteres musculares fueron observados directamente, y algunos fueron tomados de la literatura. Igualmente, los externos fueron tomados en su mayoría de las descripciones originales, 107 Phylogenetic analysis whithin the Pristimantis unistrigatus

y otros tomados de la literatura también. Se construyeron dos matrices, pues las especies que pertenecen al grupo han cambiado en los últimos años con respecto a lo propuesto inicialmente cuando el grupo fue definido. Los resultados nos llevan a concluir que el grupo no es monofilético, aunque existen algunas relaciones que vale la pena explorar, pues se mantienen en los cladogramas definitivos obtenidos para ambas propuestas. Se sugiere que deben explorarse más estas relaciones en particular, y la del grupo en general, aumentando los caracteres y el número de taxones que pertenecen a P. unirtsigatus. Igualmente se deja como interrogante si en realidad vale la pena mantener estos grupos informales dentro de los géneros de anuros.

Palabras clave. Ranas neotropicales, Cordillera Oriental, músculos superficiales, Pristimantis.

INTRODUCTION hypotheses of relationships in Colombia, because it is the richest in species number Frost et al. (2006) doubted the existence of the in this country. From the perspective of genus Eleutherodactylus Duméril and Bibron, phylogenetic systematics of the genus 1841 as monophyletic, and suggested that it Pristimantis, the work that has a growing should be divided into many monophyletic number of Colombian species for the genera. Heinicke et al. (2008) divided the unistrigatus group (sensu Lynch & Duellman former genus Eleutherodactylus plus the genus 1997) of the Cordillera Oriental of Colombia Brachycephalus Fitzinger, 1826 in three Clades: has been that of Galindo (2004), based on the South American Clade, the Caribbean external characters. However, the papers of Clade and the Middle American Clade. The Duellman & Simmons (1977), Lynch (1973, Eleutherodactylus unistrigatus group was 1975a, 1975b, 1978, 1981, 1983, 1984, 1994a, included in the South American Clade using 1994b, 1998, 2003), Lynch & Duellman the available name Pristimantis Jiménez de la (1980), Lynch & Suárez (2002 ) and Pyburn Espada, 1870. This group is one of the richest & Lynch (1981) have revealed potential Pristimantis groups, with members occurring relationships for species of the unistrigatus in the Andes and the Amazonian basin, ranging group, some with formal cladograms, but from Bolivia to Colombia (Hedges et al. 2008, the majority without them. Lynch (1984) Padial et al. 2009). proposed a hypothesis of relationship for the unsitrigatus group without including any of In the literature related to the diagnosis of the the species used here, but some belonging unistrigatus group species of the Cordillera to the “assemblages” pyrrhomerus and Oriental of Colombia, potential kinship myersi, suggesting these should be combined and characteristics in common are referred by a series of synapomorphies found in (Duellman & Simmons 1977, Lynch1973, external characters. Finally, recently Padial 1978, 1981, 1983, 1984, 1994a, 1994b, et al. (2009) considered that the Pristimantis 1998, 2003, Lynch & Duellman1980, Lynch unistrigatus group is not monophyletic. & Duellman 1997, Lynch & Suarez 2002, Pyburn & Lynch1981). However, there have Other groups, which have launched hypotheses been no formal relationship hypotheses of relationship, either with or without explicit supported by cladograms. cladograms are: discoidalis (Lynch 1989) (with explicit cladogram), using external It is perhaps for this Pristimantis group and cranial characters, and sulcatus (Lynch that were proposed the highest number of 1997) (with explicit cladogram), finding for 108 Hoyos et al. these relationships nine characters from the analyses are brief, for the first time, possible skull. As this paper aims to use morphology synapomorphies for the genus Pristimantis as a fundamental element to establish kinship (intrageneric relationships) based on the hypothesis, the discussion will not be based on muscles of hands and feet were revealed. specific morphological differences between Lynch (1984) proposed some characteres what we found and what other authors have but for relationships between species groups, found, but between the relationships proposed based on osteological characters and some of here based on potential synapomorphies, the muscles (but not hands and feet). and other hypotheses and characters used to construct them. Hedges et al. (2008) redefined thePristimantis unistrigatus group changing the species From morphology’s point of view, the included to it, assigning 193 species to contributions are all innovative in the sense this group, changing some of the species that no previous research has been dedicated previously identified into the group by Lynch solely to the study of muscles of hands (1976a, Lynch & Duellman 1997, Duellman and feet for the phylogenetic analysis of & Pramuk 1999), and by Lynch & Duellman the Pristimantis genus. Similarly, although (1997) and Acosta (2000) for Colombia. there are some previous descriptions about hand muscles of some species of the genus Taking into account that there are two (e.g.Burton 1998a, Dunlap 1960), these were different classifications for the Pristimantis mainly for comparative purposes (Dunlap unistrigatus group, our aim is to compare 1960, Salgar et al. 2009), or with the aim our results in using morphological characters to find characters to establish hypotheses of from hand and foot muscles in both proposals, relationships (Burton 1998a) but there was not because there are not enough consensus a genus-specific analysis performed. about which is the best classification. It is for this reason that both the number of species Previous studies in the literature, using both and characters in both analyses differs (see muscles of hands and forearms and feet to results). The only classification that include propose phylogenetic relationships in frogs are all species of our study is that by Lynch & very rare; in fact, there is only one (Faivovich Duellman (1997). The number of species 2002), but in the genus Scinax (Hylidae). in classification by Hedges et al. (2008) is The most comprehensive descriptive work lower, because they do not consider some of in this regard have been those of Gaup the species by Lynch & Duellman (1997) as (1886), Dunlap (1960), Andersen (1978) belonging to the P. unistrigatus group. Owing and, more recently, Burton (1996,1998a,b, to this account, the number of characters also 2001, 2004). Most of them propose (tacitly changed. In comparing our results with those or explicitly) some assumptions about the by Heinicke et al. (2008), and Hedges et al. potential systematic value of characters taken (2008), we can conclude that they agree that from hands and feet in anuran amphibians, P. unistrigatus group is not monophyletic. without reaching cladistic analysis themselves. Only in one of his most recent work, Burton MATERIALS AND METHODS (2004) developed a cladistic analysis using only the muscles of the feet in the family A total of 42 adult specimens (Table 1) Hylidae. The only existing descriptions of this representing 17 species of Pristimantis musculature in Colombian species of frogs of the Cordillera Oriental of Colombia has been conducted by Salgar (2003) and and one of Craugastor were dissected. Salgar et al. (2009). Although these last two Specimens belong to the Museo de Historia 109 Phylogenetic analysis whithin the Pristimantis unistrigatus

Natural (MUJ) of the Pontificia Universidad Table 1. List of species. M: Male. F: Javeriana in Bogotá, Colombia, and to the Female. Instituto de Ciencias Naturales (ICN) of Specimen Sex Species the Universidad Nacional de Colombia in MUJ 36 M P. elegans Bogotá. Taking into account the species MUJ 052 F P. bogotensis MUJ 465 M P. bogotensis belonging to the unistrigatus group sensu MUJ 481 M P. bogotensis Hedges et al. (2008), we decided to compare MUJ 687 F P. bogotensis results considering the former classification MUJ 23 F P. nervicus made by Lynch & Duellman (1997).We chose MUJ 1176 F P. nervicus the following two ingroups: sensu Lynch & MUJ 1507 F P. lynchi MUJ 2036 F P. lynchi Duellman (1997) (LD): Pristimantis anolirex MUJ 2420 M P. taeniatus (Lynch, 1983): ICN 33522, 15444; P. bacchus MUJ 2452 F P. taeniatus (Lynch, 1984): ICN 33164, 33170, 34232; MUJ 1480 F P. lynchi P. bicolor ( Rueda-Almonacid and Lynch, MUJ 1538 M P. lynchi 1983): ICN 14474, 26321; P. bogotensis ICN 33522 F P. anolirex ICN 15444 F P. anolirex (Peters, 1863): MUJ 052, 465, 481, 687; P. ICN 26321 M P. bicolor elegans (Peters, 1863): MUJ 036; P. frater ICN 50062 F P. miyatai (Werner, 1899): ICN 40579, 49911; P. ICN 50065 M P. miyatai jorgevelosai (Lynch, 1994): ICN 15299, ICN 50069 F P. miyatai ICN 33170 F P. bacchus 15318; P. lynchi (Duellman and Simmons, ICN 34232 M P. bacchus 1977): MUJ 1480, 1538, 1507, 2036; ICN ICN 33164 M P. bacchus 01932, 01937; P. merostictus (Lynch, 1984): ICN 12445 M P. spilogaster ICN 34233, 34235; P. miyatai (Lynch, 1984): ICN 12446 F P. spilogaster ICN 50062, 50065, 50069; P. nervicus ICN 1932 F P. lynchi ICN 1937 F P. lynchi (Lynch, 1994): MUJ 023,1176; P. nicefori ICN 14474 F P. bicolor (Cochran and Goin, 1970): ICN 20994, ICN 40579 M P. frater 21119; P. prolixodiscus (Lynch, 1978): ICN ICN 49911 M P. frater 10104, 10107; P. pugnax (Lynch, 1973): ICN ICN 15299 M P. jorgevelosai 22981, 23187; P. spilogaster (Lynch, 1984): ICN 15318 M P. jorgevelosai ICN 10107 M P. prolixodiscus ICN 12445, 12446; P. taeniatus (Boulenger, ICN 10104 F P. prolixodiscus 1912): MUJ 2452, 2420; P. tamsitti (Cochran ICN 22981 F P. pugnax and Goin, 1970): ICN 22949, 22951, 23638; ICN 23187 F P. pugnax Sensu Hedges et al. (2008) (HEA): P. anolirex ICN 21119 M P. nicefori (Lynch, 1983), P. bacchus (Lynch, 1984), P. ICN 20994 F P. nicefori ICN 22949 F P. tamsiti bogotensis (Peters, 1863) , P. elegans (Peters, ICN 22951 F P. tamsiti 1863), P. lynchi (Duellman and Simmons, ICN 23638 F P. tamsiti 1977), P. merostictus (Lynch, 1984), P. ICN 34233 F P. merostictus nervicus (Lynch, 1994), P. nicefori (Cochran ICN 34235 F P. merostictus and Goin, 1970), P. pugnax (Lynch, 1973), P. spilogaster (Lynch, 1984), P. taeniatus The new classification proposed by Hedges (Boulenger, 1912), P. tamsitti (Cochran and et al. (2008) for some of the species formerly Goin, 1970) . assigned to unistigatus group is as follows: P. frater: frater group; P. bicolor: species series Outgroups: Craugastor raniformis: MUJ P. (Hypodiction) ridens; P.miyatai: P. frater 2871, 2689 for the LD matrix, and P. frater group; P. jorgevelosai: species series P. for the HEA matrix (Tables 2 and 3). (Hypodiction) ridens; and P. prolixodiscus: lacrimosus group. 110 Hoyos et al.

Muscles External characters

We observed about 90 small hand and These were taken from taxonomic feet muscles, both superficial and deep, of descriptions after Cochran & Goin (1970), which about 50 superficial were chosen. Lynch (1983), Lynch (1984), Rueda & Lynch For choosing the characters and states, we (1983), Dunn (1944), Hoyos (1991), Pyburn did comparisons between muscles from & Lynch (1991), Lynch (1994), Duellman ten species of Pristimantis used for the & Simmons (1997), Lynch (1998), Lynch study with respect to different aspects that (1978), and Lynch & Duellman (1980), and are related below. Dissections were done after our own observations. in hands, feet, and legs and forearms in order to examine those muscles associated Phylogenetic analysis morphologically with the hands and feet. We used a dissecting microscope and a For the phylogenetic analysis, we compared reversible Weigert’s iodine solution that each result taking into account two matrices stains muscles (Bock & Shear1972) to see (see below). We did heuristic searches in the direction of the fibers, obtaining a clear TNT, performing TBR branch swapping differentiation of muscles and tendons, on 100 addition replicates. Matrices were including the origins and insertions. For running considering characters with the same all specimens both right and left hands weight. We did not find many intraspecific and feet superficial muscles were dissected (polymorphic) variations or many multistate and examined, following the protocols characters, however we use all of those proposed by Raikow et al. (1990). For each we found for the phylogenetic analyses. muscle we identified: a. origin; b. insertion; Polymorphic characters were coded according c. spatial arrangement with respect to the to Faivovich (2002) as follows: A=0/1; B=0/2, hand and foot, and d. spatial relationship and C=1/2, and multistate characters were all with adjacent muscles. Although we considered nonadditive and unordered. We did explored mainly superficial musculature, not get whether retention (RI) or consistency we also dissected some deep muscles indices (CI), given that we are comparing considering that we took into account groups of different sizes (e.g. Conrad 2008), so characters proposed by Burton (1998a, the value for the CI will be greater in the HEA 2004). The nomenclature was taken from ( Sensu Hedges et al., 2008) group than in the that proposed by Dunlap (1960), Andersen LD (sensu Lynch & Duellman, 1997) group (1978), Burton (1998a, 2004), and Salgar because we considered less species in the first et al. (2009). We used for most cases, at group than in the second one, and the RI will least one male and one female of each be less in the HEA for the same reason. species to the extent that the number of specimens of the collection allowed We choose one outgroup species for each it, to determine the existence of sexual analysis, following recommendation by dimorphism. Finally, we drew pictures Nixon & Carpenter (1993: 419) about the of characters considered as potential fact that “…a third pervasive myth, the no- synapomorphies using photographs tion that more than one outgroup is somehow and the Adobe Illustrator software. required for correct polarity determination”, For the accuracy of the origin and insertion and taking into account that “Outgroups and of muscles, we made use of clearing and polarity are the same problem” (Nixon Nixon staining skeletons of P. bogotensis. & Carpenter 1993: 420). We only show the unambiguous synapomorphies. 111 Phylogenetic analysis whithin the Pristimantis unistrigatus 0 0 0 1 0 0 0 1 0 0 0 0 0 0 0 1 0 0 2 0 1 1 0 1 0 0 0 1 0 0 0 0 0 0 0 1 0 0 A 2 0 1 0 A 0 0 12 0 0 0 0 A 1 1 2 A 1 0 0 0 1 1 10 0 0 A 0 1 1 1 1 0 0 1 0 0 1 0 0 1 A 0 0 A1 1 0 0 1 0 A1 0 1 0 A1 A 1 0 0 2 1 0 0 0 0 0 0 0 0 0 1 0 0 1 0 1 1 0 A 0 1 2 0 0 1 0 1 0 1 A 0 0 2 0 1 1 1 0 0 1 0 1 0 0 1 0 1 1 1 0 1 0 1 1 0 0 0 0 1 1 1 0 0 1 0 0 10 0 0 2 0 1 1A 1 1 0 1 0 1 0 1 0 0 0 0 0 1 1 0 0 0 1 0 0 1 0 1 0 1 2 0 0 1 0 1 0 1 1 0 0 0 1 1 0 1 0 1 0 0 1 0 0 0 0 0 0 1 0 0 0 0 0 1 1 1 0 0 1 0 0 1 0 0 0 1 1 0 2 0 1 1 0 1 0 1 0 0 0 0 0 0 0 0 0 1 0 0 0 0 0 1 0 1 0 1 2 0 0 00 0 0 1 1 0 2 0 1 1 0 10 1 0 1 1 0 0 0 0 0 0 10 0 0 0 0 1 1 1 0 0 11 0 0 0 1 1 1 0 0 2 1 1 1 0 1 0 0 1 10 0 0 1 0 0 0 0 0 1 1 1 0 1 0 1 1 1 2 0 0 1 0 1 0 1 0 0 0 0 1 1 1 1 0 1 0 1 0 0 0 1 0 1 0 1 0 0 0 0 0 1 0 0 0 1 2 0 0 0 0 1 0 1 1 0 0 0 1 1 0 1 1 0 1 1 1 0 0 0 0 1 0 0 0 0 1 0 1 1 0 1 0 1 2 0 0 1 0 1 0 1 0 0 1 0 1 1 0 0 0 1 0 1 0 0 0 0 0 0 0 1 0 0 0 0 1 1 1 1 0 1 0 0 1 00 0 0 0 0 0 0 0 1 1 0 10 0 0 1 0 0 0 1 0 0 0 00 1 1 0 1 0 1 1 0 01 1 1 1 1 0 1 1 0 1 0 0 1 1 1 1 0 0 0 1 1 1 0 1 0 1 0 0 0 0 1 0 0 1 0 1 0 1 0 0 0 0 0 1 0 1 0 0 1 0 1 1 0 1 0 2 0 1 0 0 0 0 0 0 0 1 0 0 1 0 0 1 0 1 1 0 1 0 0 10 1 1 1 1 0 2 0 1 1 0 00 1 2 1 0 0 0 1 0 1 0 10 0 1 0 0 1 1 1 1 1 0 0 0 1 0 0 0 1 1 0 0 0 2 1 0 1 0 ? 0 10 0 0 0 0 1 0 1 0 0 1 0 A 1 1 0 0 0 0 0 0 0 0 A 1 0 1 1 0 2 0 0 0 1 0 0 0 1 2 1 0 0 0 0 1 1 0 1 0 0 species group. 1 0 0 1 0 1 1 0 1 0 1 1 0 1 0 1 0 1 0 1 0 1 0 1 0 1 0 1 1 0 1 0 1 species group.

0 1 0 1 0 1 0 1 0 1 1 0 0 1 0 1 0 1 0 1 0 C 1 0 1 0 1 0 0 1 0 1 0 1 1 0 1 A 1 0 P. unistrigatus P. P. unistrigatus P. 1 0 1 0 0 1 0 0 1 0 1 0 0 1 0 1 0 1 0 0 1 1 0 0 1 0 0 0 1 0 1 0 1 1 0 0 0 0 1 0 1 0 0 0 1 0 0 0 1 0 1 0 0 0 1 0 0 1 1 0 1 0 0 1 0 1 1 0 0 1 0 1 0 1 0 1 0 0 1 1 0 0 1 1 0 1 0 0 1 1 0 0 1 1 0 0 1 0 1 0 1 B 0 1 0 1 0 0 1 0 0 0 1 0 1 0 0 0 1 0 0 1 0 1 0 1 0 2 0 1 1 0 0 1 A 1 2 A 1 0 A 0 A 1 0 0 1 2 1 0 A 1 0 A A 1 0 A A 1 0 0 1 1 0 0 1 1 0 1 A A 1 0 B 0 1 0 1 0 0 0 0 0 0 0 0 0 0 0 0 1 11 1 1 01 1 0 A 0 1 2 A 1 01 0 1 0 B 11 A A 1 0 1 0 1 0 0 1 2 0 1 0 1 1 0 2 11 A 0 1 10 1 0 1 1 0 2 A 1 0 1 2 0 1 1 0 0 1 01 10 2 0 0 0 A A 0 0 1 2 A 1 0 1 0 2 1 1 0 1 0 0 0 0 0 1 0 0 1 0 1 0 1 0 1 1 0 1 1 0 1 0 0 0 10 1 A 2 0 1 0 A 0 1 2 A 2 0 1 A 1 0 10 2 1 0 2 0 1

2 0 2 0 1 2 0 2 0 1 2 0 1 A 2 0 2 0 1 0 1 0 1 0 2 0 0 1 0 2 0

2 0 2 0 1

Data matrix employed in the cladistic analysis of LD Data matrix employed in the cladistic analysis of the HEA Data matrix employed in the cladistic analysis of HEA Table 2. Table Craugastor raniformis Pristimantis taeniatus Pristimantis tamsitti Pristimantis pugnax Pristimantis bacchus Pristimantis miyatai Pristimantis merostictus Pristimantis spilogaster Pristimantis anolirex Pristimantis jorgevelosai Pristimantis nicefori Pristimantis bicolor Pristimantis frater Pristimantis prolixodiscus Pristimantis bogotensis Pristimantis elegans Pristimantis nervicus Pristimantis lynchi 3. Table Pristimantis frater Pristimantis taeniatus Pristimantis tamsitti Pristimantis pugnax Pristimantis bacchus Pristimantis merostictus Pristimantis spilogaster Pristimantis anolirex Pristimantis nicefori Pristimantis bogotensis Pristimantis elegans Pristimantis nervicus Pristimantis lynchi 112 Hoyos et al.

Results leaving away P. nicefori, and the second one is a character shared by P. prolixodiscus and We scored 81 characters, from which 64 are P. bicolor. muscle characters (44 from the manus and 20 from the pes), and 17 external characters. Four HEA P. unistrigatus group (Appendix 2) manus characters were taken from Burton (1998a), and five pes characters from Burton We took 60 characters for the analysis (2004). Based on this matrix, we eliminated (Appendix 2). The cladistic analysis resulted those characters according to changes in the in 20 most-parismonious trees of 139 steps. taxa for each matrix as we describe below. The strict consensus tree (Fig. 3) shows a node 14 not resolved, consisting of taxa P. lynchi, P. Phylogenetic analysis nervicus, P. elegans, P. bogotensis, P. nicefori, P. anolirex, P. bacchus, and P. taeniatus. The LD P. unistrigatus group only unambiguous putative synapomorphy (found in all trees) is the character state 44(1) By eliminating those characters with only one (Origin of the tendo superficialis hallucis character state present in the outgroup and from the aponeurosis plantaris including ingroup, we took 70 from the 81 characters some fibers of the m. lumbricalis brevis (Appendix 1). The cladistic analysis resulted hallucis) (Fig. 4) relating P. spilogaster and in one fully resolved most-parismonious P. merostictus. tree (Fig. 1) with a length of 202. We found character states 62(0) (First finger shorter DISCUSSION than second) and 23(1) (The origin of the m. adductor policis is not covered by the As shown in the cladograms obtained for both aponeurosis palmaris) (Fig. 2) as the only unistrigatus groups, the LD (Fig. 1) group is two unambiguous putative synapomorphies. more resolved than the strict consensus of the The first one included almost all species HEA (Fig. 3), but there are no synapomorphies

a b

Figure 1. The only most-parsimonious cladogram of LD P. unistrigatus group. a. Tree without showing nodes numbers, and showing the character states 23 (1) and 62 (0) as unambiguous synapomorphies. b. Tree showing nodes numbers.

Abreviations: Cran: Craugastor raniformis, Pano: Pristimantis anolirex, Pbac: P. bachus, Pbic: P. bicolor, Pbog: P. bogotensis, Pele: P. elegans, Pfra: P. frater, Pjor: P. jorgevelosai, Plyn: P. lynchi, Pmer: P. merostictus, Pmiy: P. miyatai, Pner: P. nervicus, Pnic: P. nicefori, Ppro: P. prolixodiscus, Ppug: P. pugnax, Pspi: P. spilogaster, Ptae: P. taeniatus, Ptam: P. tamsitii. 113 Phylogenetic analysis whithin the Pristimantis unistrigatus that support it, except partially for nodes 21 the proposed LD unistrigatus group can be and 32 in LD, and exclusively for node16 in considered stronger if we take the fact that HEA, this means that all topologies are weakly we have got a single tree completely resolved; supported by synapomorphies. In node 22 of DL no synapomorphies were found, whereas a for the other nodes, potential synapomorphies are actually reversals or convergences.

Taking into account the results and species considered by HEA and LD, we could say that the assigned HEA unistrigatus group is weaker because the relationships are less resolved, even if the number of steps is smaller (it has a smaller number of taxa). On the other hand, even with a larger number of species involved,

Figure 3. Strict consensus of the 20 most- parsimonious cladograms of HEA P. b unistrigatus group. a. Tree without showing nodes numbers, and showing the character state 44 (1) as an unambiguous synapomorphy Figure 2. a. Ventral view of the manus b. Tree showing nodes numbers. showing the two conditions of the character 23 in the LD matrix: a. origin of the m. adductor Abreviations: Cran: Craugastor raniformis, Pano: policis is not covered by the aponeurosis Pristimantis anolirex, Pbac: P. bachus, Pbic: P. palmaris (23-1). P. bicolor (ICN 26321. b. bicolor, Pbog: P. bogotensis, Pele: P. elegans, Pfra: P. frater, Pjor: P. jorgevelosai, Plyn: P. lynchi, Pmer: P. Origin of the m. adductor policis is covered merostictus, Pmiy: P. miyatai, Pner: P. nervicus, Pnic: P. by the aponeurosis palmaris. C. raniformis nicefori, Ppro: P. prolixodiscus, Ppug: P. pugnax, Pspi: (23-0). MUJ 2871 P. spilogaster, Ptae: P. taeniatus, Ptam: P. tamsitii. 114 Hoyos et al. this does not mean that we are clear about the (44, including four of Burton) than in the putative synapomorphies involved, as we saw feet (21, including six of Burton). Similarly, above. It is remarkable that the relationships after making a clearance based on external between P. pugnax and P. tamsitti, and descriptions and original observations, we between P. spilogaster and P. merostictus are obtained 26 characters. Faivovich (2002) retained in both cladograms, meaning that used 14 characters from the manus and these species maybe considered sisters, but seven from the pes, from which he found two without finding putative synapomorphies to synapomorphies for Scinax Wagler, 1830 , support the relationship between P. pugnax both from the manus. However, although we and P. tamsitti. did not find any useful characters taken from hand and foot musculature, this does not mean Limb muscles and phylogeny that these characters should be dismissed from phylogenetic studies in frogs. Burton All characters and character states obtained (1998a) found some hand muscle character were chosen bearing in mind character conditions useful to support intrafamiliar distribution and conditions deemed to be relationships in Leptodactylids. Likewise, worth for the analysis. Burton’s (1998a, 2004) Burton (1998b) found a hand muscle character characters and character states were used just condition within Ranidae Rafinesque, 1814, as he defined them, but in some cases we stressing that superficial muscles can vary believed that some had to be redefined due more than deep ones, but that these muscle to the fact that they were not clearly named. can contribute to get frog relationships. It is Then, we think that most of these characters apparently historically clear that superficial are new and very useful for future analyses hand muscles may play a role more important in frog morphology and phylogeny studies. to resolve frog relationships than superficial It is fascinating how many characters have feet muscles, although we found muscles of found on the hands and feet, although only both that could be putative synapomorphies. the superficial muscles have been observed. More characters were found in the hands External characters

Phylogenetic analysis including external characters is rare in frogs, although identification of most of the species groups and all species have been based on these characters. The results show that characteristics of the fingers are still important, and can be used in establishing kinship, but were obviously not essential to support the monophyly of the unistrigatus group. Apparently, the other external characters are not useful because, in any case, it was established that they could participate as potential synapomorphies. Figure 4. Ventral view of the pes showing the Maybe we should pay attention to the apomorphic state of the character state 44 (1). condition of tympanic annulus and to the Origin of the tendo superficialis hallucis from dorsal view of the snout because for the the aponeurosis plantaris including some two analyses that show that these characters fibers of the m. lumbricalis brevis hallucis. appear as potential synapomorphies (nodes P. mersotictus. ICN 34235 21 and 23 respectively in LD group, and node 115 Phylogenetic analysis whithin the Pristimantis unistrigatus

15 in HEA group) , although actually they are LITERATURE CITED homoplasies. Acosta, A. R. 2000. Ranas, Salamandras To compare our results with those by Heinicke y Caecilias (Tetrapoda: Amphibia) de et al. (2008) and Hedges et al. (2008) is difficult, Colombia. Biota Colombiana 1 (3): 289- because most of the species included in our 319. work were not used by these authors. However, Andersen, M. 1978. The comparative our results agree in identifying the unistrigatus myology and osteology of the carpus and group as non monophyletic. Heinicke et al. tarsus of selected anurans. Volumes I and (2008) only included six spececies belonging II. Ph.D. Dissertation.University of Kansas, to the former P. unistrigatus group (Lynch, Department of Systematics and Ecology. 1976; Lynch et al., 1997). Taking into account Kansas, USA. 310pp. this classification, their results showed that Bernal, X. & F. Guzmán. 1999. The this group is monophyletic. However, based advertisement calls of three Eleutherodactylus on Hedges et al. (2008) classification P. species (Anura: Leptodactylidae) in ockendeni, in the P. unistrigatus group by Colombian highland community. Revista Heinicke et al. (2008), belongs to the P. frater Academia Colombiana de Ciencias Exactas group; in this sense, the group would become Físicas y Naturales 23(suplemento especial): not monophyletic. 261-264. Bock, W. & O.R. Shear. 1972. A staining CONCLUSIONS method for gross dissection of vertebrate muscles. Anatomischer Anzeiger 130: Although the muscles of hands and feet and 222-227. external characters useful in taxonomy failed Burton, T.C. 1996. Adaptation and evolution in to identify the Pristimantis unistrigatus as the hand muscles of Australo-Papuan Hylid monophyletic, this does not mean they are Frogs (Anura: Hylidae: Pelodryadinae). not useful for establishing relationships Australian Journal of Zoology 44: 611- within the genus. It is possible that including 623. more unistrigatus group species in the study, Burton, T.C. 1998a. Variation in the hand and the situation changes, however, it seems superficial throat musculature of neotropical also essential to include the deep muscles of Leptodactylidae frogs. Herpetologica 54: the hands and feet and external characters 53-72. as well. It is possible that other characters Burton, T.C. 1998b. Pointing the way: facilitate to define relationships among the distribution and evolution of some species within the genus. However, it is characters of the finger muscles of frogs. necessary to explore deeper the relationship American Museum Novitates 3229: 1-13. P. spilogaster-P. merostictus, and P. pugnax- Burton, T.C. 2001. Variation in the foot muscles P. tamsitti, because we need whether to of frogs of the family Myobatrachidae. confirm or falsify if these clades actually are Australian Journal of Zoology 49: 539- monophyletic. By detecting the difficulties 559. that several researchers (e,g, Hedges et al. Burton, T.C. 2004. Muscles of the pes of 2008; Padial et al. 2009) and our own study Hylid frogs. Journal of Morphology 260: have found to discover the monophyly of the 209-233. P. unistrigatus group and other ones in frogs, Cochran D.M. & C.J. Goin 1970. Frogs of the question we pose is: Is it worthwhile to Colombia. United States National Museum keep this informal taxonomic hierarchy called Bulletin 288, Smithsonian Institution Press, group in systematics of frogs? Washington D.C. 655 pp. 116 Hoyos et al.

Conrad, J.L. 2008. Phylogeny and systematics Glaw, F.M., M. Vences, F. Andreone & D. of Squamata (Reptilia) based on morphology. Vallan. 2001. Revision of the Boophis Bulletin of the American Museum of Natural majori group (Amphibia : Mantellidae) History 310: 1-182. from Madagascar, with the description of Duellman, W.D. & J.B. Pramuk. 1999. five new species. Zoological Journal of the Frogs of the Genus Eleutherodactylus: Linnean Society133: 495-529. Anura: Leptodactylidae) in the Andes of Hedges, S.B., W.E. Duellman & M.P. Northern Peru. Scientific Papers, Natural Heinicke. 2008. New World direct- History Museum, The University of Kansas developing frogs (Anura: Terrarana): (13):1-78. Molecular phylogeny, classification, Duellman, W.D. & J.E. Simmons. 1977. A biogeography, and conservation. Zootaxa new species of Eleutherodactylus: Anura: 1737: 1-182 pp Leptodactylidae) from the Cordillera Oriental Heinicke, M.P., W.E. Duellman & S. B. of Colombia. Proceedings of the Biological Hedges. 2007. Major Caribbean and Central Society of Washington 90: 60-65. American frog faunas originated by ancient Dunlap, D. 1960. The comparative miology oceanic dispersal. Proceedings of the of the pelvic appendage in Salientia. Journal National Academy of Sciences of United of Morphology 160: 1-76. States of America 104 (24): 1092-1097. Dunn, E. R. 1944. Herpetology of Bogotá Hoyos, J.M. 1991. Aspectos taxonómicos y Area. Revista de la Academia Colombiana patrones de diseño de Eleutherodactylus de Ciencias Exactas, Físicas y Naturales bogotensis del Parque Nacional Natural 6: 68-81. Chingaza. Cuadernos Divulgativos 18: Egan, D. 1995. The comparative osteology 1-12 and miology of the carpus of the Australian K w e t , A. & J. FAIVOVICH. 2001. Leptodactylidae. Honors Dissertation. Protoceratophrys bigibbosa species group University of Adelaide. Adelaide, Australia. (Anura: Leptodactylidae), with description 58pp. of new species. Copeia 2001: 203-215. Faivovich, J. 2002. A cladistic analysis of L y n c h , J.D. 1973. A new species Scinax (Hylidae). Cladistics 18: 367-393. of Eleutherodactylus (Amphibia, Frost, D., T. Grant, J. Faivovich, R. Bai, A. Leptodactylidae) from Andean Ecuador. Haas, C. Haddad, R. De Sá, A. Channing, M. Bulletin of the Southern California Academy Wilkinson, S. Donnellan, C. Raxworthy, J. of Sciences 72: 107-109. Campbell, B. Blotto, P. Moler, R. Drewes, Lynch, J.D. 1975a. A review of the broad- R. Nussbaum, J. Lynch, D. Green & W. headed eleutherodactyline frogs of South Wheeler 2006. Tree of life. The amphibian America (Leptodactylidae). Occasional tree of life. Bulletin of the American Papers of the Museum of Natural History Museum of Natural History 297: 1–370. University of Kansas 38: 1-46. Galindo, D. 2004. Filogenia de especies del Lynch, J.D. 1975b. The identity of the frog grupo unistrigatus (sensu Lynch), del género Eleutherodactylus conspicillatus (Günther) Eleutherodactylus (Amphibia: Anura: with description of two related species from Leptodactylidae) de la cordillera Oriental de Northwestern Soth America (Amphibia, Colombia. Trabajo de Grado de Pregrado. Leptodactylidae). Natural History Museum Pontificia Universidad Javeriana, Bogotá, of Los Angeles Contributions in Science Colombia. 190 pp. 272: 1-19. Gaupp, E.1896. A. Ecker’s und R. Wiedersheim’s Lynch, J.D. 1976.The species groups of Anatomie des Frosches. Friedrich View und the South American frogs of the genus Sohn, Braunschweig. Eleutherodactylus (Leptodactylidae). 117 Phylogenetic analysis whithin the Pristimantis unistrigatus

Occasional papers of the Museum of Natural southern Boyacá. Revista de la Academia History 61: 1-24. Colombiana de Ciencias Exactas, Físicas y Lynch, J.D. 1978. A new Eleutherodactyline Naturales 22: 429-432. frog from the Andes of Northern Colombia Lynch, J.D. 2001. Four osteological (Leptodactylidae). Copeia 1978: 17-21. synapomorphies within Eleutherodactylus Lynch, J.D. 1981. Two new species of (Amphibia: Leptodactylidae) and their Eleutherodactylus from Western Colombia bearing on subgeneric classifications. (Amphibia, Anura, Leptodactylidae). Revista de la Academia Colombiana de Occasional Papers of the Museum of Zoology, Ciencias Exactas, Físicas y Naturales 25: University of Michigan 697: 1-12. 127-136. Lynch, J.D. 1983. A new Leptodactylid frog Lynch, J.D. 2003. New species of frogs from the Cordillera Oriental of Colombia. (Eleutherodactylus: Leptodactylidae) In: Rhodin, A. G. J. & Williams, E. E. from the Cordillera Oriental of Norte de Advances in Herpetology and evolutionary Santander and Santander, Colombia. Revista biology: 52-57. Museum of Comparative de la Academia Colombiana de Ciencias Zoology, Cambridge, Massachussetts. Exactas, Físicas y Naturales 27: 449-460. Lynch, J.D. 1984. A new species of Lynch, J.D. & M.C. Ardila-Robayo. 1999. Eleutherodactylus (Amphibia, Anura: The Eleutherodactylus of the taeniatus Leptodactylidae) from Southern Andean complex in Western Colombia: taxonomy Colombia. Herpetologica 40: 234-237. and distribution. Revista de la Academia Lynch, J.D. 1989. Intrageneric Relationships Colombiana de Ciencias Exactas, Físicas y of Mainland E l e u t h e ro d a c t y l u s Naturales 23(89): 615-624. (Leptodactylidae) I. A Review of the Lynch, J.D. & W. Duellman. 1980. The Frogs Assigned to the Eleutherodactylus Eleutherodactylus of the Amazonian discoidalis species group.Contributions in slopes of the Ecuadorian Andes (Anura, Biology and Geology, Milwaukee Public Leptodactylidae). The University of Kansas, Museum 79: 1-25. Lawrence, Kansas 69: 1-86. Lynch, J.D. 1994a. A new species of Lynch, J.D. & W. Duellman. 1997. Frogs High-altitude frog (Eleutherodactylus of the genus Eleutherodactylus in Western Leptodactylidae) from the Cordillera Ecuador. Special publication, No. 23, Oriental of Colombia. Revista de la Natural History Museum, The University Academia Colombiana de Ciencias Exactas, of Kansas, Lawrence, Kansas. Físicas y Naturales 19: 195-203. Lynch, J.D. & B. Myers. 1983. Frogs of Lynch, J.D. 1994b. A new species of frog the fitzingeri group of Eleutherodactylus (Genus Eleutherodactylus: Leptodactylidae) in Eastern Panama and Chocoan South from the cloud forest in Departamento America (Leptodactylidae). Bulletin of the de Santander, Colombia. Revista de la American Museum of Natural History 175: Academia Colombiana de Ciencias Exactas, 484-568. Físicas y Naturales 19: 205-20. Lynch, J.D. & A.M. Suárez. 2002. A new frog Lynch, J.D. 1997. Intrageneric relationships (Eleutherodactylus: Leptodactylidae) from of mainland Eleutherodactylus II. A review the Southern part of the Cordillera Oriental of the Eleutherodactylus sulcatus group. de Colombia. Revista de la Academia Revista de la Academia Colombiana de Colombiana de Ciencias Exactas, Físicas y Ciencias Exactas, Físicas y Naturales 21: Naturales 24: 289-293. 353-372. Nixon, K. & J.M. Carpenter. 1993. On Lynch, J.D. 1998. A new frog (Genus outgroups. Cladistics 9: 413-426. Eleutherodactylus) from cloud forests of 118 Hoyos et al.

Padial, J. M., S. Castroviejo-Fisher, J. Rueda, J.V. & J.D. Lynch. 1983. Una nueva Köhler, C. Vilá, J. C. Chaparro & I. de especie de Eleutherodactylus (Amphibia: la Riva. 2009. Deciphering the products of Leptodactylidae) para la Cordillera Oriental evolution at the species level: the need for de Colombia. Lozania 42: 1-6. an integrative taxonomy. Zoologica Scripta Salgar, L. 2003. Miología de manos y 38: 431-447. pies de Hyla labialis (Peters, 1883) y de Pyburn, W.F. & J.D. Lynch. 1981. Two Eleutherodactylus bogotensis (Peters, little-known species of Eleutherodactylus 1883). Trabajo de Grado, Departamento de (Amphibia, Leptodactylidae) from the Sierra Biología, Pontificia Universidad Javeriana de la Macarena, Colombia. Proceedings of Bogotá D.C. 258 pp. the Biological Society of Washington 94: Salgar, L., J.M. Hoyos & A. Acosta. 2009. 404-412. New conditions and intraspecific variations Raikow, R., A. Bledsoe, B. Myers & C. in hand and foot muscles in Pristimantis Welsh. 1990. Individual variation in bogotensis (Peters, 1863) (Amphibia:Anura: avian muscles and its significance for the Strabomantidae). Zootaxa 2025: 56-68. reconstruction of phylogeny. Systematic Zoology 39: 362-370. Reichle, S., S. Lötters & I. de la Riva. 2001. A new species of the discoidalis group of Eleutherodactylus (Anura, Leptodactylidae) fron Inner-Andean Dry Valley of Bolivia. Recibido: 12/12/2011 Journal of Herpetology 35: 21-26. Aceptado: 25/02/2014

Appendix 1. List of characters and character states for the LD matrix.

Manus 0 The m. palmaris longus is partially covered by 1. Number of insertions of the m. extensor digitorum the m. palmaris longus communis longus 1 The m. palmaris longus is not covered by the m. 0 Four palmaris longus 1 Three 8. Relationship between the m. epitrochleocubitalis and 2 Two the m. epicondylocubitalis 2. Number of origins of the m. epicondylocubitalis 0 The fibers fuse partially on the external surface 0 Two, one slip in common with the m. extensor of the radioulna carpi ulnaris and the other slip free. 1 The fibers completely fuse on the external 1 One slip, not in common with the m. extensor surface of the radioulna carpi ulnaris 9. Relationship between the internal slip of the m. 3. Relationship between the m. epicondylocubitalis and extensor digitorum communis longus and the m. the m. epitrochleocubitalis extensor carpi radialis 0 Bipennated 0 Proximally partially fused 1 Parallel 1 Completely fused 4. Type of origin of the m. epitrochleocubitalis 2 Not fused 0 Double 10. Relationship betweem the m. extensor brevis 1 Single superficialis DigitiV and the m. abductor brevis 5. Insertion of the m. epitrochleocubitalis Digiti V 0 Not covered by the m. palmaris brevis 0 They are partially fused 1 Covered by the m. palmaris brevis 1 They are not fused 6. Insertion of the m. epicondylocubitalis in relation 11. Number of insertion of the m. extensor brevis with the m. palmaris brevis superficialis DigitiV 0 It does not cover the m. palmaris brevis 0 One 1 It covers the m. palmaris brevis 1 Two 7. Relationship between the m. epitrochleocubitalis and 12. Origin of the m. abductor brevis Digiti V the external slip of the m. palmaris longus 0 From the distal carpal 5-4-3 119 Phylogenetic analysis whithin the Pristimantis unistrigatus

1 From the ulnare 0 Short and wide with respect to the muscle 13. Presence of the m. abductor ulnocarpalis 1 Long and thin with respect to the muscle 0 Present 26. Origin of the m. lumbricalis brevis Digiti III in 1 Absent relation with the tendo superficialis & caput. 14. Origin of the m. palmaris longus profundum Digiti III 0 By two slips 0 Dorsal 1 By one slip 1 Laterodorsal 15. Insertion of the m. palmaris longus 27. Presence of fibers of thetendo superficialis & 0 Simple caput. profundum Digiti III 1 Double 0 Only on the external surface of the tendon 16. Connection between the tendons of the m. palmaris 1 On the external and internal surface of the longus and the flexor tendons tendon 0 Single tendon that divides to become tendones 2 Only on the internal surface of the tendon superficiales(TS) IV and V 28. Nature of the m. lumbricalis brevis Digiti IV 1 Two tendons that cross over before becoming TS 0 Two slips IV and V 1 Only one slip 17. Relationship between the fibers of them. palmaris 29. Position of the insertion tendons of the m. brevis and the m. palmaris longus lumbricalis longus Digiti IV with respect to the 0 The fibers of them. palmaris brevis are dorsal tendo superficialis aponeurosis palmaris prodigiti with respect to the tendons of insertion of the m. IV palmaris longus 0 Lateral to the tendo superficialis aponeurosis 1 The m. palmaris brevis is between the two palmaris prodigiti IV tendons of insertion of the m. palmaris longus 1 The insertion is via the tendo superficialis 18. Relationship between the insertion of the m. aponeurosis palmaris prodigiti IV palmaris brevis and the tendons of insertion of the 30. Origin of the m. lumbricalis longus Digiti V m. palmaris longus 0 From the aponeurosis palmaris and the TS V 0 m. palmaris brevis articulates with the tendons of 1 Only from the TS V insertion of the m. palmaris longus 31. Number of insertions of m. lumbricalis longus 1 The m. palmaris brevis does not articulate with Digiti V the m. palmaris longus 0 One 19. Relationship between the m. palmaris longus and 1 Two the m. flexor carpi ulnaris 32. Number of slips that compose the m. lumbricalis 0 They are partially fused brevis Digiti V 1 They are not fused 0 One 20. Origin of the m. adductor pollicis 1 Two 0 From the ventral surface of the carpal 5-4-3 1 From the internal surface of the distal carpal 5-4-3 Pes 21. Insertion of the m. adductor pollicis on the 33. Type of insertion of the m. extensor digitorum prepollex communis longus 0 Adjacent to the insertion of the m. abductor 0 Single pollicis 1 Multiple 1 Covers the insertion of the m. abductor pollicis 34. Number of insertions of the m. extensor brevis 22. Relationship between the m. adductor pollicis and superficialis hallucis the m. abductor pollicis 0 Two 0 Both muscles are independent 1 Three 1 They are fused 2 Single 23. Relationship between the origin of the m. adductor 35. Number of insertions of the m. extensor brevis pollicis and the aponeurosis palmaris superficialis Digiti II 0 The origin is covered by the aponeurosis 0 One palmaris 1 Two 1 The origin is not covered by the aponeurosis 36. Relationship between the insertions of the m. palmaris extensor brevis superficialis Digiti II and the m. 24. Condition of the proximal end of the m. lumbricalis extensor digitorum communis longus brevis indicis 0 Common insertion between m. extensor brevis 0 Dorsal to the m. adductor pollicis superficialis Digiti II and the internal slip of m. 1 Contiguous to the m. adductor pollicis extensor digitorum communis longus 25. Nature of the origin of the tendon of the m. 1 Inependent insertions lumbricalis brevis Digiti III 37. Presence of the m. extensor brevis medius hallucis

120 Hoyos et al.

0 Absent 0 From both, the ulnare and distal carpals 5-4-3 1 Present 1 From the ulnare 38. Presence of the m. extensor brevis medius Digiti II 0 Absent Pes 1 Present 51. Origin of the tendo superficialis hallucis 39. Number of insertion slips of the m. extensor brevis 0 From the aponeurosis plantaris superficialis Digiti III 1 From the aponeurosis plantaris including some 0 One fibers of them. lumbricalis brevis hallucis 1 Two 2 From a muscle that originates from the distal 40. Relationship between the insertions of the m. tarsal 2-3 extensor brevis superficialis Digiti III and the m. 52. Origin of the tendo superficialis Digiti II extensor digitorum communis longus 0 From the aponeurosis plantaris 0 Common insertion between m. extensor brevis 1 It serves as the insertion tendon of the m. superficialis Digiti III and any slip of m. extensor transversus plantae distalis digitorum communis longus 53. Origin of the tendo superficialis Digiti III 1 Independent insertions 0 From the m. flexor digiti brevis superficialis only 41. Origin of the m. extensor brevis superficialis Digiti 1 From the aponeurosis plantaris IV 2 From a superficial tendon that emerges centrally 0 By two independent slips on the plantar surface of the aponeurosis plantaris 1 By two proximally fused slips 54. Number of insertion tendons of the m. lumbricalis 2 By one slip logissimus Digiti IV 42. Presence of the m. extensor brevis superficialis 0 Two Digiti V 1 One 0 Absent 1 Present External characters 43. Origin of the m. abductor prehallucis 55. Skin on venter 0 From the internal surface of the aponeurosis 0 Areolate plantaris 1 Smooth 1 From the distal internal surface of the tarsus 56. Presence of dorsolateral folds 44. Presence of the m. abductor brevis plantaris 0 Present hallucis 1 Absent 0 Absent 57. Nature of tympannic anulus 1 Present 0 Complete 45. Origin of the m. lumbricalis brevis Digiti II 1 Incomplete 0 From the aponeurosis plantaris 58. Dorsal view of snout 1 From the proximal end of the tendo superficialis 0 Rounded hallucis 1 Subacuminate 46. Type of origin of the m. lumbricalis brevis Digiti III 2 Acuminate 0 Tendinous 59. Shape of snout in profile 1 Fleshy 0 Rounded 47. Relationship between the external slip of the m. 1 Truncated lumbricalis brevis Digiti IV and the internal slip of 2 Acute m. lumbricalis brevis Digiti V 60. Presence of cranial crests 0 Fused only in the origin 0 Present 1 Fused beyond its origin 1 Absent 61. Width of the upper eyelid in relation with the inter- Burton characters orbital distance 0 Narrower than the inter-orbital distance Manus 1 As wide as the inter-orbital distance 48. Presence of the internal slip of the m. lumbricalis 62. Length of fingers I and II brevis Digiti III 0 First finger shorter than second 0 Absent 1 First and second finger equally or almost equally 1 Present long 49. Number of slips of the m. lumbricalis longus Digiti 2 First finger longer than second IV 63. Presence of finger pads or discs 0 Two 0 Present 1 Only one, the internal slip 1 Absent 50. Origin of the m. extensor brevis superficialis Digiti V 64. Presence of skin variations on the ulnar region

121 Phylogenetic analysis whithin the Pristimantis unistrigatus

0 Present 1 Absent 1 Absent 68. Presence of webbed toes 65. Presence of skin variations on the tarsals 0 Present 0 Present 1 Absent 1 Absent 69. Presence of toe pads or discs 66. Presence of skin variations on the heel 0 Present 0 Present 1 Absent 1 Absent 70. Presence of lateral fringes on toes 67. Presence of supernumerary plantar tubercles 0 Present 0 Present 1 Absent

Appendix 2. List of characters and character states for the HEA matrix. Manus 11. Origin of the m. abductor brevis Digiti V 1. Number of insertions of the m. extensor digitorum 0 From the distal carpal 5-4-3 communis longus 1 From the ulnare 0 Three 12. Presence of the m. abductor ulnocarpalis 1 Two 0 Present 2. Relationship between the m. epicondylocubitalis and 1 Absent the m. epitrochleocubitalis 13. Origin of the m. palmaris longus 0 Bipennated 0 By two slips 1 Parallel 1 By one slip 3. Type of origin of the m. epitrochleocubitalis 14. Insertion of the m. palmaris longus 0 Double 0 Simple 1 Single 1 Double 4. Insertion of the m. epitrochleocubitalis 15. Connection between the tendons of the m. palmaris 0 Not covered by the m. palmaris brevis longus and the flexor tendons 1 Covered by the m. palmaris brevis 0 Single tendon that divides to become tendones 5. Insertion of the m. epicondylocubitalis in relation superficiales(TS) IV and V with the m. palmaris brevis 1 Two tendons that cross over before becoming TS 0 It does not cover the m. palmaris brevis IV and V 1 It covers the m. palmaris brevis. 16. Relationship between the fibers of them. palmaris 6. Relationship between the m. epitrochleocubitalis and brevis and the m. palmaris longus the external slip of the m. palmaris longus 0 The fibers of them. palmaris brevis are dorsal 0 The m. epitrochleocubitalis is partially covered with respect to the tendons of insertion of the m. by the m. palmaris longus palmaris longus 1 The m. epitrochleocubitalis is not covered by the 1 The m. palmaris brevis is between the two m. palmaris longus. tendons of insertion of the m. palmaris longus 7. Relationship between the m. epitrochleocubitalis and 17. Relationship between the m. palmaris longus. and the m. apicondylocubitalis the m. flexor carpi ulnaris 0 The fibers fuse partially on the external surface 0 They are partially fused of the radioulna 1 They are not fused 1 The fibers completely fuse on the external 18. Origin of the m. adductor pollicis surface of the radioulna 0 From the ventral surface of the carpal 5-4-3 8. Relationship between the internl slip of the m. 1 From the internal surface of the distal carpal 5-4- extensor digitorum communis longus. and the m. 3 extensor carpi radialis 19. Insertion of the m. adductor pollicis. on the 0 Proximally partially fused prepollex 1 Completely fused 0 Adjacent to the insertion of the m. abductor 2 Not fused pollicis 9. Relationship between the m. extensor brevis 1 Covers the insertion of the m. abductor pollicis superficialis DigitiV and the m. abductor brevis 20. Relationship between the m. adductor pollicis and Digiti V the m. abductor pollicis 0 They are partially fused 0 Both muscles are independent 1 They are not fused 1 They are fused 10. Number of insertion of the m. extensor brevis 21. Condition of the proximal end of m. lumbricalis superficialis DigitiV brevis indicis 0 One 0 Dorsal to the m. adductor pollicis 1 Two 1 Contiguous to the m. adductor pollicis 122 Hoyos et al.

22. Origin of the m. lumbricalis brevis Digiti III in 0 Absent relation with the m. tendo superficialis & caput 1 Present profundum Digiti III 35. Number of insertion slips of the m. extensor brevis 0 Dorsal superficialis Digiti III 1 Laterodorsal 0 One 23. Presence of fibers of thetendo superficialis & 1 Two caput. profundum Digiti III 36. Relationship between the insertions of the mm. 0 On the external and internal surface of the extensor brevis superficialis DigitiIII and the m. tendon extensor digitorum communis longus. 1 Only on the internal surface of the tendon 0 Common insertion between mm. extensor brevis 24. Nature of the m. lumbricalis brevis Digiti IV superficialis DigitiIII and any slip of m. extensor 0 Two slips digitorum communis longus 1 Only one slip 1 Independent insertions 25. Position of the insertion tendons of the m. 37. Origin of the m. extensor brevis superficialis Digiti lumbricalis longus Digiti IV with respect to the IV tendo superficialis aponeurosis palmaris prodigiti 0 By two independent slips IV 1 By two proximally fused slips 0 Lateral to the tendo superficialis aponeurosis 2 By one slip palmaris prodigiti IV 38. Origin of the m. abductor prehallucis 1 The insertion is via the tendo superficialis 0 From the internal surface of the aponeurosis aponeurosis palmaris prodigiti IV plantaris 26. Origin of the m. lumbricalis longus Digiti V 1 From the distal internal surface of the tarsus 0 From the aponeurosis palmaris. and TS V 39. Presence of the m. abductor brevis plantaris 1 Only from TS V hallucis 27. Number of insertions of m. lumbricalis longus 0 Absent Digiti V 1 Present 0 One 40. Type of origin of the m. lumbricalis brevis Digiti III 1 Two 0 Tendinous 28. Number of slips that compose the m. lumbricalis 1 Fleshy brevis Digiti V 41. Relationship between the external slip of m. 0 One lumbricalis brevis Digiti III and the internal slip of 1 Two m. lumbricalis brevis Digiti V 0 Fused only in the origin Foot 1 Fused beyond its origin 29. Type of insertion of the m. extensor digitorum communis longus Burton characters 0 Single 1 Multiple Manus 30. Number of insertions of the m. extensor brevis 42. Presence of the internal slip of m. lumbricalis superficialis hallucis brevis Digiti III 0 Two 0 Absent 1 Three 1 Present 2 Single 43. Number of slips of the m. lumbricalis brevis Digiti 31. Number of insertions of the m. extensor brevis IV superficialis Digiti II 0 Two 0 One 1 Only one, the internal slip 1 Two 32. Relationship between the insertions of the m. Pes extensor brevis superficialis Digiti II and the m. 44. Origin of the tendo superficialis hallucis extensor digitorum communis longus. 0 From the aponeurosis plantaris 0 Common insertion between m. extensor brevis 1 From the aponeurosis plantaris including some superficialis DigitiII and the internal slip of the m. fibers of them. lumbricalis brevis hallucis extensor digitorum communis longus 2 From a muscle that originates from the distal 1 Inependent insertions tarsal 2-3 33. Presence of the m. extensor brevis medius hallucis 45. Origin of the tendo superficialis Digiti II 0 Absent 0 From the aponeurosis plantaris 1 Present 1 It serves as the insertion tendon of the m. 34. Presence of the m. extensor brevis medius Digiti II transversus plantae distalis

123 Phylogenetic analysis whithin the Pristimantis unistrigatus

External characters 53. Length of fingers I and II 46. Skin on venter 0 First finger shorter than second 0 Areolate 1 First and second finger equally or almost equally 1 Smooth long 47. Presence of dorsolateral folds 2 First finger longer than second 0 Present 54. Presence of finger pads or discs 1 Absent 0 Present 48. Nature of tympannic anulus 1 Absent 0 Complete 55. Presence of skin variations on the ulnar region 1 Incomplete 0 Present 49. Dorsal view of snout 1 Absent 0 Rounded 56. Presence of skin variations on the heel 1 Subacuminate 0 Present 2 Acuminate 1 Absent 50. Shape of snout in profile 57. Presence of supernumerary plantar tubercles 0 Rounded 0 Present 1 Truncated 1 Absent 2 Acute 58. Presence of webbed toes 51. Presence of cranial crests 0 Present 0 Present 1 Absent 1 Absent 59. Presence of toe pads or discs 52. Width of the upper eyelid in relation with the inter- 0 Present orbital distance 1 Absent 0 Narrower than the inter-orbital distance 60. Presence of lateral fringes on toes 1 As wide as the inter-orbital distance 0 Present 1 Absent

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