Anura) Proveniente Da Caverna Versalles (Pleistoceno Final/ Holoceno Inicial), Apiaí, SP, Com Comentários Sobre Paleobiogeografia E Paleoambiente Da América Do Sul

UNIVERSIDADE FEDERAL DO ABC

PROGRAMA DE PÓS-GRADUAÇÃO EM EVOLUÇÃO E DIVERSIDADE

Lucas Almeida Barcelos

Descrição e posicionamento filogenético de um Ceratophryideo (Anura) proveniente da Caverna Versalles (Pleistoceno Final/ Holoceno Inicial), Apiaí, SP, com comentários sobre paleobiogeografia e paleoambiente da América do Sul

Santo André – SP

2019

Lucas Almeida Barcelos

Descrição e posicionamento filogenético de um Ceratophryideo (Anura) proveniente da Caverna Versalles (Pleistoceno Final/Holoceno Inicial), Apiaí, SP, com comentários sobre paleobiogeografia e paleoambiente da

América do Sul

Dissertação apresentada ao Programa de Pós-graduação em Evolução e Diversidade da Universidade Federal do ABC, como requisito para a obtenção do título de Mestre em Evolução e Diversidade. Linha de pesquisa: Sistemática e Biogeografia.

Orientadora: Profa. Dra. Vanessa Kruth Verdade

Co-orientador: Prof. Dr. Charles Morphy Dias dos Santos

Santo André – SP

2019

Lucas Almeida Barcelos

Descrição e posicionamento filogenético de um Ceratophryídeo (Anura) proveniente da Caverna Versalles (Pleistoceno Final/Holoceno Inicial),

Apiaí, SP

Essa dissertação foi julgada e aprovada para a obtenção do grau de mestre em Evolução e Diversidade pelo Programa de Pós-graduação em Evolução e Diversidade da Universidade Federal do ABC.

Santo André – SP, 21 de maio de 2019.

Prof. Dr. Gustavo Muniz Dias

BANCA EXAMINADORA

______

Profa. Dra.Vanessa Kruth Verdade

Orientadora

______

Prof. Dr. Helio Ricardo da Silva

Universidade Federal Rural do Rio de Janeiro

______

Prof. Dr. Ivan Sergio Nunes Silva Filho

Universidade Estadual Paulista “Júlio de Mesquita Filho” Campus Litoral Paulista

Agradecimentos

Agradeço à FAPESP (Fundação de Amparo à Pesquisa do Estado de São Paulo) pela concessão da bolsa de mestrado (processo 2017/04849-1) que possibilitou minha participação em congressos, visita a coleções, contato com outros grupos de pesquisa e com isso me ajudou a chegar a uma melhor versão do trabalho que realizo. Agradeço a UFABC por fomentar minha pesquisa em meus primeiros meses de mestrado e por todo o apoio logístico durante a pós-graduação. Agradeço ao Prof. Dr. Hussam El Dine Zaher por ter permitido acesso a coleção de anfíbios e ao microtomógrafo do Museu de Zoologia da Universidade Federal de São Paulo - MZUSP e à Dra. Aline Staskowian Benetti e Dr. Alberto Barbosa de Carvalho, por terem viabilizado a consulta ao material e realização das tomografias dos espécimes utilizados para estudos comparativos. Agradeço ao Dr. Martín D. Ezcurra por ter cedido acesso a coleção de Paleontologia de Vertebrados do Museo Argentino de Ciencias Naturales “Bernardino Rivadavia” (MACN), pela presteza, simpatia dispensada a mim ao longo do período que visitei o museu e por dispor de seu tempo para me proporcionar uma visita guiada da exposição do MACN. À Dra. Laura Nicoli e ao Prof. Dr. Julian Faivovich do MACN pelas conversas enriquecedoras a respeito de fósseis de Anura e histórico de autores de trabalhos envolvendo Ceratophryidae. Ao Prof. Dr. Marcelo Reguero, curador da coleção de Paleontologia de Vertebrados da Universidad Nacional de La Plata, por ter permitido acesso à coleção, pela simpatia e presteza dispensada a mim ao longo do período que visitei o museu. Agradeço a Dra. Addison Wynn, responsável pela coleção de anfíbios e a Dra. Rayna Bell, curadora da coleção de anfíbios e répteis do National Museum of Natural History - Smithsonian Institution por me permitir acesso a fotos, imagens de raio-x de espécimes de Ceratophrys stolzmanni. Agradeço a Profa. Dra. Sandra D. Chapman, curadora da coleção de répteis e aves do British Natural History Museum, por fornecer informações e fotos do espécime de Ceratophryidae descrito por Günther (1859). Agradeço ao Dr. Damien Germain, curador da coleção de Paleontologia de Vertebrados do Muséum National d'Histoire Naturelle da França por ter realizado a difícil tarefa de rastrear fósseis de Ceratophryidae sem número de tombo, descritos na década de sessenta e por ter fornecido informações e fotos de tais espécimes. Agradeço ao Especialista em Recursos Minerais Dr. Rodrigo da Rocha Machado por me permitir acessar a coleção de Paleontologia do Museu de Ciências da Terra - DNPM (Departamento Nacional de Produção Mineral) e pela presteza dispensada ao longo do período que visitei a coleção.

Agradeço a Mónica Analía Hidalgo da Biblioteca Florentino Ameghino, da Universidad Nacional de La Plata por me permitir ter acesso à Tese de Perí (1994), bibliografia indispensável ao desenvolvimento de qualquer trabalho versando sobre o estudo morfológico-comparativo de Ceratophryidae. Agradeço também a Dra. Silvia Isabel Perí por ter feito tantos avanços no estudo osteológico desta família de Anura. Agradeço especialmente a Prof. Dra. Florencia Vera Candioti, por ser uma pessoa incrível e ter se desdobrado algumas vezes para nos ajudar na Argentina, em vários momentos importantes da vida científica, agradeço também a Bel. Ana Sofia Duport Bru, pela disposição em ajudar e por ter fornecido imagens de espécimes importantes de Ceratophryidae. Agradeço ao Dr. Luke Welton, curador da coleção de Herpetologia do University of Kansas Biodiversity Institute por gentilmente ceder fotos de espécimes de Ceratophryidae. Agradeço ao Dr. Sven Kullander, curador da coleção de Herpetologia do Naturhistoriska Riksmuseet de Estocolmo, Suécia pela atenção dispensada a mim e por fornecer informações acerca do holótipo de C. testudo. Ao Prof. Dr. Rondinelli Donizetti Herculano da UNESP – Araraquara, por possibilitar a datação do fóssil de Ceratophryidae (ZUFABC 037). A busca por bibliografia foi um desafio considerável no decorrer deste trabalho, principalmente porque existem diversos trabalhos clássicos sobre este tópico de estudo. Mas consegui a grande maioria dos trabalhos que solicitei, agradeço a equipe de bibliotecários documentalistas da Biblioteca da UFABC, que conseguiram muitos trabalhos, via COMUT, inclusive um artigo de morfologia clássica escrito em bósnio... Agradeço a Nishon Hawkins, Anschutz Library - Kansas University por disponibilizar a Tese de Eric Wild (1997), à Renata Grun da Biblioteca de Geociências da Universidade Federal do Rio Grande do Sul por disponibilizar a Dissertação de Stela Máris Pires Gayer (1984), a Dra. Christine Appia da Societe Geologique de France, ao Prof. Dr. Ulyses F. J. Pardiñas do Instituto de Diversidad y Evolución Austral, a Annemarie Ohler do Muséum National d'Histoire Naturelle, ao pessoal do grupo do Facebook WikiPaleo e WikiHerps que me forneceram alguns artigos importantes. Agradeço ao Martín L. Saleme do Centro de Información Geo-Biológico del NOA, Fundación Miguel Lillo. Agradeço especialmente ao Dr. Kleber S. Vieira pela disposição em fornecer artigos, informações acerca de espécimes e pesquisadores que se dedicaram ao estudo dos ceratophryideos. Agradeço a plataforma morphosource.org por disponibilizar arquivos de tomografia gratuitamente e a todos museus envolvidos neste esforço. Agradeço especialmente aos professores: Prof. Dr. Ivan Sergio Nunes Silva Filho, ao Prof. Dr. Ricardo Jannini Sawaya

e ao Prof. Dr. Helio Ricardo da Silva por todas as sugestões e críticas que fizeram na Qualificação e Defesa desta Dissertação.

Agradeço imensamente a Profa. Dra. Vanessa Kruth Verdade por ter apostado em mim e nas minhas ideias, por ter me ajudado em várias etapas e me aconselhado cientificamente. Gostaria de agradecer ao meu coorientador Prof. Dr. Charles Morphy Dias dos Santos por ter cedido a estrutura do Laboratório de Sistemática e Diversidade e pelos auxílios ao longo do mestrado. Agradeço especialmente ao Dr. Alessandro Marques de Oliveira egresso da UFABC e membro do Espeleo Grupo de Rio Claro, coletor do fóssil objeto foco desta Dissertação. Ao pessoal do Laboratório de Evolução e Diversidade por toda ajuda, especialmente ao hermano Me. Diego Almeida-Silva e pela amizade, parceria, e por toda a força que me deu ao longo do desenvolvimento deste trabalho e a Me. Adriana Hiromi Yukimitsu, e Bel. Michaella Pereira Andrade pela amizade e por toda força e energia boa que me deram para continuar no caminho. Agradeço a Me. Lucy Gomes de Souza pela amizade, pelos vários toques científicos, por ser minha orientadora da vida, e por ser um exemplo de pessoa e profissional que quero seguir! Agradeço aos meus familiares e amigos, em especial a Maria de Lourdes de Almeida Barcelos, minha mãe que acredita e investe nos meus sonhos, mesmo o Brasil não sendo o melhor lugar para se almejar a carreira de cientista, devido aos vários impedimentos culturais e políticos. Agradeço a minha namorada e companheira Tamara Jarosi Handajevsky por estar sempre a meu lado, e dividir os bons momentos e os difíceis também. Agradeço à Tatiana Ivanovna Handajevsky (Baba) e Vladimir Handajevsky (Dieda), por serem minha família na grande São Paulo.

Este trabalho se concretiza devido ao apoio de todos vocês, muito obrigado!

O presente trabalho foi realizado com apoio da Coordenação de Aperfeiçoamento de Pessoal de Nível Superior – Brasil (CAPES) – Código de Financiamento 001.

“São destemidos e reagem á qualquer attaque,

correndo de bocca aberta atraz de seus perseguidores;

e emittem nesta occasião um grito

algo parecido com o choro de uma creança.

Apanham tudo quanto lhes cáe ao alcance e que se mova.”

Miranda-ribeiro sobre Ceratophrys aurita

Miranda-ribeiro (1920:797)

"l'avenir de la systematique des Amphibiens git, pour une large part, dans la connaissance de leur squelette." “o futuro da sistemática dos anfíbios reside, em grande parte, no conhecimento de seu esqueleto” (Laurent, 1942:6)

“[…] a vida não é um game Mas passa o tempo e sobe o nível.” Insanidade – PrimeiraMente

Resumo

Descrevemos uma nova espécie fóssil pertencente ao gênero vivente Ceratophrys, baseados na morfologia comparada do crânio. A nova espécie Ceratophrys sagani, proveniente da caverna Versalles (Pleistoceno superior/Holoceno inferior) do sudeste de São Paulo-Brasil, diferencia-se das outras espécies pela presença de dois forâmens formados na sutura entre o esfenetmoide e o vômer, em vista ventral; eminências epióticas com uma concavidade cotiloide, parcialmente recoberta pelo processus posterior do frontoparietal; Pars interna plectri da columela com três cristas curvas. Nossa análise filogenética inclui todas as espécies viventes de Ceratophryidae (exceto C. testudo) e seis espécimes fósseis. Recuperamos C. ameghinorum no grupo de espécies cornuta, C. rusconii como grupo-irmão de todas as outras espécies de Ceratophrys, Ceratophrys sagani como grupo-irmão de C. aurita + C. cornuta BMNH 18895 e C. cornuta BMNH 18895 como um espécime fóssil de C. aurita. Corroboramos C. ornata MLP 86.VIII.1.4 como um espécime de C. ornata, e L. australis como uma espécie extinta de Lepidobatrachus. Comentamos a respeito da paleodistribuição e a retificação taxonômica de alguns especimes fósseis de Ceratophryidae.

Palavras-chave: Lissamphibia; Paleontologia; Sistemática; PETAR

Abstract

We describe a new fossil species belonging to the extant genus Ceratophrys, based on skull comparative morphology. The new species Ceratophrys sagani, from late Pleistocene/early Holocene os Southeastern São Paulo-Brazil, is distinguished from the other species in the presence of two foramina formed on the suture of sphenethmoid with vomer, in ventral view; epiotic eminences of exoccipital with a cotyloid concavity, partially covered by processus posterior of frontoparietal; Columella pars interna plectri with three curved crests. Our phylogenetic analysis includes all extant species of Ceratophryidae (except C. testudo) and six fossil specimens. We recovered C. ameghinorum in the cornuta species group, C. rusconii as sister to all Ceratophrys species, Ceratophrys sagani as sister clade to C. aurita plus C. cornuta BMNH 18895, C. cornuta BMNH 18895 as a C. aurita fossil specimen, we corroborate C. ornata MLP 86.VIII.1.4 as a C. ornata fossil specimen, and L. australis as a Lepidobatrachus extinct species. We comment on the paleodistribution and on the taxonomic rectification of some Ceratophryidae fossil specimens.

Key-words: Lissamphibia; Paleontology; Systematics; PETAR

SUMÁRIO

Introdução...... 14

Capítulo 1

Introduction ...... 15

Material and Methods ...... 16

Results ...... ,...... 18

Discussion ...... ,...... 27

References ...... ,...... 30

Figures ...... 37

Tables ...... 48

Appendix I ...... 51

Appendix II ...... 52

Capítulo 2

Introduction ...... 53 Material and Methods ...... 54 Results ...... 56 Discussion ...... 59 References ...... 67 Figures ...... 76

Appendix I ...... 79

Appendix II ...... 80 Appendix III ...... 96 Conclusão ...... 100 Atividades realizadas durante o mestrado...... 102

INTRODUÇÃO

A seguir encontram-se os resultados finais das atividades que realizei ao longo de meu mestrado. Planejamos apresentar a Dissertação em dois capítulos, que correspondem a dois manuscritos que serão submetidos à publicação. O Capítulo 1 versa a respeito da descrição de uma nova espécie de Ceratophryidae fóssil, com comentários sobre a paleodistribuição da família. Os resultados deste trabalho foram recentemente apresentados no XIX Congreso Argentino de Herpetologia. O Capítulo 2 concerne na análise filogenética da família Ceratophryidae incluindo todas as espécies viventes (exceto C. testudo) e seis representantes fósseis da família, juntamente a comentários a respeito da taxonomia de alguns espécimes fósseis de Ceratophryidae. Optamos por apresentar o texto da Dissertação na formatação requerida pelo periódico que pretendo submeter após revisão das sugestões da banca (Zootaxa). 15

CAPÍTULO 1

Description of the first Brazilian Ceratophrys fossil species (Anura: Ceratophryidae) from late Pleistocene-early

Holocene of Versalles Cave, Apiaí, São Paulo, Brazil

INTRODUCTION

Ceratophryidae is one of the most frequent anuran clade found in the fossil record. The family is recovered as a well supported clade in most recent phylogenetic hypotheses proposed from molecular data sets (e.g. Frost et al. 2006; Pyron & Wiens 2011; Feng et al. 2017; Sabbag et al. 2018; Streicher et al. 2018) and analysis of total evidence using morphological and molecular data (Faivovich et al. 2014). The living species of Ceratophryidae (sensu Favovich et al. 2014; Frost 2019) are arranged in three genera: Ceratophrys (8 spp.), Chacophrys (1 sp.) and Lepidobatrachus (3 spp.). The Ceratophrys is the most diverse genus presenting two species groups: The C. cornuta group, including C. calcarata, C. testudo, C. cornuta, and C. stolzmanni; and the C. aurita group including C. aurita, C. cranwelli, C. joazeirensis, and C. ornata (Lynch 1982; Faivovich et al. 2014).

Ceratophryidae are Neotropical anurans characterized by a robust, medium to large body size (SVL — snout-vent length 40–50 mm) (Evans et al. 2008; Fabrezi 2006), short limbs, broad skull, adductor muscles of mandible inserting in a large bony surface, disproportionately large jaws in relation to SVL, nonpedicellated and fanglike teeth in the maxilla and premaxilla (Fabrezi 2006). The skeleton is hyperossified (Wild 1997a), resulting from peramorphic heterochrony (Reilly et al. 1997; Fabrezi 2006).

The osteology of Ceratophryidae is well known, available in long term academic works (e.g. Lynch 1971; Gayer 1976; Perí 1994; Wild 1997b; Vieira 2012), and in many papers (e.g. Reig & Cei 1963; Barrio 1968; Lynch 1982; Gayer 1984; Wild 1997a; 1997b; 1999; Vieira et al. 2006). Many authors accessed the taxonomy and systematics of the family from adult (Lynch 1982; Perí 1994; Wild 1997b) and larval (Lynch 1982; Wild 1997b) phenotypic characters and from molecular markers (Faivovich et al. 2014).

Recent works changed the time origin of Ceratophryidae (e.g. Feng et al. 2017; Hutter et al. 2017) from late Cretaceous to late Miocene, and phylogenetic analyses including 16 fossil species corroborated the new scenario (e.g. Nicoli et al. 2016; Báez & Gómez 2017). Baurubatrachus pricei and Beelzebufo ampinga (late Cretaceous) and Wawelia gerholdi (early Miocene) were known as the most ancient representatives of Ceratophryidae, but during the last years their phylogenetic allocation was consistently doubted (e.g. Agnolin 2012; Faivovich et al. 2014). Nicoli et al. (2016) recovered in their analysis B. pricei and W. gerholdi as non-Ceratophryidae, and B. ampinga as Ceratophryidae. Báez & Gómez. (2017) recovered none of them as members of Ceratophryidae. Therefore, the older Ceratophryidae species currently accepted is Ceratophrys sp. MD-CH-06-165 from the Arroyo Chasicó Formation, Buenos Aires Province - Argentina and dates back to late Miocene (Nicoli et al. 2017).

There are five Ceratophryidae fossil recognized as new species: Ceratophrys ameghinorum Fernicola 2001, C. ensenadensis Rusconi 1932, C. prisca species inquirenda Ameghino 1899, C. rusconii Agnolín 2005 and Lepidobatrachus australis Nicoli 2015; and nearly thirty Ceratophryidae fossil determined only to genus level (see Günther 1859; Perí 1993; Marshall & Patterson 1981; Stoessel et al. 2008; Vergnaud-Grazzini 1968; Rinderknecht 1998; Fernícola 2001; Pardiñas 2001; Mercadal de Barrio & Barrio 2002; Turazzini 2015; Nicoli 2014, 2016, 2017; Nicoli et al. 2017; Tab. 1). Herein we describe a new fossil species belonging to the genus Ceratophrys, C. aurita group, based on skull comparative morphology.

MATERIAL AND METHODS

The fossil material is a well-preserved, almost complete skull showing no signs of deformation found with Megafauna fossils and other Anura remains, all disarticulated and fragmented. We removed mechanically the mud and carbonatic incrustation covering the skull. The age of the fossil is unknown. However, Oliveira (2018) dated Nothrotherium maquinense found in the same paleontological expedition and cave. The calibrated age obtained for the fossil remains of N. maquinense is between 13,037 and 12,790 BP (before present). The experiment to obtain the age of the fossil ZUFABC 037 and other Anura remains is currently in course in the Electron Paramagnetic Resonance X-band FCF (Faculdade de Ciências Farmacêuticas) -Araraquara, UNESP (Universidade Estadual Paulista Júlio de Mesquita Filho). 17

The fossil skull was measured with a Vernier caliper to the nearest of 0.01 mm on fossil specimens and ImageJ software on µCT-scanned specimens. The morphometric measurements follow Reig & Cei (1963), with the addition of diameter of supratemporal fenestra (U): length of the condylepremaxillae (A), maximum skull diameter anteroposteriorly (B), maximum skull transverse diameter (C), maximum diameter between otic flange (D), distance from the posterior edge of the orbit to the posterior edge of the squamosal (E), diameter of anteroposterior orbit (F), maximum diameter bi-orbital (G), maximum diameter bi-narinal (H), maximum diameter bi-premaxillary (K), maximum diameter inter-pterygoids (L), length of suspensorium projection (N), length from the quadrate to the anterior edge of premaxillae (P), length of the posterolateral coanae (Q), minimum interorbital diameter (R), height of premaxillae alary process (S), maximum cranium height (T) (Tab. 2). The osteological nomenclature follows Lynch (1971) and Wild (1997b).

We compared skull morphology to the skull of at least one specimen from each Ceratophryidae living species, except C. testudo, and with fossil specimens of Ceratophryidae (Appendix I). We prepared and studied specimens under a stereomicroscope Zeizz Stemi V11, besides the direct study of specimens, we used µCT-scanning images provided by the Laboratory of Computerized Tomography at Museu de Zoologia da USP (MZUSP). The µCTscanning images from MZUSP were prepared in v tome x m microfocus µCT scanner Version 2.3.0.1032 (General Eletric Company, Wunstorf, DE). All specimens were scanned using a tungsten target, a background medium of air, no filter and were rendered as 16-bit TIFFs. The scan data were analyzed in the free viewer software MyVGL (Volume Graphics, Heidelberg, DE) and 3DSlicer software, version 4.10.1 (Fedorov et al. 2012). Photos of the fossils were taken with a LGK10 cell phone 13 MP, f/2.2, AF (LTE model). The specimens were measured directly with a Vernier caliper to the nearest of 0.01 mm and through ImageJ Fiji software when measures were taken from µCT-scanned images and photos of specimens. 18

RESULTS

Systematic palaeontology Ceratophrys sagani (Figures 1, 2, 3a, b and 4a) Class Amphibia Linnaeus, 1758 Order Anura Duméril, 1805 Family Ceratophryidae Tschudi, 1838 Genus Ceratophrys Wied-Neuwied, 1824. Type Species Ceratophrys varius Wied-Neuwied, 1824 (currently C. aurita [Raddi 1823]). Extant taxon, Neotropical distribution. Ceratophrys sagani Barcelos, Santos, Almeida-Silva & Verdade

Holotype (ZUFABC 037): Nearly complete skull, lacking the right premaxilla and right columella.

Type locality: Versalles cave, Apiaí municipality, State of São Paulo, Brazil UTM 22J 737457/7294530, 715m (Datum WGS) (Oliveira 2015). Ceratophrys sagani was collected in the Versalles cave, inside a private area ownership of Purical Mining Company, approximately 16 km northeast of the city of Apiaí, São Paulo State (Fig. 5). The area belongs to the Alto do Ribeiro region, part of the austral portion of Ribeira Belt (De Almeida et al. 1973), one of the three geotectonic provinces that border San Francisco Craton, part of the Brazilian Atlantic Shield (Theodorovicz 2014). The area presents typical geomorphological and speleological features, with high speleomorphological variety (more than 200 recognized caves) and important paleontological sites of the Quaternary (Karmann & Ferrari 2002). The caves at that region present estimated age between 2 – 1.7 million years ago (Karmann & Ferrari 2002). The sediments found in these caves present evidence of reworking of the stratigraphic horizon, made by floods (Dias Neto et al. 1984). The sediments present low maturity and low degree of selection, reaching several sizes of granulometry in consonance with decomposing organic matter, blackish coloration and carbonate cementation at some levels (Dias Neto et al. 1984). The fossils found are usually disjointed and well fragmented due to the high degree of reworking (Ghilardi et al. 2011).

Diagnosis: Diagnosable as Ceratophrys by the combination of characters 1) palatal shelf of premaxilla lacking; 2) facial lobe of premaxillae very deep; 3) palatal shelf of maxilla not evident anteriorly, posteriorly forming a large pterygoid process fused to pterygoid; 4) nasals large, fused medially; 5) nasal fused to maxilla, nasal-pterygoid relationship not evident; 6) 19 nasals broadly fused to frontoparietals; 7) temporal arcade forming supratemporal fenestrae, temporal arcade notched posteriorly; 8) epiotic eminences prominent, concealed dorsally by temporal arcade; 9) cristae paroticae narrow and long. And diagnosable as a new species in the genus by presenting 1) two foramina formed by the suture of sphenethmoid with vomer in ventral view; 2) epiotic eminences of exoccipital with a cotyloid concavity, partially covered by processus posterior of frontoparietal; 3) Columella pars interna plectri with three curved crests; anteriormost portion of sphenethmoid wide and rounded.

Derivation of name

Specific epithet is, sagani, in genitive singular, in honor to Carl Sagan, who did much for Scientific communication and inspire the future generations of scientists.

Distribution: Known only from the type locality.

Fossil Description

Ceratophrys sagani (ZUFABC 037) is an adult based on the hyperossification of the skull, with synostosis of skull bones.

The sphenethmoid is in the central cranium axis, it is not exostosed and presents three portions, which articulates conspicuously to adjacent bones. The anterior one is a wide and rounded projection directed anteriorly (Fig 3a); below this projection is located two olfactory nerve foraminae (I). Dorsally, the back of the anterior portion articulates with the anterior edge of nasals. Laterally, the back of sphenethmoid articulates with the pars facialis of maxilla. The sphenethmoid articulates with anterior edge of vomer, forming a medial foramen (Figs. 1b; 3b). The medial portion of the sphenethmoid is reduced, triangular, and slightly concave. It contacts synostotically the vomer antero and laterally, and the palatine, posteriorly, at the level of the planum antorbitale. The posterior portion of the sphenethmoid is a bifurcated element, with broad laminae. The anterior portion of the laminae bears an orbitonasal foramen, and the posterior portion of the laminae bears an optic nerve foramen (II). It articulates anteriorly with the proximal portion of palatine, laterally with the lamina perpendicularis of frontoparietals, and posteriorly with otoccipital. Its lateral articulation with the anterior edge of cultriform process of parasphenoid occurs along its length. All sutures are obscured by the extensive synostosis. There is no dorsal exposure of sphenethmoid. 20

The prootics and the exoccipitals are continuous and indistinguishable forming an unite characterizing the otoccipital (Lynch 1971). It is composed by a thin solid wall, but spongy internally. Ventrally, it articulates with the parasphenoid alae and the medial ramus of pterygoids. Dorsally, it articulates with the frontoparietal lamina perpendicularis. Anteriorly, the prootics articulates with the posterior portion of sphenethmoid, where the optic nerve foramina (II) is formed. The sutures between those bones are obscured by synostosis. The posterior portion of otoccipital forms the posteromedial parts of the auditory capsules and the occipital condyles that are juxtaposed (Type II; Lynch 1971), massive, and lack a constricted base, its articular surfaces are rounded and ventromedially to dorsolaterally angled. Above the occipital condyle there are the jugular foramina (IX, X and XI), and the auditory nerve foramina (VII). Above the foramen magnum, there is the epiotic eminences, which is a triangular protuberance in dorsal view, and in posterior view, present a cotyloid concavity partially covered by the processus posterior of frontoparietals.

The plectral apparatus, also known as columella is a slender, cylindrical bone, enlarged at its proximal edge. It is located below the otic ramus of the squamosal and associated with the fenestra ovalis of the otoccipital. Three portions are recognized from proximal to distal end: pars interna plectri, pars media plectri, and pars externa plectri. The pars interna plectri is composed of a complex trochanter or apophysis, which presents a concavity at its ventral surface, and three curved crests (Fig. 6). One crest is dorsal, initiating in the posteromedial portion of pars interna plectri and extending until its anterodorsal end, one crest is ventral, following the same conformation of the first one, but forming one of the axis which structure the concavity, and the third crest extends through the posteromedial portion of collumella pars interna plectri until the ventral axis structuring the concavity, forming an accentuated curve. The pars media plectri and the pars externa plectri forms together a distal anterolateral stylus. The pars externa plectri is not entirely preserved in the fossil.

The frontoparietals are paired, symmetrical, wide, and heavily ossified bones fused to each other all along their contact. There is a tuberculated ornamentation on its dorsal surface. Dorsally, there is an interocular crest along the frontoparietals suture, which become clearer between the medial portion of interocular region and the anterior edge of frontoparietals, where they articulate to the posterior portion of nasals. The articulation between the frontoparietals and the nasals form part of the infraocular flanges. The posterior portion of frontoparietal invests the otic ramus of squamosal dorsally, forming the 21 supraorbital flanges. The posterior-most portion of frontoparietal invests the exoccipital posteriorly and cover partially the epiotic eminences. At this posterior portion, the frontoparietal process present acute edges. The posterior-most portion of frontoparietals articulates synostotically with the otic ramus of squamosals, forming the crista parotica, which is the posterior flange of supratemporal fenestrae. Ventrolaterally, along the entire length of frontoparietal, there is a lamina perpendicularis in the dorsolateral region of the braincase. The throclear nerve foramen (IV) is present in the ventral view, between the orbits and the optic nerve foramen (II).

The nasals are flat, wide, symmetrical and paired bones, which has an anteriorly tapering with dermostosis on its dorsal surface. It articulates synostotically with each other medially. It presents a maxillary process laterally that sutures with the pars facialis of maxilla and the zygomatic ramus of squamosal. Posteriorly, the nasals articulate with the anterior edge of frontoparietals. There is a restrained groove in the posterior extremity, following the suture axis between the nasals. The ventral portion of nasals articulates anteriorly with the back of sphenethmoid producing a suture with low degree of ossification. The posterior edge of nasals forms the infralocular flange, which suture with the zygomatic ramus of squamosals generates a pronounced oblique groove where there is no dermostosis. The groove links the orbits to the choanae. The choanae aperture is oblong and directed to the medial axis of the skull. It is formed anteriorly by the contact between the pars facialis of maxillae and the anterior portion of sphenethmoid, posteriorly by the contact between the maxillary process of nasals and the zygomatic ramus of squamosal, and laterally and posteriorly, by the pars facialis of maxillae, and the nasals. The bones forming the choanae appertures are well ossified and synostotic, exceptionally by the visible suture between nasals and the sphenethmoid.

The parasphenoid is a “T-shaped” bone, formed by two symmetrical laterally oriented alae, and a single anteriorly oriented cultriform process. The alae articulate broadly anteriorly and posteriorly with otoccipital and anterolaterally with the pterygoid. The alae are well developed and present a crest along its anterior edge, resulting from their contact with the medial ramus of pterygoid. There is a flange curved anteroventrally, with the concavity directed posteriorly at the posterior portion of alae, resulting from the articulation of the alae with the posterior portion of otoccipital, where there is the auditory nerve foramina (VIII) on the right concavity and the jugular nerve foramen (IX, I and XI) on the left concavity. There is a protuberance at distal edge of alae, resulting from the alae contact 22 with otoccipital and medial ramus of pterygoid. The cultriform process of parasphenoid articulates laterally with the lamina perpendicularis of frontoparietals, medial portion of sphenethmoid and otoccipital. The articulation between parasphenoid and otoccipital forms two foramina. The optic nerve foramen (II), anterior and the optic-motor foramen (III) posteriorly. The sutures are not clear because of the high degree of synostosis. The vomers are paired bones, anterolaterally oriented, and present an anterior and a posterior portion. The anterior portion of vomers articulate with the sphenethmoid, along its entire width. The suture is not heavily ossified, as observed in all the other skull bones’ sutures. There is a foramen formed by the articulation between the anterior portion of the sphenethmoid and the vomer. It seems to be a new character for Anura. The distal edge of vomers invests the pars facialis of the maxillae. The posterior portion of vomer presents the pars odontoides, a lamina laterally directed, initiating medially and extending until two thirds of the vomer length, as seen in other species of Ceratophryidae. Its extremities are not preserved in the fossil, preventing further description and comparison. The posterior portion of the vomer articulates medially with the palatine, the final edge of the cultriform process of parasphenoid, and the medial portion of sphenethmoid. The articulations are obscured by synostosis. Vomerine teeth or dental ridges are absent.

The septomaxillae are absent in the fossil and it is in the olfactory capsule. The palatines are elongated, laterally oriented, and do not meet medially. The proximal edge of palatines contacts anteriorly the sphenethmoid and vomers with high degree of synostosis. The distal edge of palatines expands and invest the pars facialis of the maxilla ventrally and articulate with the pterygoid dorsally.

The maxillae present three portions: the pars dentalis, pars palatina and the pars facialis. The pars dentalis is compact and bears small hollows internally, corresponding to the pulp cavity and teeth. The teeth are not well preserved, but it is possible to count the teeth root preserved in the fossil. The maxillae could bear at least 40 teeth. Only eight teeth from the right maxilla and nine from the left maxilla are in good preservation condition. The other are worn and deteriorated. The “V-shaped” distal edge of pars dentalis invests the quadratojugal. The pars palatina is reduced, investing dorsally, the pars facialis, and present an internal hollow along its entire length. The pars facialis also presents dermostosis on its dorsal surface. It articulates dorsally with the nasals and with the zygomatic ramus of the squamosals; and ventrally with the anterior portion of sphenethmoids, lateral edges of vomers, palatine and pterygoids. 23

The premaxillae are pared bones. Only the right premaxilla is preserved in the fossil. The pars dentalis of premaxilla presents ten pedicellate teeth, but only seven are well preserved. The pars facialis present dermostosis on its dorsal surface. The alary process is perpendicular to the pars dentalis axis, but slightly divergent. The alary process of the premaxilla contacts ventrally the anterior portion of sphenethmoid. The distal edge of the alary process is not preserved in the fossil. The maxillary ramus of the premaxilla is long, “V-shaped”, and contacts ventrally the pars palatina of maxillae.

The pterygoids are paired triradiate, laminate and symmetrical bones. It presents three well developed rami, the anterior, medial and posterior rami. The anterior ramus articulates laterally with the pterygoid process of the maxillae in broad contact, and, medially, with the distal edge of the palatine and the planum antorbitale of the sphenethmoid. The anterior ramus forms the lateral edge of interpterygoid vacuity. The medial portion of anterior ramus of pterygoid presents a dorsoventral concavity along its length, which become plain at its distal edge. The medial ramus of the pterygoid is a tapering portion which articulates diagonally with the distal edge of parasphenoid alae and with the ventrolateral edge of the otoccipital and forms the posterior edge of interpterygoid vacuity. The posterior ramus of pterygoid is a laterally curved portion, which articulates laterally with the medial edge of the ventral ramus of the squamosal and invests the pars articularis of the quadrate posteroventrally.

The squamosals are paired well developed bones, presenting dermostosis on its dorsal surface. The squamosals present three rami. The ventral ramus is wide, elongated and flat antero-posteriorly, present a posterolateral orientation and it is concave anteriorly. It articulates synostotically with the posterior ramus of pterygoid, with the quadrate and maxilla. Anteriorly, the otic ramus articulate with the lateral edge of the frontoparietal forming the anterior edge of the supratemporal fenestra. The otic ramus present a medially directed projection, which articulates the otoccipital forming the crista parotica. Ventrally, this projection participates in the configuration of the auditory capsule. The otic ramus of squamosals also presents the otic plate (sensu Lynch 1971), a sub-quadrangular bone projection, posterolaterally directed which is well expanded and tapering in dorsolateral view, following the axis of three squamosals convergent crests. The otic plate is highly curved dorsolaterally and presents a dorsal concavity. 24

The zygomatic ramus of the squamosals articulate with the pars facialis of the maxillae, and with the anterior ramus of the pterygoid, anteriorly. Medially, it articulates with nasals and with the pars facialis of the maxillae. From these contacts, two well developed crests are formed. The most anterior crest, called lateral crest, results from the contact among the bifurcated anterior ramus of pterygoid, the pars facialis of the maxilla and the anterior edge of the zygomatic ramus of the squamosal. The other crest, oblique crest, is formed by the contact between the zygomatic ramus of squamosal and the nasal, this crest begins at the anterolateral flange of the orbit and extends, diagonally oriented, until it connects with the lateral crest. The medial portion of the otic ramus of squamosal articulate with the frontoparietal and forms the supraorbital flange. At this bone contact another well- developed crest arise, the supraorbital crest. The supraorbital crest is laterally oriented and fuses with the other crests of the zygomatic ramus of squamosals at this distal portion, originating a crest that follows the axis of the lateral edge of the otic plate of otic ramus of squamosal. There are two depressions in the orbital arch between the crests of the zygomatic ramus.

The quadrate is a discrete bone, with a medial constriction and expanded extremities. It is heavily synostosed with the quadratojugals, the posterior ramus of the pterygoids and with the ventral ramus of the squamosals, the articulations are obscured by synostosis. There is a condyle in the medial portion of quadrate, result of the articulation with the posterior ramus of the pterygoid and posterolaterally with the ventral ramus of the squamosals. The suture between the posterior portion of pars palatina and pars facialis of maxilla, quadratojugal, ventral ramus and zygomatic ramus of squamosal forms the subtemporal fenestra, an ellipsoid opening at the lateral of the cranium. The quadratojugals are paired bones, massive and “V-shaped”, in lateral view. Its anterior portion is cylindrical and articulates anteriorly with the pars dentalis and pars palatina of maxillae and its posterior portion articulates with the quadrates and with the squamosals. 25

Comparison with living species of Ceratophrys

The comparison of the fossil skull of Ceratophrys sagani with the skulls of the living species of Ceratophryidae indicated that Ceratophrys sagani present all diagnostic characters of the Ceratophryidae and Ceratophrys. The comparative anatomy using the Ceratophrys sagani and the living and fossil species of Ceratophrys revealed the presence of unique characters that justify the determination of the fossil as a new species, probably belonging to the C. aurita group.

The C. cornuta species group present elongated and subtriangular otic plates (except C. stolzmanni) and species of the C. aurita group present sub-quadrangular otic plates. The otic plate of Ceratophrys sagani is sub-quadrangular, but differs in format from all species in the C. aurita group, except for C. aurita species itself.

The squamosals of Ceratophryidae are well developed and in Ceratophrys, all three rami are broad and elongated. However, the zygomatic ramus varies in width among species. Ceratophrys ornata and C. joazeirensis present a narrow zygomatic ramus, while Ceratophrys sagani and C. aurita, specially, present wider zygomatic ramus and crests on its dorsal surface. Additionally, C. aurita and Ceratophrys sagani share the presence of two depressions in the orbital arch between the crests of the zygomatic ramus of squamosal.

In dorsal view, the epiotic eminences of Ceratophrys sagani resembles the C. aurita elliptical ones, differing from the slender ones of C. joazeirensis (Vieira et al. 2006), In posterior view, the posterior-most surface of epiotic eminences of all Ceratophrys species are flat, in some species there are dorsolateral protuberances, recorded here in some specimens of C. aurita, C. calcarata, C. cornuta and C. joazeirensis. In Ceratophrys sagani the dorsolateral protuberances are absent, and this fossil species is unique by presenting a concavity on the posterior surface of epiotic eminences, a morphological novelty for Ceratophryidae.

Ceratophrys sagani present the anteriormost portion of sphenethmoid wide and rounded while, C. joazeirensis and C. cranwelli presents this portion narrow and rounded. This portion is wide and flat in C. aurita and narrow and flat in C. cornuta (Figs. 3a, b). Ceratophrys sagani differs from all other Ceratophryidae by bearing two foramina on the suture between the anterior portion of the sphenethmoid and the anterior edge of vomer. Our comparison indicates it is a morphological novelty for Ceratophryidae. Topologically, at this 26 suture we noted a concavity in C. joazeirensis (MZUSP 142284) and a discreet crest in C. aurita (cas:herp:84998) specimen (Fig. 3c).

The Ceratophrys. sagani alary process of premaxilla is not as narrow as in C. joazeirensis, resembling C. aurita and C. cranwelli. There is a rounded dermostosis at the external surface of pars facialis of premaxilla (near the alary process axis) in Ceratophrys sagani; a similar dermostosis is sometimes recorded in C. aurita specimens (see Gayer 1984). Ceratophrys sagani and C. joazeirensis present crests on the pars interna plectri of the columella, C. joazeirensis presents only the ventral crest. Ceratophrys sagani differs from all other Ceratophryidae by presenting three crests on the pars interna plectri of columella (two ventral and one dorsal; Fig. 6).

Comparison with fossils of Ceratophryidae

Ceratophrys sagani is the second fossil species in size, being, C. ameghinorum (MACN 14318; Fig. 7), the largest. Ceratophrys sagani is larger in six bone length measurements, though: H) Maximum bi-narial diameter (6.9%); K) Maximum bi-premaxillary diameter (3.04%); R) Minimum interorbital diameter (10%); S) Height of premaxillae alary process (9,16%); T) Maximum cranium height (5%); and U) Diameter of supratemporal fenestrae (18,75%). Ceratophrys sagani differs from C. ameghinorum by presenting a curved shape of the posterior edge of otic plate (straight, well-developed and dorsally projected in C. ameghinorum).

Ceratophrys sagani and C. rusconii (MACN 19744; Fig. 8) present supratemporal and subtemporal fenestrae, epiotic eminences partially covered by processus posterior of frontoparietals and subquadrangular otic plate of squamosal and differs by the presence of a well-developed and ornamented anterior portion of sphenethmoid in Ceratophrys sagani (reduced in C. rusconii); the otic plate of Ceratophrys sagani is curved dorsally at its distal edge (concave ventrally in C. rusconii, resembling C. ornata).

Ceratophrys sagani resembles the Ceratophrys fossil specimen (BMNH 18895), from Pleistocene of Lagoa Santa (Minas Gerais, Brazil) erroneously classified by Günther (1859) as Ceratophrys cornuta, proposed as close related to [C. aurita-C. joazeirensis] clade by Faivovich et al. (2014). The fossil classified by Günther (1859) urge revision and will be referred as Ceratophrys cf. C. aurita (BMNH 18895). Ceratophrys cf. C. aurita (BMNH 18895) resembles C. aurita due the presence of the following characteristics: posterior 27 margin of dermocranium indented, supratemporal fenestrae, sub-quadrate otic plate of squamosal, three crests on the squamosal and maxilla.

Additionally, Ceratophrys sagani differs from Ceratophrys cf. C. aurita (BMNH 18895), C. ameghinorum, C. rusconii and all other Ceratophrys fossil species by the presence of epiotic eminences surface with concavity in posterior view; columella pars interna plectri with three curved crests; the anterior-most portion of sphenethmoid wide and rounded; and foramen on the contact of vomer with sphenethmoid.

Taxonomic attribution

Ceratophrys sagani is assigned to the genus Ceratophrys based on the following characters: Posterior margin of dermocranium indented, supratemporal fenestrae, and fusion of most skull bones into an akinetic unit (Lynch 1971; 1982). The presence of sub- quadrangular supratemporal fenestrae allow assignment of Ceratophrys sagani to the C. aurita species group. Additionally, due the share of characteristics presents in both C. aurita species (e.g. presence of crests and depressions on the maxilla and squamosal), and C. joazeirensis species (e.g. epiotic eminences with a slender format) Ceratophrys sagani could be hypothesized as close related to those two species.

Ceratophrys sagani is assigned as a new species based on combination of the following characters: two foramina formed on the suture of sphenethmoid with vomer in ventral view; the anteriormost portion of sphenethmoid wide and rounded, epiotic eminences of exoccipital with a cotyloid concavity, partially covered by processus posterior of frontoparietal; Columella pars interna plectri with three curved crests.

DISCUSSION Ceratophrys sagani represent a new Ceratophrys fossil species close related to C. aurita species group, based on the presence of all the diagnostic characters proposed for the genus, and for being unique among the Ceratophryidae species by presenting: 1) two foramina formed on the suture of sphenethmoid with vomer, in ventral view; 2) epiotic eminences of exoccipital with a cotyloid concavity, partially covered by processus posterior of frontoparietal; 3) Columella pars interna plectri with three curved crests. Due the systematic update of Baurubatrachus pricei as not related to Ceratophryidae (Nicoli et al. 2016; Báez & Gómez 2017), Ceratophrys sagani could be recognized as the first fossil species of Ceratophryidae and the second fossil specimen of the family in Brazil. 28

Contribution to character variation Despite many authros have studied Ceratophryidae skull morphology there are still some not completely explored sets of characters. The sphenethmoid anterior portion is recognized by Gayer (1976) and Perí (1994) and is absent in Wild (1997b) description, that recognizes the anterior portion of sphenethmoid as part of vomer. Perí (1994) recognizes this anterior portion as well developed in Ceratophrys and Lepidobatrachus, and the presence of this morphology maybe be a result of a densely ossified skull (Gayer 1976). The sphenethmoid and vomer are fused in Ceratophryidae, and some species present ornamentation on this region, however this is not recognized in the literature. Through the aid of the µCT-scan we could apply lower bone density values on the Ceratophrys specimens under analysis, and we found these foramina are present, internally, in some Ceratophrys species but were covered by extensive exostosis. The species under analysis which present this characteristic are: C. aurita, C. joazeirensis, and C. cornuta., in the same region C. aurita and C. joazeirensis present those ornamentations. These foramina maybe could be accessory nerve foramina result of a larval character retention; an investigation is in course to better understand the function of this concavity in C. aurita juvenile specimens.

The epiotic eminences of the otoccipital are probably a bone surface for muscle insertion (Gayer 1984). The variation present on it was not well accessed in the literature, except for Vieira et al (2006), differing C. joazeirensis and C. aurita in dorsal view. In dorsal view, C. aurita, C. calcarata, C. cornuta and C. joazeirensis present dorsolateral protuberances, yet not described in the literature. In posterior view, the epiotic eminences surfaces are flat in Ceratophryidae species, except for Ceratophrys sagani differing from all others by presenting a concavity on the posterior surface of epiotic eminences (Fig. 2d).

The columella morphology known for Ceratophryidae is a pars interna plectri concave/globous, without any ornamentation (e.g. Lepidobatrachus and Chacophrys), with an anterior and posterior plectral apophysis (e.g. C. cranwelli and C. ornata) or a ventral crest (C. joazeirensis). The development degree of the apophyses depends on the age or gender of Ceratophrys specimens (Perí 1993). Ceratophrys sagani is unique among Ceratophryidae by presenting a pars interna plectri ornamented with three curved crests (Fig. 5).

Ceratophrys fossils distribution Ceratophrys sagani represents one of the most northern records of Ceratophrys fossil species (Fig. 9). The great majority of Ceratophrys fossils are located in the southern South America. We could hypothesize three possible explanations for that paleodistribution: 1) the ancient distribution of Ceratophrys species were more southern in the continent; 2) the environment these species lived in northern lands forbid fossil preservation (e.g. Amazonian and Atlantic Forests); or 3) maybe in Argentina there are more Anura fossil collection effort. The distribution of the majority of the fossils belonging to Ceratophrys are included inside the distribution of living species of the genus, specifically the distribution of C. aurita group. However, there are some outliers: Ceratophrys sp. (MLP 247, late Miocene-early Pliocene), Ceratophrys sp. (MNHN 1560, Pleistocene), Ceratophrys sp. (Vergnaud- Grazzini 1968, Pleistocene), Ceratophrys sp. (MD-CH-06-165) located west of the distribution of the living species Ceratophrys cranwelli, and Ceratophrys sp. (PVSJ 284, late Miocene) located south of distribution of C. ornata. The distribution of all Lepidobatrachus fossil specimens, i.e. Lepidobatrachus sp. (IANIGLA-PV 112, late Miocene), Lepidobatrachus australis (MMH FMH 85-12-2a, Pliocene) and Lepidobatrachus sp. (GHUNL-Pam 8633, late Miocene) is more than 250 km southern from the extant species distribution (Fig. 10). Vieira et al. (2018) hypothesized C. cranwelli and C. ornata and Lepidobatrachus species presented a wider distribution during the Last Glacial Maximum (~8,000 BP). The distribution of fossils can corroborate this assertion and is an indicative that, since late Miocene, the paleodistribution of some Ceratophryidae species was different than the current one, maybe fossils close related to Ceratophryidae could have presented a wider distribution, western and southern directed.

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Figure legends

Figure 1: Illustration of Ceratophrys sagani. a) dorsal view; b) ventral view; c) frontal view; d) posterior view; e) lateral view. Abbreviations: c, choana; co, occipital condyles; cp, crista parotica; ee, epiotic eminence fp, frontoparietal; lc, lateral crest; lp, lateral plate; m, maxilla; n, nasal; nf, nerve foramina I-XI; oc, oblique crest; op, otic plate; otc, otoccipital; ps, parasphenoid; pl, palatine; pt, pterygoid; ptf, postemporal fenestra; q, quadrate; qj, quadratejugal; sbf, subtemporal fenestra; sf, supratemporal fenestra; soc, supraorbital crest; sph, sphenethmoid; sq, squamosal; v, vomer.

Figure 2: µCT-scanned images of Ceratophrys sagani in a) frontal view, b) dorsal view, c) lateral view, d) posterior view, and e) ventral view.

Figure 3: Ventral and dorsal view A) Ceratophrys sagani; B) C. aurita; B) C. joazeirensis.

Figure 4: Frontal view A) Ceratophrys sagani; B) C. aurita; B) C. joazeirensis

Figure 5: Location (red marker) of the Versalles cave, Apiaí municipality, State of São Paulo, Southeastern Brazil (UTM 22J 737457/7294530, 715m (Datum WGS)). Gray: Unidades de conservação. PETAR: Parque Estadual Turístico do Alto do Ribeira; PEI: Parque Estadual Intervales; EE Xitué: Estação Ecológica de Xitué.

Figure 6: Distribution of fossil specimens of Ceratophrys, following the data of Table 1.

Figure 7: Distribution of fossil specimens of Lepidobatrachus, following the data of Table 1

Figure 8: Columella of Ceratophrys sagani.

Figure 9: Holotype of the fossil species C. ameghinorum MACN 14318 in dorsal view, ventral view, frontal view, posterior view and lateral view.

Figure 10: Holotype of the fossil species C. rusconii MACN 19744 in dorsal view, ventral view, frontal view, posterior view and lateral view. 38

Figure 1

Figure 2

Figure 3

Figure 4 42

Figure 5 43

Figure 6

Figure 7

Figure 8 46

Figure 9

Figure 10 48

Table 1: Fossil specimens of Ceratophryidae n° N° de tombo Taxonomy Geographical Age References Provenance 1 MMH 84.1.15 Ceratophrys ameghinorum Farola de Monte Pliocene Fernícola 2001 Hermoso, Buenos Aires 2 MMP 664.S Ceratophrys ameghinorum Barranca de los Lobos, Pliocene Fernícola 2001 Buenos Aires 3 MMP 892.M Ceratophrys ameghinorum Monte Hermoso, late Miocene- Fernícola 2001 Buenos Aires early Pliocene 4 MMP 664.M Ceratophrys ameghinorum Playa de Los Lobos, Pliocene Fernícola 2001 Chapadmalal 5 MMP 1063.M Ceratophrys ameghinorum Playa de Los Lobos, Pliocene Fernícola 2001 Chapadmalal 6 MLP Ceratophrys ameghinorum Miramar, Barranca Pliocene Fernícola 2001 88.VII.20.1/2 Parodi, Buenos Aires 7 MACN 19731 Ceratophrys ameghinorum Farola Monte Pliocene Fernícola 2001 Hermoso, Buenos Aires 8 MACN 14317-9 Ceratophrys ameghinorum Monte Hermoso, late Miocene- Rovereto 1914 Montehermosense early Pliocene Fernicola 2001 9 MACN 14323-6 Ceratophrys ameghinorum Monte Hermoso, late Miocene- Rovereto 1914 Montehermosense early Pliocene Fernícola 2001 10 BMNH 18895/6 Ceratophrys aurita Lagoa Santa, Minas Pleistocene Günther 1859, this Gerais issue 11 ZUFABC 038 Ceratophrys aurita Versalles cave, São late Pleistocene- Barcelos & Verdade Paulo early Holocene 2019 12 PVL 699 Ceratophrys ensenadensis Olivos, Buenos Aires Pleistocene Rusconi 1932 13 ICNU (MNHN) Ceratophrys ornata Chuquisaca ,Bolivia Pleistocene Vergnaud-Grazzini 1968 14 MMH 85.3.8 Ceratophrys ornata Monte Hermoso, Holocene Pardiñas 2001 Buenos Aires 15 MMH 85.2.11 Ceratophrys ornata Farola de Monte Holocene Fernícola 2001, Hermoso, Buenos Pardiñas 2001 Aires 16 MMH 88.2.5 Ceratophrys ornata Monte Hermoso, Holocene Pardiñas 2001 Buenos Aires 17 MMH 90.2.1 Ceratophrys ornata Monte Hermoso, Holocene Pardiñas 2001 Buenos Aires 18 MLP Ceratophrys ornata G. Chávez, Los Tres late Pleistocene- Perí 1993 86.VIII.1.4 Reyes early Holocene 19 MMP 692-S Ceratophrys ornata Chapadmalal, Buenos Pliocene Reig 1958 Aires 20 MMP 4846 Ceratophrys ornata Daireaux District, late Pleistocene Perez-ben et al. 2019 Buenos Aires 21 unkown Ceratophrys prisca Monte Hermoso, Miocene Ameghino 1899 Buenos Aires 22 MACN 19744 Ceratophrys rusconii Mar Chiquita, Playa Pleistocene Agnolin 2005 Dorada, Buenos Aires 23 ZUFABC037 Ceratophrys sagani Versalles cave, São late Pleistocene- Barcelos et al. 2019 Paulo early Holocene 24 MLP Ceratophrys sp. G. Chávez Laguna Los late Pleistocene- Perí 1993 86.VIII.1.5/6 Tres Reyes early Holocene 25 PVSJ 284 Ceratophrys sp. Ullum Valley, San Late Miocene Contreras & Acosta Juan 1998, Nicoli 2016 49

26 FCS.SA1.M3.59 Ceratophrys sp. San Antonio (Río late Holocene Stoessel et al. 2008 0.1 Colorado) Buenos FCS.SA1.M3.59 Aires 0.2 FCS.SA1.S1.48 6.1 FCS.SA1. S1.506.1 FCS.SA1.S1.52 5.1 FCS.SA1.S1.53 4.1 27 MLP Ceratophrys sp. Miramar, Mar del Sur, late Holocene Frenguelli 1921, 52.IX.27.11 Buenos Aires Fernícola 2001, MB & B 2002 28 MD-CH-06-165 Ceratophrys sp. Arroyo Chasicó late Miocene Urrutia & Rosset 2006, Nicoli et al. 2017 29 ICNU (MZUSP) Ceratophrys sp. Gêmeo Abyss, late Pleistocene- Araújo 2010 Iporanga, São Paulo early Holocene 30 MMH 85.12.10 Ceratophrys sp. Monte Hermoso, late Pleistocene not described Buenos Aires 31 MMH 79.2.3 Ceratophrys sp. Monte Hermoso, late Pleistocene not described Buenos Aires 32 MLP Ceratophrys sp. Olavarría, Arroyo Holocene Mercadal de Barrio & 86.III.25.150/1 Tapalqué Barrio 2002 33 MLP Ceratophrys sp. Gral. La Madrid, Fortín Holocene Mercadal de Barrio & 96.V.18.12 Necochea Barrio 2002 34 MLP 88.V.20.1 Ceratophrys sp. Gral. Pueyrredón, Mar Pleistocene Mercadal de Barrio & del Plata Barrio 2002 35 MACN 18074 Ceratophrys sp. Chapadmalal, Buenos Pliocene Mercadal de Barrio & Aires Barrio 2002 36 MACN 17585 Ceratophrys sp. Arroyo Lobería Pliocene Mercadal de Barrio & Barrio 2002 37 MACN 17936 Ceratophrys sp. Pta. Lobería Pliocene Mercadal de Barrio & Barrio 2002 38 MLP 34.V.10.8 Ceratophrys sp. Punta Varohué Pliocene Mercadal de Barrio & Barrio 2002 39 MLP Ceratophrys sp. Quequén Salado Pliocene Mercadal de Barrio & 94.II.1.171 Barrio 2002 40 MLP Ceratophrys sp. Monte Hermoso late Miocene- Fernicola 2001, MB 48.XII.16.195 early Pliocene & B 2002 41 MACN 14322 Ceratophrys sp. Monte Hermoso late Miocene- Fernicola 2001 early Pliocene 42 MLP 136 Ceratophrys sp. Monte Hermoso, late Miocene- Fernicola 2001 Buenos Aires early Pliocene 43 MLP 247 Ceratophrys sp. Monte Hermoso, late Miocene- Fernicola 2001 Buenos Aires early Pliocene 44 FMNH P 14402 Ceratophrys sp. Corral Quemado, late Miocene- Marshall & Patterson, Catamarca early Pliocene 1981, Báez 1986 45 MNHN 1560 Ceratophrys sp. Colonia, Uruguay late Pleistocene Rinderknecht 1998 46 ICNU (MNHN) Ceratophrys sp. Tarija, Bolivia Pleistocene Vergnaud-Grazzini 1968 47 MMH FMH Lepidobatrachus australis Farola Monte Pliocene Tomassini et al. 2011 85.12.2a Hermoso, Buenos Nicoli 2015 Aires 48 GHUNL Pam Lepidobatrachus sp. Quehué, La Pampa late Miocene Scanferla & Agnolín 8633 2015, Nicoli 2017 49 IANIGLA.PV Lepidobatrachus sp. Fm. Huayquerías, late Miocene Turazzini 2015 112 Mendoza

Table 2: Cranial morphometric measurements of fossil specimens of Ceratophrys. Approximated measurements in red.

Fossil specimen A B C D E F G H K L N P Q R S T U Ceratophrys sp. ZUFABC037 5.2 5.4 7 5.1 1.5 1.1 3.4 2.3 2.3 3.8 2.1 5.8 2 1.1 1.2 3.2 0.8 Ceratophrys sp. MACN 17911 ? ? ? 4.5 1.5 ? 3 ? ? ? ? ? ? ? ? ? 0.6 Ceratophrys sp. MLP 86-viii-1-5 ? ? ? 4.4 1.2 1.2 3 1.8 ? 3.2 ? ? ? 1 ? 2.2 0.4 Ceratophrys sp. MLP 86-viii-1-6 ? ? ? ? ? ? ? ? ? 1.5 ? ? ? 0.9 ? ? ? Ceratophrys sp. MLP 86-viii-1-4 3.1 3.6 5.85 4.6 1.2 1.1 2.98 1.85 1.68 3.1 1.9 3.9 1.8 1.05 ? 2.5 0.51 C. prisca MACN 14323 ? ? ? ? 1.25 1.05 ? ? ? ? ? ? ? 0.9 ? ? 0.52 Ceratophrys sp. MACN 14328 3.6 ? ? 4.2 1.35 1.03 2.8 1.8 1.2 ? ? ? 1.55 0.95 0.9 ? 0.5 C. ameghinorum MACN 14318 3.82 4.72 7.2 5.7 1.89 1.22 3.65 2.14 2.23 4.46 2.34 5.15 2.02 0.99 1.09 3.04 0.65 C. ameghinorum MACN 19731 3.6 3.48 ? 2.6 1.24 1 2.7 1.6 1.6 2.8 1.4 4 1.37 0.95 1.06 2.14 0.59 C. ameghinorum MACN 14317 ? ? ? 4.1 1.2 0.9 2.7 1.4 ? 2.8 ? ? 1.5 0.9 ? ? 0.47

C. rusconii MACN 19744 ? ? 2.8 4 1 1 2.6 1.4 ? 2.8 1.6 ? 1.6 0.9 ? 2.2 0.69

APPENDIX I

Examined specimens. Acronyms are: CM-HERPS (Carnegie Museum of Natural History – Herpetology) Cas-herp (California Academy of Sciences – Herpetology); DNPM-MP (Departamento de Produção Mineral – Museu de Paleontologia); MACN (Museo de Ciencias Naturales de Buenos Aires “Bernardino Rivadavia”); MCZ:Herp (Museum of Comparative Zoology, Harvard University); MLP (Museo de Ciencias Naturales de La Plata); MZUSP (Museu de Zoologia da Universidade de São Paulo); UF-H (Florida Museum of Natural History – Herpetology); USNM (National Museum of Natural History, Smithsonian Institution)

Dry skeletons

Ceratophrys aurita MZUSP 22976* MZUSP 61114(2); DNPM-MP 1-AR.

Cleaned and stained

Chacophrys pierottii MZUSP 16274

Fossils

Ceratophrys† MLP 86-viii-1-5/4, 86-viii-1-5/6, 86-viii-1-5/7; C. ameghinorum† MACN 14317 (paratype), 14318, 19731 (paratype); C. prisca† MACN 14323, C. rusconii† MACN 19744; Ceratophrys sp.† MACN 14328; Ceratophrys cornuta (Günther 1859)† BMNH 18895.

µCTscanned images

Alsodes nodosus CM-HERPS-68395; Ceratophrys aurita MZUSP 31367; and Cas-herp- 84998; C. calcarata MZUSP 99657; C. cornuta MZUSP 70548; uf:herp:63162; MCZ: Herp: A-17499; C. cranwelli MZUSP 99320; MCZ: Herp: A-35360; C. joazeirensis MZUSP 142284; C. ornata MZUSP 70791; C. stolzmanni USNM 160970; Chacophrys pierottii KU191932, MZUSP 99329; Lepidobatrachus asper MZUSP 94595; L. laevis MZUSP 94681; UF-H-12347; L. llanensis MZUSP 74347; Telmatobius thompsoni UF- H-39734. 52

APPENDIX II

Acronyms are: BMNH (British Museum of Natural History, London, UK); FCS (Facultad de Ciencias Sociales, Universidad Nacional del Centro de la Provincia de Buenos Aires); FMNH-P (Field Museum of Natural History, Geology Department, Chicago, Illinois); GHUNLPam (Facultad de Ciencias Exactas y Naturales, Universidad de La Pampa, Santa Rosa, La Pampa, Argentina); IANIGLA-PV (Instituto Argentino de Nivología, Glaciología y Ciencias Ambientales-Paleovertebrados); ICNU (Institutional collection number of material unreported); MACN (Museo de Ciencias Naturales de Buenos Aires “Bernardino Rivadavia”); MD-CH (Museo Municipal de Ciencias Naturales ‘Carlos Darwin’, Arroyo Chasicó collection, Punta Alta, Buenos Aires Province, Argentina); MLP (Museo de Ciencias Naturales de La Plata); MMH (Museo Municipal de Ciencias Naturales Vicente Di Martino, Monte Hermoso); MMP (Colección de Paleontología de Vertebrados Museo Municipal de Ciencias Naturales de Mar del Plata “Lorenzo Scaglia”, Mar del Plata); MHNM (Museo de Historia Natural de Montevidéo); MNHN (Muséum national d'Histoire naturelle, Paris, France); MZUSP (Museu de Zoologia da USP); PVL (Colección de Paleontología de Vertebrados del Instituto Miguel Lillo, San Miguel de Tucumán); PVSJ (Museo de Ciencias Naturales, Facultad de Ciencias Exatas, Físicas y Naturales, Universidad Nacional de San Juan); ZUFABC (Coleção de anfíbios, Universidade Federal do ABC, São Bernardo, Brazil).

Capítulo 2

A new head-to-head proposal of Ceratophryidae (Anura) phylogeny including fossil and living species

INTRODUCTION

Ceratophryidae is a well-supported clade (Frost et al. 2006; Fabrezi 2006; Fabrezi & Quinzio 2008; Pyron & Wiens 2011; Faivovich et al. 2014; Feng et al. 2017; Sabbag et al. 2018; Streicher et al. 2018) including large, robust, wide-mouthed frogs. The family (sensu Favovich et al. 2014; Frost 2019) embraces three genera: Ceratophrys (8 spp.), Chacophrys (1 sp.) and Lepidobatrachus (3 spp.). Ceratophrys is the most diverse genus presenting two species groups: the cornuta group, including C. calcarata Boulenger, 1890, C. cornuta (Linnaeus 1758), C. stolzmanni Steindachner 1882, and C. testudo Andersson, 1945; and the aurita group including C. aurita (Raddi 1823), C. cranwelli Barrio 1980, C. joazeirensis Mercadal de Barrio 1986, and C. ornata (Bell 1843; Lynch 1982; Faivovich et al. 2014).

The systematics of the family was exhaustively studied, accessed from phenotypic adult characters (Lynch 1982; Perí 1994; Wild 1997; Fabrezi 2006; Fabrezi & Quinzio 2008; and Vieira 2012), from adult and larval phenotypic characters (Lynch 1982; Wild 1997; Vieira 2012), from immunological data (Maxson & Ruibal 1988), and from molecular markers (Faivovich et al. 2014). The family is consistently considered monophyletic. However, the internal relationships are not completely congruent in the analysis. The three genera are considered monophyletic but the collateral relationships are not consensual: Ceratophrys is recovered as sister to the clade Chacophys plus Lepidobatrachus (Maxson & Ruibal 1988; Perí 1994; Fabrezi 2006; Faivovich et al. 2014; Frazão et al. 2015); Chacophys is recovered as sister to the clade Lepidobatrachus plus Ceratophrys (Lynch 1982; Wild 1997; Fabrezi & Quinzio 2008; Faivovich et al. 2014); and Lepidobatrachus is recovered as sister to the clade Chacophys plus Ceratophrys (Frost et al. 2006; Grant et al. 2006; Vieira 2012).

The fossil record of Ceratophryidae ranges from the late Miocene (11–5 million years bp) to late Holocene (11,000 ybp), with nearly 50 fossil specimens recognized (see Nicoli 54

2014, Barcelos et al. submitted). Six are determined to species level: Ceratophrys ameghinorum Fernícola 2001, C. ensenadensis Rusconi 1932, C. prisca Ameghino 1899, C. rusconii Agnolín 2005, Ceratophrys sagani Barcelos, Almeida-Silva, Santos and Verdade (submitted), and Lepidobatrachus australis Nicoli 2015. Seven are assigned as fossil representatives of extant species: Ceratophrys cf. C. ornata (Reig 1958; Vergnaud- Grazzini 1968; Perí 1993; Pardiñas 2001) and Ceratophrys cf. C. aurita (Günther 1859; Barcelos et al. submitted). More than 30 are not determined, assigned to Ceratophrys or Lepidobatrachus.

The high number of fossils in the family, the low number of specimens determined to species level, and some inaccurate classifications motivated many authors to review the classification of these fossils (e.g. Fernícola 2001; Nicoli 2015, 2016a, 2016b, 2017; Nicoli et al. 2016, 2017; Báez & Gómez 2017; Barcelos et al. submitted). Nevertheless, the fossil specimens of Ceratophryidae were never considered together with living species in a phylogenetic analysis. The goals of this paper are to (1) propose a phylogeny including extant species and extinct specimens taxonomically assigned as members of Ceratophryidae, (2) re-access the phylogeny of Ceratophryidae using morphological characters, (3) use a phylogenetic analysis to assert the position of fossil species within Ceratophryidae first proposed based on overall similarities.

MATERIAL AND METHODS

We restricted our analysis to skull characters, as most fossils are known only from preserved skulls. The characters were extracted from direct examination of specimens, and we also used Morphosource data (http://morphosource.org/), radiography, alizarin red cleared-and-stained specimens from museum collections, and µCT-scanning images (Appendix I).

We included in our sample at least one specimen from each Ceratophryidae living species (except for Ceratophrys testudo) and six fossil specimens, i.e., Ceratophrys ameghinorum, Ceratophrys cf. C. ornata Perí 1993 (included here as Ceratophrys sp. 86.VIII.1.4 to test Perí species hypothesis), C. rusconii, Ceratophrys sagani Barcelos et al., Ceratophrys cf. C. aurita BMNH 18895 (instead of C. cornuta of Günther 1859, see Barcelos et al. Cap. I) and Lepidobatrachus australis. The specimens were direct examined under a stereomicroscope Zeizz Stemi V11 at UFABC, and under the available 55 devices in visits to the collections of Museo de Ciencias Naturales de La Plata (MLP) and Museo Argentino de Ciencias Naturales “Bernardino Rivadavia” (MACN). All fossil specimens were directly examined, except for Lepidobatrachus australis coded using photos available in Tomassini et al. (2011) and Nicoli (2015); and Ceratophrys cf. C. aurita BMNH 18895, coded using photos ceded by British Museum. We supplemented our firsthand observations with descriptions and figures from the literature (Appendix I).

The µCT-scanning images were provided by the Laboratory of Computerized Tomography at Museu de Zoologia da USP (MZUSP), prepared in v tome x m microfocus µCT scanner Version 2.3.0.1032 (General Eletric Company, Wunstorf, DE). All specimens were scanned using a tungsten target, a background medium of air, no filter and were rendered as 16-bit TIFFs. The scan data were analyzed in the free viewer software MyVGL (Volume Graphics, Heidelberg, DE) and 3DSlicer software, version 4.10.1 (Fedorov et al. 2012). Photos of the fossils were taken with a LGK10 cell phone 13 MP, f/2.2, AF (LTE model). The specimens were measured directly with a Vernier caliper to the nearest of 0.01 mm and through ImageJ Fiji software (Schindelin et al. 2012) when measures were taken from µCT-scanned images and photos of specimens.

We used the Mesquite software, version 2.75 (Maddison and Maddison 2011) to construct the character data matrix, and used the Tree Analysis Using New Technology software (TNT version 1.5; Goloboff et al. 2008; Goloboff & Catalano 2016) to run the phylogenetic analysis. We performed traditional search for random addition sequences with a ‘random seed’ value of one, 10,000 replications, and 10 cladograms saved per replication. The branch swapping algorithm used was ‘Tree Bisection Reconnection’ (TBR), the trees were collapsed after the search. The characters were unordered (Caetano-Anollés et al. 2018). The Parsimony Jackknife (Farris et al. 1996), absolute frequencies were estimated using new technology (1,000 replicates) and Bremer support (Bremer 1994) was calculated using the TNT (10,000 replicates). We calculated the consistence index (CI; Kluge & Farris 1969) and retention index (RI; Farris 1989) in TNT software.

The phylogenetic analysis was performed with a data matrix including 70 morphological characters. Most characters were obtained from Lynch (1982), Perí (1994), Wild (1997), Fabrezi & Quinzio (2008), Nicoli (2015) and Báez & Goméz (2017), sometimes rephrased following Sereno (2007), but 19 are new characters. The 56 data matrix includes 17 terminals in the ingroup (11 extant species and 6 fossil specimens). The sister relationship of the Ceratophryidae to other Anura clades is not set, and would require a large sample of clades, that is beyond our scope. We consider Alsodes nodosus (Alsodidae) and Telmatobius thompsoni (Telmatobiidae) as outgroup, based on results presented by Lynch (1971), Zang et al. (2013), Faivovich et al. (2014), and Sabbag et al. (2018). We present the phylogenetic characters and coding for the taxa in Appendix II. A Nexus file of the matrix is available as Supplementary Data 1.

RESULTS

Phylogenetic analysis

We present a representative phylogenetic analysis for the internal relationships of Ceratophryidae based on morphological data, using skull-from only characters. We coded 70 characters for 19 terminals. The phylogenetic analysis performed resulted in six equally lengthy cladograms, summarized in the strict consensus (Fig 1), with a best score of 155 steps (CI 0.610; RI 0.753). The topology recovers Ceratophryidae (node 24) as monophyletic group, and all genera also as monophyletic (nodes 13, 22, and 33). The strict consensus tree is solved, except for the internal relationships of Lepidobatrachus. There are homoplasies, but most of them are informative as we can see from the tree indexes. Most clades are recovered under high Bremer values, indicating congruence among data.

List of synapomorphies

The synapomorphies that support Ceratophryidae (node 24, Bremer = 11) are: Skull, bones hyperossification and exostosis (1-1); Nasal and frontoparietal, contact (9- 1); Frontoparietal, degree of contact (11-1); Frontoparietal and squamosal, contact (15- 1); Parasphenoid, alae (27-1); Vomer, pars odontoides (31-1); Sphenethmoid, anterior portion, ossification degree (38-1); Sphenethmoid, planum antorbitale, mineralization (41-1); Columella (50-1); Tooth shape (52-1); Teeth condition (53-1); Premaxillae, pars palatina (56-1); Maxilla, pars palatina, pterygoid process (61-1); Fang-like, symphyseal ectopic ossifications (66-1); Choanae edges, ossification degree (69-1).

Chacophrys is recovered as the sister clade to Lepidobatrachus plus Ceratophrys (Bremer = 11). The autopomorphies that support Chacophrys (node 13) are: 57

Parasphenoid, overlapping the medial ramus of Pterygoid (28-0); Otoccipital artery channel, format (46-1). The synapomorphies that support Ceratophrys as sister clade to Lepidobatrachus (node 23) are (Bremer = 11): Squamosal, otic plate, extension (24-1); Vomer, vomerine teeth (30-1); Sphenethmoid, dorsal exposition (39-1); Otoccipital, suprapterygoid fenestra (45-0); Occipital artery channel, format (46-1); Maxilla, pars facialis, orbital flange participation (60-1); Maxillary and quadratojugal, fusion (63-1).

The synapomorphies supporting Lepidobatrachus (node 33) are (Bremer = 1): Frontoparietal, interorbital and postorbital width (14-0); Squamosal and maxilla, width (17- 2); Parasphenoid, cultriform process, extension (29- 2). The internal relationships of Lepidobatrachus are not solved, but the autapomorphies for L. asper (node 14) are: Vomer, pars odontoides, shape (32-1); Otocciptal, epiotic eminences, shape in dorsal view (49-2); Columella, pars interna plectri (51-2). The autapomorphies for L. laevis (node 16) are: Sphenethmoid, planum antorbitale, mineralization (41-0); Otoccipital, epiotic eminences, shape in dorsal view (49-3); Maxilla, pars facialis in the orbital region, shape (61-2). The autapomorphy for L. llanensis (node 17) is: Maxilla, pars facialis in the orbital region, shape (61-3). The autapomorphies for L. australis clade (node 15) are: Skull, covering of dermal roof bones by exostosis (2-0); Vomer, pars odontoides (31-0); Postemporal fenestrae (64-0).

The synapomorphies for Ceratophrys (node 22) are (Bremer = 11): Skull, bones hyperossification forming a single akinetic unit (4-1); Skull, bones hyperossification overlapping subtemporal fenestra (5-0); Orbits, position relative to skull (6-1); Frontoparietal, supraorbital flange, shape (13-1); Frontoparietal and squamosal, portion the contact occur (16-1); Squamosal, otic ramus, shape (23-3); Palatine, odontoids or ridges on ventral surface (35-1); Crista parotica, position in relation to dorsal edge of epiotic eminences (43-0).

We recovered Ceratophrys rusconii as the sister clade to all Ceratophrys species (node 21, Bremer = 3), which synapomorphies are: Squamosal and maxilla, lateral crest (18-1); Squamosal, supraorbital crest (20-1); Squamosal, otic plate, extension (24-2). The clade “Ceratophrys except C. rusconii” is split in two additional clades, corresponding grossly to the C. aurita and C. cornuta species group.

The synapomorphies that support C. cornuta species group (node 20; Bremer = 2) are: Squamosal, otic ramus shape (23-2); Squamosal, otic ramus, lateral plate (25-1); 58

Otoccipital, epiotic eminences, ornamentation in posterior view (48-2). The autapomorphies that support C. calcarata (node 4) are: Quadrate, position relative to occipital condyles (65-1); Fang-like symphyseal ectopic ossifications (66-0). The autapomorphies that support C. cornuta (node 5) are: Squamosal and maxilla, lateral crest (18-0); Palatine, odontoids or ridges on ventral surface (35-0); Premaxillae, alary process (54-1); Maxilla, pars facialis in the orbital region, shape (61-2). The autapomorphy that support C. ameghinorum (node 1) is: Squamosal and maxilla, width, evaluated as the width of the suture region of the two bones (WSM) in relation with the diameter of choanae (CD) (17-1).

Ceratophrys stolzmanni, considered a member of C. cornuta species group, was recovered as sister to the aurita species group in our analysis, the synapomorphies that support C. stolzmanni plus aurita species group (node 30, Bremer = 2) are: Vomer, vomerine teeth (30-0); Vomer, pars odontoides length (33-1).

The synapomorphies that support the C. aurita species group (node 29, Bremer = 2) are: Otoccipital, occipital condyles (47-1); Maxilla, pars palatina pterygoid process (62-2); Dorsal shield (67-1). The internal relationships of C. aurita species group are solved. There are two clades in the group: one including Ceratophrys cranwelli, C. ornata, and Ceratophrys sp. 86.VIII.1.4 (node 31); and other including C. joazeirensis, C. aurita, Ceratophrys sagani, and Ceratophrys cf. C aurita BMNH 18895 (node 28). The synapomorphies that support node 31 (Bremer = 1) are: Parasphenoid overlapping the medial ramus of Pterygoid (28-0); and Sphenethmoid, anterior portion, shape (40-1). The synapomorphies that support node 28 (Bremer = 1) are: Skull proportions, evaluated as the proportion between the skull height in relation to skull transverse diameter (3-2); Frontoparietals, processus posterior edge, shape (12-1); Squamosal, otic ramus, lateral plate (25-1).

Within node 31, the autapomorphies that support C. cranwelli are: Squamosal, lateral margin of orbit, oblique crest (19-1); Sphenethmoid, anterior portion, ossification degree (38-0); Crista parotica, overlapping degree by frontoparietal and squamosal, evaluated as the overlayed portion extension (OP) relative to the distal length (DL) (42- 2); Columella, pars interna plectri, ornamentation (51-0); Premaxillae, alary process, angle (54-1). The synapomorphies that support C. ornata plus Ceratophrys sp. 86.VIII.1.4 (node 32, Bremer = 3) are: Squamosal and maxilla, width (17-1); Squamosal, 59 supraorbital crest (20-0); Squamosal, otic plate, extension (24-1); Parasphenoid, cultriform process, extension (29-0); Otoccipital, epiotic eminences, ornamentation in posterior view (48-1). Ceratophrys sp. 86.VIII.1.4 is recovered as sister clade to C. ornata. The autapomorphy that supports the Ceratophrys sp. 86.VIII.1.4 (node 12) is: Otoccipital, epiotic eminences, shape in dorsal view (49-1).

Within node 28, the autapomorphies that supports C. joazeirensis (node 7) are: Squamosal, supraorbital crest (20-0); Squamosal, otic plate, extension (24-1); Vomer, pars odontoides, shape (32-1); Vomer, pars odontoides, angled (34-2). The synapomorphies that support the clade (node 27, Bremer = 4) C. aurita, Ceratophrys sagani and Ceratophrys cf. C. aurita BMNH 18895 are: Squamosal and maxilla, width (17-1); Squamosal, lateral margin of orbit, oblique crest (19-1); Otoccipital, epiotic eminences, shape in dorsal view (49-1); Premaxillae, alary process (54-1); Choanae, shape (70-1); there are no autapomorphies that support Ceratophrys sagani. The synapomorphy that support C. aurita plus Ceratophrys cf. C. aurita BMNH 18895 (node 26, Bremer = 1) is: Vomer, vomerine teeth (30-1). Autapomorphies of Ceratophrys cf. C. aurita BMNH 18895 (node 3) are: Vomer, pars odontoides, shape (32-1); Vomer, pars odontoides, length (33-0); Otoccipital, epiotic eminences, ornamentation in posterior view (48-2). There is no autapomorphy for C. aurita.

DISCUSSION

Ceratophryidae was recovered monophyletic as expected, and the topology recovered for internal relationships in the family (Figs. 1, 2) is congruent to those of Faivovich et al., (2014), Lynch (1982), Wild (1997), and Fabrezi & Quinzio (2008), the first, based on molecular markers; the other, proposals based on morphological datasets. The points of incongruence among the different analyses and the phylogenetic positioning of fossil species are further discussed below.

Faivovich et al. (2014) proposed the most recent and complete phylogenetic analysis focusing on Ceratophryidae and recovered Chacophrys as sister clade to Lepidobatrachus when using direct optimization, and Chacophrys as sister clade to Ceratophrys plus Lepidobatrachus when using static parsimony analysis. Our phylogeny corroborates the collateral relationships obtained from the static parsimony analysis. The Ceratophryidae clade is supported in our analysis with 100% Jackknife frequency. The 60 collateral relationship of Lepidobatrachus and Ceratophrys is supported with 100% Jackknife frequency.

Taxonomic accounts

Chacophrys Chacophrys is a monotypic genus, the status of C. pierottii was doubted by Lynch (1982) but corroborated by Maxson & Ruibal (1988) from immunological data and from the description of its tadpole and ontogeny (Wild 1999). We present two non-exclusive synapomorphies that supports Chacophrys as a distinct genus.

Ceratophrys Ceratophrys is recovered as a clade supported with 99% jackknife frequency. This clade includes, beyond the living species, three fossil species and two fossil specimens. We recovered partially the species groups proposed by Lynch (1982), and we consider the cornuta species group as: C. ameghinorum, C. calcarata, and C. cornuta; and consider the aurita species group as: C. aurita, Ceratophrys cf. C aurita BMNH 18895, C. cranwelli, C. joazeirensis, C. ornata, and Ceratophrys sp. 86.VIII.1.4. Our topology is mostly congruent to the ones presented by Faivovich et al. (2014) and Lynch (1982), except for C. stolzmanni recovered as sister to aurita species group. The species appears as sister to C. cornuta plus C. calcarata in the works of Faivovich et al (2014) and Lynch (1982). Our topology is also consistent to that recovered by Perí (1994) on internal relationships of Ceratophrys, with the exception of C. calcarata, recovered as close related to C. aurita species group in the aforementioned work. Mercadal (1981) described and included C. joazeirensis in a non-quantitative phylogenetic proposal. C. joazeirensis was recovered as sister clade to C. ornata. In this issue we recovered a different topology, C. stolzmanni as close related to C. aurita plus C. cranwelli, C. joazeirensis and C. ornata.

Ceratophrys rusconii is a fossil species described by Agnolín (2005). The author considered the species as closely related to C. ornata and C. cranwelli. Our results (Fig. 1) indicate C. rusconii as the sister clade to all Ceratophrys species. There are no autopomorphies for this clade. We recognize the autapomorphies presented by Agnolín and present other two possible autapomorphies: the anteriormost portion of sphenethmoid reduced, and a short and anterior angled palatine. This palatine condition was not commented by Agnolín (2005), and was considered, until now, an exclusive condition of Lepidobatrachus. 61

Ceratophrys ameghinorum was recovered as part of C. cornuta species group, supported by 84% Jackknife frequency. The specimen was first described by Rovereto (1914) and assigned to C. prisca. Rovereto (1914) classified four additional specimens as: C. prisca var. subcornuta, C. prisca var. intermedia, C. prisca var. gigantea, and C. prisca plesiotype. The specimens classified by Rovereto (1914) were re-accessed by Fernícola (2001), and described as a new fossil species, C. ameghinorum in honor to Carlos and Florentino Ameghino. We acknowledge all the autopomorphies recognized in Fernícola (2001), corroborate the extinct species status and propose the addition of one character recovered in our analysis in its diagnose. Our observation indicates the species present the otic plate of squamosal laceolate as C. cornuta and C. calcarata, but less acute. Additionally, C. ameghinorum present the otic plate angled dorsally, and C. cornuta and C. calcarata presents this portion directed laterally. Due the history of species attribution concerning C. ameghinorum specimens mentioned above, on our exploratory analysis, we coded only the holotype. However, we are skeptical about the positioning of C. ameghinorum in cornuta species group, due the presence of some missing data in this analysis that could be important if further coded (e.g. fragment dorsal shield, premaxilla, mandible, vomer pars odontoides preserved in MACN 19731). Therefore, the position of this fossil species could possibly change with the inclusion of more specimens of C. ameghinorum in the analysis.

Ceratophrys cf. C. aurita BMNH 18895 was described in Günther (1859). It was re-accessed by Faivovich et al. (2014) and Nicoli (2016) presenting a new taxonomic proposal, assigning this fossil as close related to [C. aurita-C. joazeirensis] clade, and Barcelos (submitted) proposed this fossil is a C. aurita fossil specimen. This fossil is first included in a phylogenetic analysis in this issue. As hypothesized in Barcelos et al. (submitted), Ceratophrys cf. C. aurita BMNH 18895 is recovered here as sister clade of C. aurita. Ceratophrys sagani was first mentioned in Barcelos et al. (2018) and was described in Barcelos et al. (submitted) and hypothesized as close related to [C. aurita – C. joazeirensis] clade. Herein, we first include this species in a phylogenetic analysis. Ceratophrys sagani was recovered as sister clade to C. aurita and Ceratophrys cf. C. aurita. Mainly, due to the presence of an oblique crest on the lateral margin of orbit, and the Choanae (Fig. 2), oblong shaped, synapomorphies of the clade including Ceratophrys 62 sagani, C. aurita and Ceratophrys cf. C. aurita. We acknowledge all the autopomorphies recognized in Barcelos et al. (submitted) and corroborate the extinct species status.

Ceratophrys sp. MLP 86.VIII.1.4 was recovered as sister clade to C. ornata within the aurita species group. Ceratophrys sp. 86.VIII.1.4 was described in Perí (1993) together with two fossil specimens of Ceratophrys, assigned to the genus level. This fossil was never re-accessed and included in a phylogenetic analysis. Our topology recovers this fossil as sister clade to C. ornata. We maintain the Perí (1993) hypothesis that this fossil specimen is a representative of C. ornata extant species. Ceratophrys ornata was described by Bell (1843), and C. cranwelli was described by Barrio (1980). Perí (1993) points that she could not find any difference between the two species, except apophyses of pars interna plectra of the columella, and in Perí (1994) recovered C. cranwelli as sister clade to C. aurita plus C. ornata; Wild (1997) recovered C. cranwelli as sister clade to C. aurita plus C. calcarata and C. cornuta. Faivovich et al. (2014) recovered C. cranwelli as sister clade to C. ornata.

Note on Ceratophrys testudo

Andersson (1945) described the species. Lynch (1982) synonymized it to C. cornuta. Mercadal (1988) resurrected C. testudo. Perí (1993b) proposed this species is a C. cornuta juvenile specimen. We only had access to an x-ray and lateral view images of the holotype housed in the Naturhistoriska Riksmuseet Stockholm, Sweden. Therefore, we considered the codification of characters from the available images poor, and excluded the species of the analysis.

Note on Ceratophrys ensenadensis Rusconi (1932) described briefly the fossil species, assigning four fossil specimens as C. ensenadensis, three from Frederico Hennig collection (i.e. n° 480, n° 481 and another specimen without collection number), and one specimen without collection number from Museo de Ciencias Naturales de La Plata, Argentina identified as “Ceratophrys ? ornata” at that time. The holotype was recognized as species inquirenda by Nicoli (2014). We search for this specimen in Museo de La Plata without success and do not have record of this specimen in the collection.

Note on Ceratophrys prisca Ceratophrys prisca was described by Ameghino (1899) from Monte Hermoso, 63

Buenos Aires, Argentina. Fernícola (2001) revised the specimens of Rovereto and gave species inquirenda status to the holotype specimen of Ameghino (1899), that is currently lost (Fernícola 2001; Nicoli 2014).

Note on the Ceratophrys prisca var. subcornuta MACN 14319, 14323, 14325 and Ceratophrys sp. MACN 14322 Rovereto (1914) described and illustrated the specimen MACN 14319. Fernícola (2001) revised the Rovereto’s fossil specimens and included new fossil specimens in the sample (e.g. MACN 14325) assigning them all to C. ameghinorum. We could verify through firsthand observation that the specimen MACN 14319 is clearly deformed, possibly, product of a taphonomic artefact. This was not mentioned in Rovereto (1914) and neither in Fernícola (2001). The morphometric measurements, and some bones shape observations obtained from this specimen should not be used for further comparisons. MACN 14323 was first mentioned in Mercadal de Barrio & Barrio (2002) and assigned as Ceratophrys prisca var. subcornuta, MACN 14319 and MACN 14325 received the same classification. The specimen Ceratophrys sp. MACN 14323 is highly fragmented skull that was grossly restored and was never described neither illustrated. We do not agree with the Mercadal de Barrio & Barrio (2002) classification of this fossil, there are clear differences in the skull bones differing MACN 14323 from all other specimens assigned to the same species (e.g. the medial portion of nasals without exostosis). The fossil specimens MACN 14322 (maxilla, squamosal and quadrate fragments) and MACN 14325 (other skull bones) are crassly restored together with epoxy-like material filling in the whole ventral portion (Appendix III). Fernícola (2001) do not mentioned the specimen MACN 14322, it was only studied in Mercadal de Barrio & Barrio (2002). The problem concerning these specimens is that MB&B proposed MACN 14325 fossil specimen present 4n (2n=4x) polyploidy and considered it as a Ceratophrys prisca var. subcornuta specimen, and MACN 14322 is a diploid, assigned as Ceratophrys sp. Our first hand observation indicate the specimens present two collection number MACN 14322 in red marker (4444 in black marker) and MACN 14325 in red marker (4799 in black marker). This could indicate the fossils were recorded in the collection in different occasions. We suppose there are three possible answer for this urgent issue: (1) the crass restoration were made after Fernícola’s times and was mistaken; (2) The 64

polyploidy proposition of MB&B (2002) could have mistaken the specimens; (3) There are two fossil specimens with the same collection number. Several authors recognized as invalid the characters that supported the C. prisca species hypothesis (Reig 1958; Perí 1993; Fernícola 1994), rejected all variations proposed by Rovereto and recommended that the species should be reviewed (Gasparini & Báez 1975; Báez & Gasparini 1977; Fernícola 2001). Mercadal de Barrio & Barrio (2002) did not follow any recommendation and assigned the specimens MACN 14319, MACN 14323, MACN 14324, and MACN 14325 as C. prisca. Due the various problems mentioned, we propose these fossils as Ceratophrys sp. We reinforce the necessity of proper restoration of the fossils and suggest the specimens should be µCT-scanned to allow the study of ventral view of MACN 14325, understand the fossil fragments glued together and describe properly the fossils.

Lepidobatrachus Lepidobatrachus is recovered as monophyletic. However, with the characters we choose, the internal relationships in Lepidobatrachus was not resolute. The history of taxonomic classification of Lepidobatrachus species was marked by some important confusions (Budgett 1899; Boulenger 1919; Nieden 1923; Vellard 1948; Cei 1958; Reig & Cei 1963). Barrio (1968) revised the genus and the propositions made on this work remained. Perí (1994) and Vieira (2012) proposed Lepidobatrachus laevis as sister clade to L. asper plus L. llanensis. Faivovich et al. (2014) proposed Lepidobatrachus asper as sister clade to L. laevis plus L. llanensis. We propose eight synapomorphies for the clade.

Lepidobatrachus australis was first described in Tomassini et al. (2011) as L. laevis and was re-classified by Nicoli (2015) rising it to the extinct species status. It is included in a phylogenetic analysis for the first time in this issue. Lepidobatrachus australis is recovered a Lepidobatrachus species, but with the characters applied in our analysis we could not propose a collateral relationship in the genus. We acknowledge all the autopormophies recognized in Nicoli (2015), corroborate the extinct species status and propose the addition of the three new characters recovered in our analysis.

Taxonomic misidentification of fossils of Ceratophryidae The misidentification of fossils is a problematic and systemic issue in Paleontology and Systematics of Anura. Some issues helped to straighten out the Paleobatracology. Focusing on Ceratophryidae, Agnolín (2012), recovered Beelzebufo ampinga as a 65

Calyptocephalellidae fossil species, instead of a Ceratophryidae as proposed by Evans et al. (2008; 2014). Nicoli et al. (2016) revised Báez & Perí (1990) work and changed the Wawelia gerholdi (known Ceratophryidae) fossil species to the Calyptocephalella gerholdi (Calyptocephalellidae) species status. Báez & Gómez (2017), contributed to the phylogenetic rectification of Baurubatrachus pricei, known as one of the most ancient Ceratophryidae by Báez & Perí (1989), and in aforementioned work was hypothesized as sister clade to fossil representative of Calyptocephalellidae family.

Fernícola (2001) taxonomically assigned Ceratophrys species to the C. ameghinorum species. Fernícola & Vizcaíno (2006) mentioned the presence of a Ceratophrys specimen in the Santa Cruz Province, Fernícola & Albino (2012) revised the specimen MPM-PV 3507, an incomplete right frontoparietal, and assigned it as a Calyptocephalella sp. Nicoli (2014; 2015; 2016; 2017) and Nicoli et al. (2017) re-accessed fossil specimens and revised them taxonomically.

Günther (1858) described various extant species, including C. cornuta and three other Ceratophys species. The author commented that C. cornuta presents a bony dorsal shield. Nowadays is a consensus that C. cornuta do not present dorsal shield. On XVIII and XIX Centuries there was a taxonomic confusion, C. aurita was attributed to C. cornuta (e.g. Seba 1734; Kloetzke 1816; Wied-Neuwied 1822; and Spix 1824). One year later Günther (1859) described two fossil specimens of Ceratophrys from Lagoa Santa locality, Minas Gerais - Brazil. Therefore, these clues could indicate that Günther (1859) described the fossils of C. cornuta BMNH 18895 referring to C. aurita. Báez & Gasparini (1977) referred to C. cornuta fossil specimen from Lagoa Santa locality, of Günther (1859) as C. aurita, but without explain this change. Faivovich et al. (2014) and Nicoli (2016) commented on the taxonomic rectification of Ceratophrys cornuta BMNH 18895 (Günther 1859), addressing this fossil as close related to [C. aurita-C. joazeirensis] clade and Barcelos et al. (submitted) proposed this fossil is a fossil representative of C. aurita. Here, we corroborated this hypothesis.

Pardiñas (2001) assigned the fossil specimens MMH 85-3-8 (two frontoparietal fragments); 85-2-11 (one frontoparietal fragment); MLP s/n (two presacral vertebrae); MMH 88-2-5 (two fragmented humerus); MMH 90-2-1 (distal fragment of a humerus) to Ceratophrys cf. C. ornata. The author does not describe and presented a diagnosis supporting the species hypothesis and the fossils fragments are not sufficiently informative. 66

There are not any autopomorphic characteristics described in the literature for C. ornata representing the frontoparietal and humerus bones. Therefore, we request a judicious look to hypothesize that these fossils are representative of an extant species. We consider these fossils as species inquirenda. We tried to get photos of the specimens on Museo Municipal de Monte Hermoso, Argentina but they are currently loaned.

Vieira (2012) propose a phylogenetic analysis including C. ameghinorum and C. rusconii fossils. However, based on polyploidy data of Mercadal de Barrio & Barrio (2002), he made an unjustified taxonomic confusion, considering Ceratophrys cf. C. aurita BMNH 18895, Ceratophrys specimens MLP 86.viii.1.4, MLP 86.viii.1.5, MLP 86.viii.1.6 Perí 1993, and other fossil specimens, classified only at the genus level, as C. rusconii specimens. Additionally, the author considers the C. ameghinorum fossil specimen MACN 14325 as a C. rusconii. Our results indicate the taxonomic suggestions made in Vieira (2012) are incorrect. We corroborate, through specimens direct examination and in our phylogenetic analysis, that the specimens examined by the author represent different species, not close related, and not attributed to C. rusconii.

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Figure legends Figure 1: Strict consensus of 6 most parsimonious trees from the analysis using 19 terminals and a matrix of 70 characters from skull morpholgy. Bremer support values are indicated below branches; Values above nodes are parsimony jackknife frequencies. Nodes lacking values have < 50% jackknife frequencies.

Figure 2: Illustration of right portion of skull of Ceratophrys sagani representing the bones elements studied. a) dorsal view; b) ventral view; c) frontal view; d) posterior view; e) lateral view. Abbreviations: c, choana; co, occipital condyles; cp, crista parotica; ee, epiotic eminence fp, frontoparietal; lc, lateral crest; lp, lateral plate; m, maxilla; n, nasal; nf, nerve foramina I-XI; oc, oblique crest; op, otic plate; otc, otoccipital; ps, parasphenoid; pl, palatine; pt, pterygoid; ptf, postemporal fenestra; q, quadrate; qj, quadratejugal; sbf, subtemporal fenestra; sf, supratemporal fenestra; soc, supraorbital crest; sph, sphenethmoid; sq, squamosal; v, vomer (modified from Barcelos et al. cap I).

Figure 1

Figure 2

APPENDIX I

Specimens Examined. Acronyms are: CM-HERPS (Carnegie Museum of Natural History – Herpetology) Cas-herp (California Academy of Sciences – Herpetology); DNPM-MP (Departamento de Produção Mineral – Museu de Paleontologia); MACN (Museo de Ciencias Naturales de Buenos Aires “Bernardino Rivadavia”); MCZ:Herp (Museum of Comparative Zoology, Harvard University); MLF (Miguel Lillo Foundation); MLP (Museo de Ciencias Naturales de La Plata); MZUSP (Museu de Zoologia da Universidade de São Paulo); UF-H (Florida Museum of Natural History – Herpetology); USNM (National Museum of Natural History, Smithsonian Institution)

Dry skeletons: Ceratophrys aurita MZUSP 22976* MZUSP 61114(2); DNPM-MP 1- AR; KU 98129; KU 223818; Ceratophrys cornuta KU 222113; Ceratophrys cranwelli KU 211317; FML 5471.

Cleaned and stained: Chacophrys pierottii MZUSP 16274

Fossils: Ceratophrys† MLP 86-viii-1-4, 86-viii-1-5, 86-viii-1-6; C. ameghinorum† MACN 14317 (paratype), 14318, 19731 (paratype); C. prisca† MACN 14323, C. rusconii† MACN 19744; Ceratophrys sp.† MACN 14328.

Specimens examined through µCTscanned images: Alsodes nodosus CM-HERPS-68395; Ceratophrys aurita MZUSP 31367; and Cas-herp-84998; C. calcarata MZUSP 99657; C. cornuta MZUSP 70548; uf:herp:63162; MCZ: Herp: A-17499; C. cranwelli MZUSP 99320; MCZ: Herp: A-35360; C. joazeirensis MZUSP 142284; C. ornata MZUSP 70791; C. stolzmanni USNM 160970; Chacophrys pierottii KU191932, MZUSP 99329; Lepidobatrachus asper MZUSP 94595; L. laevis MZUSP 94681; UF-H-12347; L. llanensis MZUSP 74347; Telmatobius thompsoni UF-H-39734.

Specimens examined through photographs and literature: Ceratophrys ameghinorum Fernícola (2001); Ceratophrys aurita Gayer (1984); Ceratophrys joazeirensis Vieira (2006); Ceratophrys cornuta Wild (1997); Ceratophrys rusconii Agnolín (2005); Lepidobatrachus Reig & Cei (1963); Reig (1960); Lepidobatrachus Barrio (1968); Reig & Limeses (1963); Lepidobatrachus australis Tomassini et al. (2011); Nicoli (2015). 80

APPENDIX II

Characters list

The character list is composed of 28 original characters, 23 rephrased characters, 19 new characters. Some characters are commented to favor understanding. Symbols: *, original unmodified characters; #, rephrased characters; §, new characters.

#1. Skull, bones hyperossification and exostosis: (0) absent; (1) present. Rephrased from character 51 (Perí 1994)

#2. Skull, covering of dermal roof bones by exostosis: (0) Partially covered; (1) Completely covered. Rephrased from character 51 (Perí1994)

§3. Skull proportions, evaluated as the proportion between the maximum skull height (H- length of the quadrate level until frontoparietal roof level) in relation to maximum skull width (TD- length of lateral extremity of quadratojugal to quadratojugal): (?) not applicable; (0) H near 1/4 TD; (1) H 1/4 < H < 1/2 TD; (2) H near 1/2 TD. This character was inspired in Reig & Cei (1963) and Trueb (1974) studies. The character states were set based on variation present in Ceratophryidae.

#4. Skull, bones hyperossification: (0) Forming single akinetic unit (1) not forming single akinetic unit. Rephrased based on character 5 (Lynch 1982), and character 1 (Wild 1997).

#5. Skull, bones hyperossification overlapping subtemporal fenestra: (0) absent; (1) present. Rephrased from character 51 (Perí 1994).

*6. Orbits, position relative to skull: (0) located at the mid-length of the skull; (1) located posterior to the mid-length of the skull.

Character proposed by Reig (1961) and Nicoli (2015). It was well noted by the two authors, but this character was never added to a phylogenetic analysis.

*7. Nasals, relation to one another: (0) well separated; (1) slightly separated or in minimal contact; (2) in contact throughout most of their medial margins, or completely fused. proposed as character 11 by Perí (1994). We modified only state 1 to include another probable variable included in the outgroup of Ceratophryidae.

*8. Nasals, maxillary process, shape: (0) slender, sharp process; (1) robust, hourglass- shaped; (2) robust bar of uniform width. 81

Character proposed by Nicoli (2015)

#9. Nasal and frontoparietal, contact: (0) absent; (1) present.

*10. Nasal-frontoparietal articulation: (0) perpendicular to midline; (1) oblique to midline.

Character proposed by Nicoli (2015). It was well observed by the author, but in some specimens, the anterior edge of frontoparietal is rounded and it is difficult to define which character states best fits in this universe. We noted it in C. stolzmanni specimen and consider the rounded edges of frontoparietals perpendicular to midline.

*11. Frontoparietals, degree of contact: (0) separated; (1) in medial contact. Character 64 proposed by Fabrezi & Quinzio (2008).

*12. Frontoparietal, processus posterior edge, shape: (0) not deflected dorsally; (1) deflected dorsally. Character 3 proposed by Wild (1997).

*13. Frontoparietal, supraorbital flange, shape: (0) not angled dorsally; (1) angled dorsally. Character 4 proposed by Wild (1997).

§14. Frontoparietal, interorbital and postorbital width and length, evaluated as the proportion between the maximum width and length of interorbital portion and postorbital portion: (0) Postorbitary portion with near the same width and extension of the interorbital ones; (1) Postorbitary portion much wider and longer than the interorbital ones. Character obtained by the description of Reig & Cei (1963) and was never included in a phylogenetic analysis and was first tested in this issue.

A) Ceratophrys sagani in dorsal view; B) Lepidobatrachus llanensis in dorsal view. 82

*15. Frontoparietal and squamosal, contact: (0) absent; (1) present. Character 40 proposed by Perí (1994).

*16. Frontoparietal and Squamosal, portion the contact occur: (0) contacts above crista parotica; (1) contacts anteriorly to crista parotica. Character 40 proposed by Perí (1994).

§17. Squamosal and maxilla, width, evaluated as the width of the suture region of the two bones (WSM) in relation with the diameter of choana (CD): (0) narrow (WSM near the same size CD); (1) wide (WSM near two times the size of CD); (2) much wide (WSM > than two times the size of CD).

A) C. joazeirensis (0); B) Ceratophrys sagani (1); C) Lepidobatrachus llanensis (2) 83

*18. Squamosal and maxilla, lateral crest: (0) absent; (1) present.

Character 17 proposed by Lynch (1982); Also present in Wild (1997), as character 14. It was treated as a lateral carina in Lynch and Wild characters and here we noted there are more variation present in dermal roof bones of Ceratophryidae and created character 19 and 20 to access this unexplored variation.

A) C. cornuta; B) Ceratophrys sagani. 84

§19. Squamosal, lateral margin of orbit, oblique crest: (0) absent; (1) present.

§20. Squamosal, supraorbital crest: (0) absent; (1) present.

*21. Squamosal, zygomatic ramus, bone contact: (0) absent; (1) with the maxillar; (2) with maxillary and nasal.

Character 12 proposed by Perí (1994).

#22. Squamosal, otic ramus: (0) short, poorly developed; (1) reaching the prootic; (2) expansive and overlapping the prootic.

Rephrased from character 66 (Fabrezi & Quinzio 2008).

#23. Squamosal, otic ramus, shape: (0) in cuneiform format; (1) smooth sub-quadrangular shape; (2) lanceolate format; (3) sub-quadrangular format. 85

Rephrased from character 41 (Perí 1994). This character was rephrased from character 41 from Perí analysis mainly because we think the phylogenetic signal of this character could be improved if turn two variables independent.

§24. Squamosal, otic plate, extension: (0) anterior to the occipital condyles; (1) same level of the occipital condyles; (2) beyond the occipital condyles.

Rephrased from character 41 (Perí 1994).

#25. Squamosal, otic ramus, lateral plate: (0) absent; (1) present.

Rephrased from character 10 (Lynch 1982).

#26. Supratemporal fenestra, visibility: (0) not visible, covered by squamosal and frontoparietal; (1) visible, not covered by squamosal and frontoparietal.

Rephrased character from Nicoli (2015).

We restructured this character to highlight the focus variable in this character, that is the Supratemporal fenestrae visibility.

#27. Parasphenoid, alae: (0) not in contact with the medial ramus of Pterygoid; (1) in contact with the medial ramus of Pterygoid.

Rephrased from character 47 (Perí 1994). In Perí character 47 are two variables, not mutually dependent, degree of contact and overlapping degree. We split in two characters that best fit the universe of this analysis.

#28. Parasphenoid overlapping the medial ramus of Pterygoid: (0) absent; (1) present.

Rephrased from character 47 (Perí 1994).

*29. Parasphenoid, cultriform process, extension: (0) not reaching planum antorbitale; (1) reaching planum antorbitale; (2) beyond the planum antorbitale.

Character 13 proposed by Perí (1994).

#30. Vomer, vomerine teeth: (0) present; (1) absent.

Rephrased from character 19 (Lynch 1982).

§31. Vomer, pars odontoides: (0) absent; (1) present. 86

The pars odontoides, is a bone projection related in Gayer (1984). It is an unexplored variation, first tested in this issue. Our analysis indicates this bone projection is well conserved among specimens of the same species and a good discriminant between species.

A) Telmatobius thompsoni; B) C. aurita; C) C. joazeirensis.

§32. Vomer, pars odontoides: shape: (0) laminar, bladed; (1) threadlike.

§33. Vomer, pars odontoides, length: (0) short; (1) long.

§34. Vomer, pars odontoides, angled: (0) laterally; (1) anteriorly; (2) posteriorly.

*35. Palatine, odontoids or ridges on ventral surface: (0) absent; (1) present.

Character 28 proposed by Báez & Goméz (2017).

*36. Palatine: (0) absent; (1) present as a discrete element. 87

Character 27 proposed by Báez & Goméz (2017).

#37. Palatine, angle in relation to skull longitudinal axis: (0) perpendicular; (1) anterolateral.

Rephrased from character 9 (Wild 1997). This variation was well observed and tested by Wild, however, C. rusconii present a short and anterolateral angled Palatine, something only recorded in Lepidobatrachus species.

#38. Sphenethmoid, anterior portion, ossification degree: (0) cartilaginous;

(1) ossified.

Rephrased from character 4 (Perí 1994). We paraphrased Perí character (i.e. Sphenethmoid ossification extending beyond the planum antorbitale) to emphasize the variation we would like to explore, that is the anterior portion of sphenethmoid that contacts Vomer, together with the anteriormost edge that could present an ornamentation explored in character 40.

#39. Sphenethmoid, dorsal exposition: (0) visible; (1) hidden by nasals and frontoparietals.

Rephrased from character 63 (Fabrezi & Quinzio 2008).

§40. Sphenethmoid, anterior portion, shape: (0) reduced; (1) narrow and flat;

(2) narrow and rounded; (3) wide and rounded; (4) wide and flat.

This is a variation never explored and first tested here. We noted it could be a useful to understand the variation present in the fossil record of Ceratophrys and indicate a more assertive phylogenetic positioning of fossil specimens.

A) C. rusconii; B) C. joazeirensis; C) C. cornuta; D) Ceratophrys sagani; E) C. aurita. 88

*41. Sphenethmoid, planum antorbitale, mineralization: (0) mostly cartilaginous; (1) well ossified, at least one half of planum.

Character 33 proposed by Báez & Gómez (2017).

§42. Crista parotica, overlapping degree by frontoparietal and squamosal, evaluated as the overlayed portion extension (OP) relative to the distal length (DL):

(0) not; (1) OP < 1/2 < DL overlayed by medial portion of otic plate of squamosal;

(2) OP near 3/4 DL overlayed by medial portion of otic plate of squamosal;

(3) completely overlayed by medial portion of otic plate of squamosal.

This character explores the variation present in C. cranwelli, that present a squamosal projection overlapping almost completely the crista parotica.

A) T. thompsoni; B) C. aurita; C) C. cranwelli; D) L. llanensis. 89

*43. Crista parotica, position in relation to dorsal edge of epiotic eminences:

(0) ventral; (1) same level.

Character 45 proposed by Perí (1994).

*44. Crista parotica, degree of ossification: (0) mostly cartilaginous; (1) ossified at least one half.

Character 38 proposed by Báez & Gómez (2017).

*45. Otoccipital, suprapterygoid fenestra: (0) present; (1) absent.

Character 15 proposed by Perí (1994).

*46. Occipital artery channel, format: (0) groove; (1) duct.

Character 17 proposed by Perí (1994).

#47. Otoccipital, occipital condyles: (0) without continuum attachment articulation surface; (1) with continuum attachment articulation surface.

Rephrased from character 20 (Perí 1994).

§48. Otoccipital, epiotic eminences, ornamentation in posterior view: (0) flat; 90

(1) flat with one protuberance dorsoposteriorly; (2) with dorsolateral protuberances;

(3) with concavity.

The variation in the epiotic eminences was never explored in a phylogenetic analysis, and neither explored in osteological description of Ceratophryidae species. Inspired on the observation of Vieira (2006) in C. aurita and C. joazeirensis species, we created the character to test the intraspecific variation.

§49. Otoccipital, epiotic eminences, shape in dorsal view: (0) narrow; (1) elliptical; (2) right angle triangle-shape; (3) horn-shape.

*50. Columella: (0) absent; (1) present.

Character 42 proposed by Báez & Goméz (2017).

§51. Columella, pars interna plectri, ornamentation: (0) concave, globous; (1) with anterior and posterior plectral apophysis; (2) with crests.

A) L. llanensis; B) C. cornuta; C) Ceratophrys sagani in C.1) ventral view and C.2) dorsal view

Perí (1993) explored this variation in C. ornata fossil specimen, and inspired on this work we created this character to test the variation present on this bone. 91

*52. Tooth shape: (0) bicuspid; (1) monocuspid.

Character 68 proposed by Fabrezi & Quinzio (2008).

*53. Teeth condition: (0) pedicellate; (1) non pedicellate.

Character 1 proposed by Lynch (1982); also present in Fabrezi & Quinzio (2008), as character 69.

*54. Premaxillae, alary process, angulation: (0) parallel; (1) divergent, in frontal view.

Character 71 proposed by Fabrezi & Quinzio (2008).

§55. Premaxilla, alary process, length in relation to premaxillae entire height:

(0) short; (1) long.

A) C. joazeirensis; B) L. llanensis.

#56. Premaxillae, pars palatina: (0) present; (1) absent.

Rephrased from character 70 (Fabrezi & Quinzio 2008). Perí (1994) noted during ontogeny, an interesting morphological novelty in C. cranwelli, the pars palatina of maxillae extends dorsally investing pars facialis. Nicoli (2017) noted this same variation in C. cranwelli and Chacophrys pierottii juvenile specimens. We noted on µCT-scanned 92 adult specimens, internally in the pars palatina region, a vacuity in the portion outgroup species present a massive bone. Therefore, we corroborate the hypothesis of Perí and Nicoli, but opted to maintain the character of Fabrezi & Quinzio and expect this variation could be thoroughly studied and understood.

#57. Premaxillae, pars palatina, configuration: (0) complete; (1) reduced to the palatine processus only.

Rephrased from character 70 (Fabrezi & Quinzio 2008). This character was split in two (56 and 57) based on the presence of two independent variables that could be best explored using two independent characters.

§58. Premaxilla, alary process, slopping: (0) do not slope dorso-posteriorly;

(1) slope dorso-posteriorly.

*59. Maxilla, transverse pillars of the pars dentalis: (0) thin; (1) long.

Character proposed by Reig (1961) and tested in Perí (1994), as character 21.

*60. Maxillae, pars facialis, orbital flange participation: (0) participates in the formation of the orbit; (1) do not participates in the formation of the orbit.

Character 1 proposed by Perí (1994).

*61. Maxilla, pars facialis in the orbital region, shape: (0) decreases gradually in height in the orbital region; (1) decreases abruptly in height in the orbital region; (2) high along the orbital region but decreases posteriorly; (3) high throughout most of its length.

Character 49 proposed by Báez & Gómez (2017).

#62. Maxilla, pars palatina, pterygoid process: (0) not differentiated; (1) developed.

Rephrased from character 46 (Perí 1994).

#63. Maxillar and quadratojugal, fusion: (0) not fused; (1) fused

Rephrased from character 52 (Perí 1994).

#64. Postemporal fenestrae: (0) absent; (1) present

Rephrased from character 39 (Perí 1994). 93

*65. Quadrate, position relative to occipital condyles: (0) anterior; (1) same level; (2) adjacent posterior; (3) Well posterior.

Character 3 proposed by Perí (1994).

*66. Fang-like, symphyseal ectopic ossifications: (0) absent; (1) present.

Character 59 proposed by Báez & Gómez (2017).

#67. Dorsal shield: (0) absent; (1) present.

Rephrased from character 30 (Perí 1994).

It was split in two characters (67 and 68) to evaluate independently two different variables, i.e. presence and composition.

#68. Dorsal shield, composition, in relation to vertebral column: (0) formed by one plate in anteroposterior disposition; (1) two plates in anteroposterior disposition; (2) more than three plates in transversal disposition.

Rephrased from character 30 (Perí 1994).

§69. Choanae edges, ossification degree: (0) cartilaginous; (1) ossified.

§70. Choanae, shape, evaluated as the maximum length (ML) in relation with the maximum height (MH) of choana: (0) rounded; (1) oblong.

A) C. joazeirensis; B) Ceratophrys sagani 94

Data matrix

Species/ Characters 1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 22 23 24 25 26 27 28 29 30 31 32 33 34 35 Alsodes nodosus 0 ? 1 0 ? 0 0 0 0 ? 0 0 0 ? 0 ? 0 0 0 0 0 0 0 0 0 ? 0 ? 1 0 0 ? ? ? 0 Ceratophrys ameghinorum 1 1 1 1 0 1 2 2 1 1 1 0 1 1 1 1 1 1 0 1 2 2 2 2 1 1 1 0 ? 1 1 0 ? ? 1 Ceratophrys aurita 1 1 2 1 0 1 2 2 1 1 1 1 1 1 1 1 1 1 1 1 2 2 3 2 1 1 1 1 1 1 1 0 1 0 1 Ceratophrys BMNH 18895 1 1 2 1 0 1 2 2 1 1 1 1 1 1 1 1 1 1 1 1 2 2 3 2 1 1 ? ? ? 1 1 1 0 0 ? Ceratophrys calcarata 1 1 1 1 0 1 2 2 1 1 1 0 1 1 1 1 0 1 0 1 2 2 2 2 1 1 1 1 1 1 1 0 0 ? 1 Ceratophrys cornuta 1 1 1 1 0 1 2 2 1 1 1 0 1 1 1 1 0 0 0 1 2 2 2 2 1 1 1 0 1 1 1 0 0 0 0 Ceratophrys cranwelli 1 1 1 1 0 1 2 2 1 1 1 0 1 1 1 1 0 0 1 1 2 2 3 2 0 1 1 0 1 0 1 0 1 0 1 Ceratophrys joazeirensis 1 1 2 1 0 1 2 2 1 1 1 1 1 1 1 1 0 1 0 0 2 2 3 1 1 1 1 1 1 0 1 1 1 2 1 Ceratophrys ornata 1 1 1 1 0 1 2 2 1 1 1 0 1 1 1 1 1 0 0 0 2 2 3 1 0 1 1 0 0 0 1 ? ? ? 1 Ceratophrys rusconii 1 1 1 1 0 1 2 2 1 ? 1 ? 1 1 1 1 0 0 0 0 2 2 3 1 0 1 1 1 ? 1 1 0 0 0 1 Ceratophrys ZUFABC 037 1 1 2 1 0 1 2 2 1 1 1 1 1 1 1 1 1 1 1 1 2 2 3 2 1 1 1 1 1 0 1 0 ? 0 1 Ceratophrys stolzmanni 1 0 1 1 1 1 2 2 1 1 1 0 1 1 1 1 0 1 0 1 2 2 3 2 0 1 1 1 0 0 1 0 1 0 1 Ceratophrys 86.VIII.1.4 1 1 1 1 0 1 2 2 1 1 1 0 1 1 1 1 1 1 0 0 2 2 3 1 0 1 1 0 0 0 1 0 ? ? 1 Chacophrys pierottii 1 0 1 0 1 0 1 0 1 1 1 0 0 1 1 0 0 0 0 0 1 1 1 0 0 ? 1 0 0 0 1 0 0 0 0 Lepidobatrachus asper 1 1 0 0 1 0 2 1 1 0 1 0 0 0 1 0 2 0 0 0 ? 2 1 1 0 0 1 1 2 1 1 1 0 ? 0 Lepidobatrachus australis 1 0 ? 0 1 0 2 1 1 ? 1 0 0 0 1 0 2 0 0 0 2 2 1 1 0 0 1 1 2 1 0 ? ? ? 0 Lepidobatrachus laevis 1 1 0 0 1 0 2 1 1 0 1 0 0 0 1 0 2 0 0 0 2 2 1 1 0 0 1 1 2 1 1 0 0 2 0 Lepidobatrachus llanensis 1 1 0 0 1 0 2 1 1 0 1 0 0 0 1 0 2 0 0 0 2 2 1 1 0 0 1 1 2 1 1 0 0 1 0 Telmatobius thompsoni 0 ? 1 0 ? 0 0 0 0 ? 0 0 0 ? 0 ? 0 0 0 0 0 0 0 0 0 ? 0 ? 1 0 0 ? ? ? 0 95

Species/ Characters 36 37 38 39 40 41 42 43 44 45 46 47 48 49 50 51 52 53 54 55 56 57 58 59 60 61 62 63 64 65 66 67 68 69 70 Alsodes nodosus 1 0 0 0 ? 0 0 1 0 1 0 0 ? ? 0 ? 0 0 0 0 0 0 0 0 0 3 0 0 0 0 0 0 ? 0 ? Ceratophrys ameghinorum 1 0 1 1 0 1 1 0 1 0 1 0 0 2 ? ? 1 1 ? ? 1 ? ? 0 1 1 1 1 1 2 ? ? ? 1 0 Ceratophrys aurita 1 0 1 1 4 1 1 0 1 0 1 1 1 1 1 1 1 1 1 0 1 ? 1 0 1 2 1 1 1 2 1 1 2 1 1 Ceratophrys BMNH 18895 1 ? 1 1 4 1 1 0 1 0 1 1 1 2 ? ? 1 1 ? ? ? ? ? ? 1 2 1 1 1 2 ? ? ? 1 1 Ceratophrys calcarata 1 0 1 1 2 1 1 0 1 0 1 0 2 2 1 1 1 1 0 0 1 ? 0 0 1 1 1 1 1 1 0 0 ? 1 0 Ceratophrys cornuta 1 0 1 1 1 1 1 0 1 0 1 0 2 2 1 1 1 1 1 0 1 ? 0 0 1 2 1 1 1 2 1 0 ? 1 0 Ceratophrys cranwelli 1 0 0 1 1 1 2 0 1 0 1 1 0 3 1 0 1 1 1 0 1 ? 0 0 1 2 1 1 1 2 1 1 2 1 0 Ceratophrys joazeirensis 1 0 1 1 2 1 1 0 1 0 1 1 0 3 1 2 1 1 0 0 1 ? 0 0 1 2 1 1 1 2 1 1 2 1 0 Ceratophrys ornata 1 0 1 1 ? 1 1 0 1 0 1 1 1 3 1 1 1 1 0 0 1 ? 1 0 1 2 1 1 1 2 1 1 2 1 0 Ceratophrys rusconii 1 1 1 1 0 1 1 0 1 0 1 0 0 3 ? ? 1 1 ? ? ? ? ? ? 1 1 1 1 1 1 ? ? ? 1 0 Ceratophrys ZUFABC 037 1 0 1 1 3 1 1 0 1 0 1 1 3 1 1 2 1 1 1 0 1 ? 1 0 1 2 1 1 1 2 ? ? ? 1 1 Ceratophrys stolzmanni 1 0 1 1 2 1 1 0 1 0 1 0 0 3 1 1 1 1 0 0 1 ? 1 0 1 1 1 1 1 2 1 0 ? 1 0 Ceratophrys 86.VIII.1.4 1 0 1 1 1 1 1 0 1 0 1 1 1 1 ? ? 1 1 ? ? ? ? ? ? 1 2 1 1 1 2 ? ? ? 1 0 Chacophrys pierottii 1 0 1 0 1 1 2 1 1 1 0 1 0 1 1 ? 1 1 1 0 1 1 0 0 0 1 1 0 0 1 1 0 ? 1 0 Lepidobatrachus asper 1 1 1 1 0 1 3 1 1 0 1 0 0 2 1 2 1 1 0 1 1 ? 1 1 1 1 1 1 1 3 1 1 0 1 0 Lepidobatrachus australis 1 1 1 1 0 1 3 1 1 0 1 0 0 1 ? ? 1 1 ? ? ? ? ? ? 1 1 1 1 0 ? ? ? ? 1 0 Lepidobatrachus laevis 1 1 1 1 0 0 3 1 1 0 1 0 0 3 1 0 1 1 0 1 1 ? 1 1 1 2 1 1 1 3 1 0 ? 1 0 Lepidobatrachus llanensis 1 1 1 1 0 1 3 1 1 0 1 0 0 1 1 0 1 1 0 1 1 ? 1 1 1 3 1 1 1 3 1 1 1 1 0 Telmatobius thompsoni 1 0 0 0 ? 0 0 1 1 1 0 0 0 0 0 ? 0 0 1 0 0 0 0 0 0 2 0 0 0 1 0 0 ? 0 ? 96

APPENDIX III

Specimens MACN 14319, MACN 14322, MACN 14323, and MACN 14325.

Figure 1: Ceratophrys sp. MACN 14319 in dorsal view, ventral view, frontal view, posterior view and lateral view.

Figure 2: Ceratophrys sp. MACN 14323 in dorsal view and ventral view.

Figure 3: Ceratophrys sp. MACN 14322 MACN 14325 in dorsal view, ventral view and posterior view. 97

Figure 1

Figure 2

Figure 3

100

CONCLUSÃO

Os dois capítulos apresentados oferecem uma ampla visão do conhecimento que temos atualmente sobre os fósseis de Ceratophryidae e os problemas taxonômicos que envolvem a família. Demonstramos que o registro fóssil de Ceratophryidae é riquíssimo, mas que ainda existem muitas lacunas a serem trabalhadas e expomos o potencial do grupo para discussões sobre paleobiogeografia. Atestamos o uso de anatomia comparada de grupos recentes como ferramenta útil a identificação e compreensão da variação observada no registro fóssil do grupo e estabelecimento de caracteres osteológicos, ainda que de um conjunto disponível relativamente restrito - como é o caso dos fósseis.

No Capítulo 1 concluímos que Ceratophrys sagani representa uma nova espécie fóssil de Ceratophrys proximamente relacionada ao grupo de espécies C. aurita, baseado na presença de todas características diagnósticas propostas para o gênero, Ceratophrys sagani é único dentre as espécies de Ceratophryidae por apresentar: 1) dois forâmens formados na sutura do esfenetimoide com o vômer, em vista ventral; 2) eminencia epiótica do otoccipital com uma concavidade cotiloide, parcialmente recoberta pelo processos posterior do frontoparietal; 3) pars interna plectri da columela com três cristas curvas. Devido a atualização sistemática de Baurubatrachus pricei como uma espécie não proximamente relacionada a Ceratophryidae (Nicoli et al. 2016; Báez & Gómez 2017), Ceratophrys sagani pode ser reconhecida como a primeira espécie fóssil de Ceratophryidae e o segundo espécime da família registrado para o Brasil. Praticamente todos fósseis de Ceratophryidae são registrados mais ao sul da América do Sul, Ceratophrys sagani é um dos poucos fósseis de Ceratophryidae registrados mais ao norte.

No Capítulo 2 propomos a primeira análise filogenética a incluir espécimes fósseis de Ceratophryidae, apresentamos uma hipótese de posicionamento filogenético de seis especimes fósseis. Discutimos problemas na taxonomia de fósseis da família e retificamos a identificação taxonômica de alguns desses espécimes. Em nossa análise filogenética recuperamos C. ameghinorum no grupo de espécies C. cornuta, C. rusconii como grupo-irmão de todas as outras espécies de Ceratophrys, Ceratophrys sagani como grupo-irmão de C. aurita + Ceratophrys cf. C. aurita BMNH 18895, Ceratophrys cf. C. aurita BMNH 18895 como um representante fóssil de C. aurita, corroboramos C. ornata MLP 86.VIII.1.4 como um representante fóssil de C. ornata, e L. australis como uma espécie extinta de Lepidobatrachus. Nossa topologia apresenta os dois grupos de espécies 101 de Ceratophrys (Lynch 1982; Faivovich et al. 2014), a exceção da espécie C. stolzmanni que nesta Dissertação é recuperada como grupo-irmão do grupo de espécies C. aurita. Apesar da quantidade de missing data presente na codificação de alguns fósseis, o consenso estrito apresenta alguns clados bem resolvidos e bem suportados. Nossa topologia revela que o monofiletismo da família e dos gêneros é bem suportado por sinapomorfias, todavia alguns caracteres são homoplásicos (e.g. formato da placa ótica, cristas craneanas). Estas características devem ser re-acessadas a posteriori diante de novas amostras fósseis e da compreensão da variação intraespecífica e ontogenética em espécies viventes. Por fim, fica claro que ainda há muito a ser investigado e compreendido a respeito do prolífico registro fóssil da família Ceratophryidae.

102

Atividades realizadas durante o mestrado

Participações em eventos durante o período 1. I Ciclo de Palestras em Evolução e Diversidade. 2017. 2. IV Workshop Evolução e Diversidade. 2017. 3. I Workshop de Genômica Comparada. 2017. 4. VIII Congresso Brasileiro de Herpetologia. Campo Grande, 2017. 5. VI Semana da Biologia UFABC. 2017. 6. VI Curso de Verão em Zoologia. Universidade de São Paulo, 2018. 7. XXXII Congresso Brasileiro de Zoologia. Foz do Iguaçú, 2018. 8. VII Semana da Biologia da UFABC. 2018. 9. XIX Congreso Argentino de Herpetología, La Plata 2018. 10. 1st Virtual Paleontology Congress, Valencia 2018. 11. V Simpósio de Zoologia e Sistemática da UFMG, Belo Horizonte, 2018. 12. IX Encontro de Biologia Comparada da FFCLRP-USP, Ribeirão Preto, 2019. 13. 9º Congresso Brasileiro de Herpetologia, UNICAMP Campinas, 2019.

Resumos apresentados em congresso 1. Barcelos L.A.; Verdade V.K. Descrição e posicionamento filogenético de um Ceratophrys (Ceratophryidae: Anura) proveniente da Caverna Versalles (Pleistoceno final/Holoceno inicial), Apiaí, SP, com comentários sobre paleobiogeografia e paleoambientes da América do Sul. IV Workshop de Evolução e Diversidade, 2017. 2. Dos Santos; Porfiri, J.D. ; Valieri J ; Riff D.S. ; Barcelos, L. A.. Nuevo registro de anuros (Grupo Neuquén: Cretácico Tardío) de la Localidad de Sierra Chata, Neuquén. In: Iº Reunión de Paleovertebrados de la Cuenca Neuquina, 2017, Neuquén. Iº Reunión de Paleovertebrados de la Cuenca Neuquina, 2017. 3. Barcelos, L. A.; Muniz, F.; Hsiou, A. S.; Riff D.S.; Verdade, V. K.. Inferring Paleoenvironment from Anura Fossils. In: XXXII Congresso Brasileiro de Zoologia, 2018, Foz do Iguaçú, PR. 4. Barcelos, L.A., Santos, C.M.D., Verdade, V.K. (2018). A new fossil of Ceratophryidae, late Pleistocene-early Holocene, Brazil. In: II Simposio de Paleontología: “Paleoherpetología: una mirada al pasado de los anfibios y los reptiles”. Libro de Resumenes. XIX Congreso Argentino de Herpetología, La Plata, 33. 103

5. Barcelos L.A. & Souza R.G. (2018). Evolutionary Convergence: the homoplasts explanatory level. Simpósio de Zoologia e Sistemática da UFMG, Belo Horizonte. 6. Barcelos L.A. & Souza R.G. (2018). Natural Selection present in Mass Extinction Events. 1st Virtual Paleontology Congress. Apresentação oral (video). Valencia, 2018. 7. Barcelos L.A. & Verdade V. K. (2019). Reassessment of Ceratophrys (Ceratophryidae: Anura) fossil species Günther (1859) from Pleistocene of Lagoa Santa, MG. Caderno de resumos do IX Encontro de Biologia Comparada, FFCLRP, USP Ribeirão Preto, Brazil. 8. Barcelos L.A. & Verdade V. K. (2019). A new Ceratophrys aurita (Ceratophryidae: Anura) fossil from late Pleistocene-early Holocene of Versalles cave, Southeastern Brazil. 9° Congresso Brasileiro de Herpetologia, UNICAMP, Campinas.

Minicurso aprovado em congresso 1. Barcelos L. A., Muniz F. P. Origem e história evolutiva de Anura contada através dos fósseis. Aprovado no 8° Congresso Brasileiro de Herpetologia e no XXXII Congresso Brasileiro de Zoologia, mas não alcançaram o quórum estabelecido pela organização.

Minicurso ministrado 1. Barcelos L.A. História da Biologia e o Latim como língua da ciência. V Semana da Biologia da UFABC 2017.

Palestra ministrada 1. Barcelos L.A. E agora que sou cientista? Desafios de se fazer e promover ciência no Brasil. UFABC para Todos, Santo André, 2018.

Organização de eventos 1. de Setta ; Centeno, D. ; Barcelos, L. A. . I Workshop de Genômica Comparada. 2017. 2. Freitas, S. R. ; Barcelos, L. A. . VI Semana da Biologia UFABC. 2017. 3. Freitas, S. R. ; Barcelos, L. A. . IV Workshop do Ppg de Evolução e Diversidade. 2017. 4. Freitas, S. R. ; Barcelos, L. A. . VII Semana da Biologia UFABC. 2018. 5. Freitas, S. R. ; Barcelos, L. A. . V Workshop de Evolução e Diversidade. 2018. 6. Freitas, S. R. ; Barcelos, L. A. . VIII Semana da Biologia UFABC. 2019.

Editor de Anais de Eventos 104

1. Freitas, S. R.; Barcelos, L. A.. IV Workshop do Programa de Pós-Graduação em Evolução e Diversidade. 2017. 2. Freitas, S. R.; Barcelos, L. A.. V Workshop de Evolução e Diversidade. 2018.

Vice-representante discente do Programa de Pós-Graduação em Evolução e Diversidade entre os anos 2017-18

Coleções visitadas: 1. Museu de Zoologia da USP. 2. Museu de Ciências da Terra, Departamento Nacional de Produção Mineral (DNPM/CPRM). 3. Museo de Ciencias Naturales de La Plata, Universidad Nacional de La Plata. 4. Museo Argentino de Ciencias Naturales “Bernardino Rivadavia”, Universidad de Buenos Aires. 5. Coleção de Paleovertebrados do Laboratório de Mastozoologia da UNIRIO.