Tracing the evolutionary history of two endemic ground frogs of temperate forest of southern Chile, through molecular and cytogenetic approaches.
Conicyt InstrumentBecas Nacionales
Eupsophus is a genus of ground frogs, endemic from Southern Chile and Argentina. The taxonomy and systematic of the genus have been controversial since its founding. Currently, Eupsophus is composed by 10 species, which have been match up into two groups: The roseus group that comprise eight species (2n=30), and the vertebralis group with two species E. vertebralis, and E. emiliopugini (2n=28). Within the vertebralis group, geographic distributio... Ver más
Eupsophus is a genus of ground frogs, endemic from Southern Chile and Argentina. The taxonomy and systematic of the genus have been controversial since its founding. Currently, Eupsophus is composed by 10 species, which have been match up into two groups: The roseus group that comprise eight species (2n=30), and the vertebralis group with two species E. vertebralis, and E. emiliopugini (2n=28). Within the vertebralis group, geographic distribution, morphologic and genetic variation remain unclear and without hypotheses about the relationships between these lineages. Moreover, morphology karyotype description made by Giemsa stained, evidenced differences on the fundamental number (FN) between E. vertebralis (FN= 54) and E. emiliopugini (FN= 56). Nevertheless, the consistency of this report at intraspecific levels and with others cytogenetics techniques remain unexplored. Thus, the goals of this thesis were, on the one hand, to evaluated phylogenetic relationships, diversification times, and to determine species limits of Eupsophus vertebralis and E. emiliopugini. On the other hand, it was proposed to determine the presences or absences of specie specific cytogenetics patterns for the vertebralis group, and test hypothesis about chromosomal rearrangements, considering individuals from different localities and sexes. For the molecular analyses, three mitochondrial (D-loop, cytb, and coI) and two nuclear (crybA1, and pomc) markers, from 91 individuals of the vertebralis group collected in 19 localities, were sequenced. Then, it was performed phylogenetics analyses with Maximum Likelihood (ML) and Bayesian Inference (IB), and species delimitation analyses using unilocus and multilocus coalescent approaches. For its part, for the cytogenetic analyses it was employed conventional (Giemsa staining, C-banding and, Ag-NOR) and cytomolecular (fluorescence in situ hybridization FISH, using telomeric and 28S ribosomal probes) techniques, on metaphasic plates of 23 individuals of the vertebralis group from 15 different localities. ML and IB phylogenetic reconstructions recovered the monophyly of the vertebralis group (Bootstrap: 100%; Bayesian posterior Probability PP: 1.0), but not the reciprocal monophyly of its species (Bootstrap: < 50; PP: < 80), exhibiting a polyphyletic pattern within the vertebralis group. Incongruence among the six species delimitation analyses carried out, detecting from one to eleven species (Max. congruence index= 0.59, for one putative species; Min. congruence index= 0.18, for eleven putative species). Also, the divergence time between E. emiliopugini and E. vertebralis (0.040 Mya) indicates an evolutionary history probably molded by the interglacial periods of Late Pleistocene. On the other hand, FN differences between E. vertebralis and E. emiliopugini were corroborated through Giemsa staining and C-banding (FN= 54 and 56, respectively). FISH with 28S rDNA probes confirmed the active NORs signal for E. vertebralis and E. emiliopugini (in accordance with the Ag-NORs staining results). Nevertheless, we found polymorphisms relative to the numbers and location of 28S rDNA (four signals) and active NORs (two signals) in one specimen of E. emiliopugini from Puyehue locality. This intraspecific chromosomal variation could be explained by chromosomal rearrangements or derived from hybridization process. FISH using telomeric probe over spreads from both species detected no interstitial fluorescent signals, but clearly stained telomeric regions. The molecular results of this study, suggest low genetic differentiation and the early diversification for the vertebralis group. Also, the cytogenetics results corroborate that the difference in FN between the karyotype of E. vertebralis and E. emiliopugini is conserved at interspecific level. Nonetheless, intraspecific differences for the NORs position were found in one sample of E. emliopugini from Puyehue locality. Therefore, the results do not provide sufficient evidence to support E. emiliopugini and E. vertebralis as one or two species. Hence, the speciation process of E. vertebralis and E. emiliopugini lineages are discussed under an integrative framework, considering the phylogenetic relationships, evolutionary history, geographic antecedents, cytogenetic, and ethologic (calls) data. Finally, new concerning lineage for conservation efforts are proposed. Ver menos
InstitutionUNIVERSIDAD AUSTRAL DE CHILE
Date de publicación2019
Nuñez, José J.
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