Molecular Phylogeny of Cryptonanus (Didelphidae: Thylamyini): Evidence for a recent and complex diversification in South American open biomes.

Systematic revisions of South American marsupials have contributed to our knowledge about genus and species diversity in the last decades, including studies of the most recently described genus Cryptonanus (Didelphidae), currently comprising four recognized species. Herein we provide the first phylogeny for these mouse opossums based on comprehensive sampling, including representatives from all nominal taxa, encompassing most of the geographic distribution of the genus while also extending its known range. The taxonomic status of Cryptonanus species was explored by analyses of multiple mitochondrial and nuclear DNA markers to assess phylogenetic relationships and to provide divergence time estimates, species delimitations and biogeographical hypotheses. Cryptonanus monophyly remained highly supported despite the inclusion of abundant new data from more than a hundred specimens, comprising 10 independent evolutionary lineages. Species-complexes within valid nominal taxa reveal higher species richness in the genus. Based on divergence estimates from a dated phylogeny, we suggest that Cryptonanus diversified along the Quaternary, with speciation events occurring well into the Pleistocene. The best supported biogeographical hypothesis endorses speciation by vicariance and subset speciation across open formations in shaping the evolutionary history of this didelphid genus, strongly associated with dry tropical landscapes of South America.

Molecular Phylogeny of Cryptonanus (Didelphidae: Thylamyini): evidence for a recent and complex diversification in South American open biomes.

Systematic revisions of South American marsupials have contributed to our knowledge about genus and species diversity in the last decades, including studies of the most recently described genus Cryptonanus (Didelphidae), currently comprising four recognized species. Herein we provide the first phylogeny for these mouse opossums based on comprehensive sampling, including representatives from all nominal taxa, encompassing most of the geographic distribution of the genus while also extending its known range. The taxonomic status of Cryptonanus species was explored by analyses of multiple mitochondrial and nuclear DNA markers to assess phylogenetic relationships and to provide divergence time estimates, species delimitations and biogeographical hypotheses. Cryptonanus monophyly remained highly supported despite the inclusion of abundant new data from more than a hundred specimens, comprising 10 independent evolutionary lineages. Species-complexes within valid nominal taxa reveal higher species richness in the genus. Based on divergence estimates from a dated phylogeny, we suggest that Cryptonanus diversified along the Quaternary, with speciation events occurring well into the Pleistocene. The best supported biogeographical hypothesis endorses speciation by vicariance and subset speciation across open formations in shaping the evolutionary history of this didelphid genus, strongly associated with dry tropical landscapes of South America.

A review of Euryoryzomys legatus (Rodentia, Sigmodontinae): morphological redescription, cytogenetics, and molecular phylogeny

The taxonomic history of Euryoryzomys legatus has been complex and controversial, being either included in the synonymy of other oryzomyine species or considered as a valid species, as in the most recent review of the genus. Previous phylogenetic analyses segregated E. legatus from E. russatus, its putative senior synonym, but recovered it nested within E. nitidus. A general lack of authoritative evaluation of morphological attributes, details of the chromosome complement, or other data types has hampered the ability to choose among alternative taxonomic hypotheses, and thus reach a general consensus for the status of the taxon. Herein we revisit the status of E. legatus using an integrated approach that includes: (1) a morphological review, especially centered on specimens from northwestern Argentina not examined previously, (2) comparative cytogenetics, and (3) phylogenetic reconstruction, using mitochondrial genes. Euryoryzomys legatus is morphologically and phylogenetically distinct from all other species-level taxa in the genus, but its 2n=80, FN=86 karyotype is shared with E. emmonsae, E. nitidus, and E. russatus. Several morphological and morphometric characters distinguish E. legatus from other species of Euryoryzomys, and we provide an amended diagnosis for the species. Morphological characters useful in distinguishing E. legatus from E. nitidus, its sister taxon following molecular analyses, include: larger overall size, dorsal fur with a strong yellowish brown to orange brown tinge, flanks and cheeks with an orange lateral line, ventral color grayish-white with pure white hairs present only on the chin, presence of a thin blackish eye-ring, tail bicolored, presence of an alisphenoid strut and a well-developed temporal and lambdoid crests in the skull, and a labial cingulum on M3. Molecular phylogenetic analyses recovered E. legatus as a monophyletic group with high support nested within a paraphyletic E. nitidus; genetic distances segregated members of both species, except for an exemplar of E. nitidus. Our integrated analyses reinforce E. legatus as a full species, but highlight that E. macconnelli, E. emmonsae, and E. nitidus each may be a species complex and worthy of systematic attention. Finally, we also evaluated the chromosome evolution of the genus within a phylogenetic context.

Marcadores genéticos e sua contribuição para a caracterização das espécies e reconstrução da história evolutiva do gênero Euryoryzomys (Rodentia: sigmodontinae)

Morphological similarities among rodents hinders the taxonomy of the group. Previously, the genus Oryzomys had its species allocated in species complexes. Cytogenetics and molecular information pointed out to the existence of cryptic entities among those species complexes. Accordingly, Oryzomys was later split in ten new genera, being Euryoryzomys one of them. Six species and another one not formally described are recognized for the genus. These species are morphologically similar, and have a broad distribution through different phytophysiognomies. Cytogenetic represents an important tool to identify some species, and the karyotype for E. legatus is still missing. Furthermore, systematic studies are also extremely relevant to recover the phylogenetic relationships of the species of the genus and their diversification evolutionary hypotheses. The aim of this study was to describe the karyotype of E. legatus, and to perform phylogenetic analyses on Euryoryzomys species. Chromosomal preparation of E. legatus were analyzed based on conventional Giemsa staining, C and G-banding patterns, and fluorescence in situ hybridization (FISH) using telomeric probes. The karyotype of E. legatus showed 2n=80, NF=86, and FISH analyses showed signals in the telomeric regions. The phylogenetic analyses using all the recognized Euryoryzomys species were based on 83 partial sequences of the mitochondrial gene cytochrome b (cyt-b) - 11 produced in this study - and 17 partial sequences of the nuclear gene interphotoreceptor retinoid binding protein (IRBP) - four obtained in this study -, besides 11 taxa that composed the outgroup. Phylogenetic analyses - applying Bayesian inference and Maximum-Likelihood methods – using concatenated dataset recovered the monophyly of the genus with low support, and the following topology: ((E. macconnelli A, ((E. macconnelli B, E. macconnelli C), E. macconnelli D)), ((E. emmonsae A, E. emmonsae B), (E. russatus, ((E. lamia, Euryoryzomys sp.), (E. nitidus B, (E. nitidus A, (E. nitidus MSB70697, E. legatus))))))). The diploid number, chromosomes morphology and banding patterns of E. legatus were similar to those of E. emmonsae, E. nitidus, and E. russatus described in the literature. Cytogenetic data are informative to distinguish E. legatus from three other species: E. lamia, E. macconnelli, and Euryoryzomys sp. The phylogenetic analyses indicated the hypothesis that E. emmonsae, E. macconnelli, and E. nitidus - and according to the literature, E. lamia - may represent species complexes (cryptic species previously revealed by the cytogenetics data or morphology and corroborated by the phylogenetic reconstruction). Three subclades (northeast, southeast and south-southeast) were recovered for E. russatus, which demonstrates that phylogeographic analyses are needed, based on the sample size. E. nitidus was recovered as polyphyletic, and its phylogenetic relationships with E. legatus must be further investigated. The phylogenetic analyses indicates that species of Euryoryzomys from humid areas should have diverged earlier than species from open and refugia areas. This study increases the knowledge of Euryoryzomys and shows that the difficulty on species delimitation and their diversification processes are still strongly related to sample gaps, given the wide distribution of the genus. Fieldwork expeditions are still required to resolve this basic deficiency. Subsequently, taxonomic revisions are also required, especially under an integrative taxonomy approach.

A acentuada semelhança morfológica entre roedores gera problemas para a sua taxonomia. O gênero Oryzomys, por exemplo, tinha suas espécies alocadas em grupos de espécies, porém, dados citogenéticos e moleculares apontavam para a existência de entidades taxonômicas crípticas nesses grupos. Posteriormente, dez novos gêneros foram reconhecidos a partir de Oryzomys, entre os quais está Euryoryzomys e para o qual, atualmente, são reconhecidas seis espécies e uma não descrita formalmente. Essas espécies são semelhantes morfologicamente e possuem ampla distribuição geográfica, ocupando diferentes fitofisionomias. Os dados citogenéticos auxiliam na identificação de algumas espécies. E. legatus é a única espécie do gênero que não possui cariótipo descrito. Além disso, estudos de sistemática com o gênero são importantes para estabelecer as relações filogenéticas de suas espécies e possíveis hipóteses evolutivas. O objetivo desse estudo foi descrever o cariótipo de E. legatus e realizar análises filogenéticas com marcadores moleculares para as espécies do gênero. Foram analisadas preparações citogenéticas de E. legatus empregando coloração com Giemsa, Banda C, Banda G e hibridação in situ fluorescente (FISH) com sondas teloméricas. O cariótipo de E. legatus apresentou 2n=80, NF=86, e a FISH evidenciou sinais nas regiões teloméricas dos cromossomos. As análises filogenéticas com todas as espécies do gênero contaram com 83 sequências parciais do gene mitocondrial citocromo b (cit-b) - 11 produzidas no presente estudo - e 17 sequências parciais do gene nuclear interphotoreceptor retinoid binding protein (IRBP) - quatro produzidas nesse estudo -, além de 11 táxons compondo o grupo externo. Os métodos de análises empregados foram inferência Bayesiana e Máxima-Verossimilhança. As análises com os marcadores concatenados (cit-b + IRBP) recuperaram o gênero monofilético com baixo suporte e a seguinte topologia: ((E. macconnelli A, ((E. macconnelli B, E. macconnelli C), E. macconnelli D)) ((E. emmonsae A, E. emmonsae B), (E. russatus, ((E. lamia, Euryoryzomys sp.), (E. nitidus B, (E. nitidus A, (E. nitidus MSB70697, E. legatus))))))). O número diploide, morfologia e padrão de bandas de E. legatus foram semelhantes aos de E. emmonsae, E. nitidus e E. russatus descritos na literatura, de modo que dados citogenéticos são informativos somente para distinguir E. legatus de E. lamia, E. macconnelli e Euryoryzomys sp. As reconstruções filogenéticas indicam que E. emmonsae, E. macconnelli, E. nitidus – e, segundo a literatura, E. lamia - possam representar complexos de espécies (espécies crípticas reveladas previamente pela informação citogenética ou morfologia e corroborada pela reconstrução filogenética). Para E. russatus foram recuperados três subclados (nordeste, sudeste e sul-sudeste), demonstrando que são necessárias análises filogeográficas, dado o tamanho da amostra. E. nitidus foi recuperado polifilético e sua relação filogenética com E. legatus deve ser melhor investigada com ampliação de amostragem. As análises sugerem também que espécies de Euryoryzomys de áreas úmidas podem ter divergido antes das espécies de áreas abertas e de refúgios. O presente estudo amplia o conhecimento de Euryoryzomys e demostra que a maior dificuldade na delimitação das espécies e compreensão dos processos envolvidos em sua diversificação ainda é a existência de muitas lacunas amostrais, dada a ampla distribuição do gênero. Expedições de coleta são necessárias para solucionar essa deficiência amostral, bem como revisões taxonômicas, especialmente à luz de uma abordagem de taxonomia integrativa, para um dos gêneros de maior distribuição da tribo Oryzomyini.