The value of updating GenBank accessions for supermatrix phylogeny: The case of the New Guinean marsupial carnivore genus Myoictis.

Erroneous taxonomic attributions in GenBank accessions can mislead phylogenetic inference and appear to be widespread within genera. We investigate the influence of taxonomic misattributions for reconstructing the phylogeny of three-striped dasyures, which include four recognized Myoictis species (Marsupialia: Dasyuridae) that are distributed across New Guinea and nearby islands. Molecular phylogenetic studies that have focused on dasyurids consistently resolve the interrelationships of these small carnivores, grouping M. leucura with M. wavicus, and placing M. wallacei and M. melas as successively deeper divergences from these. Two recent marsupial and mammalian supermatrix phylogenies instead favour an alternative Myoictis topology that is discordant with each of these relationships. We add new nuclear and mitochondrial sequences and employ randomized accession resampling that shows the supermatrix topologies are an artefact of several outdated taxonomic attributions in GenBank. Updating these accessions brings agreement across Myoictis phylogenies with randomly resampled accessions. We encourage authors to update GenBank taxonomic attributions and we argue that an option is needed for flagging accessions that are not demonstrably incorrect, but that provide anomalous results. This would serve both as a caution for future supermatrix construction and to highlight accessions of potentially significant biological interest for further study.

Niche conservatism and phylogenetic clustering in a tribe of arid-adapted marsupial mice, the Sminthopsini.

The progressive expansion of the Australian arid zone during the last 20 Ma appears to have spurred the diversification of several families of plants, vertebrates and invertebrates, yet such taxonomic groups appear to show limited niche radiation. Here, we test whether speciation is associated with niche conservatism (constraints on ecological divergence) or niche divergence in a tribe of marsupial mice (Sminthopsini; 23 taxa) that includes the most speciose genus of living dasyurids, the sminthopsins. To that end, we integrated phylogenetic data with ecological niche modelling, to enable us to reconstruct the evolution of climatic suitability within Sminthopsini. Niche overlap among species was low-moderate (but generally higher than expected given environmental background similarity), and the degree of phylogenetic clustering increased with aridity. Climatic niche reconstruction illustrates that there has been little apparent evolution of climatic tolerance within clades. Accordingly, climatic disparity tends to be accumulated among clades, suggesting considerable niche conservatism. Our results also indicate that evolution of climatic tolerances has been heterogeneous across different dimensions of climate (temperature vs. precipitation) and across phylogenetic clusters (Sminthopsis murina group vs. other groups). Although some results point to the existence of shifts in climatic niches during the speciation of sminthopsins, our study provides evidence for substantial phylogenetic niche conservatism in the group. We conclude that niche diversification had a low impact on the speciation of this tribe of small, but highly mobile marsupials.

Investigating an approach to the alliance based on interpersonal defense theory.

OBJECTIVE: Notwithstanding consistent findings of significant relationships between the alliance and outcome, questions remain to be answered about the relatively small magnitude of those correlations, the mechanisms underlying the association, and how to conceptualize the alliance construct. We conducted a preliminary study of an approach to the alliance based on interpersonal defense theory, which is an interpersonal reconceptualization of defense processes, to investigate the promise of this alternative approach as a way to address the outstanding issues. METHOD: We employed qualitative, theory-building case study methodology, closely examining alliance processes at four time points in the treatment of a case in terms of a case formulation based on interpersonal defense theory. RESULTS: The results suggested that our approach made it possible to recognize key processes in the alliance and that it helps explain how the alliance influences outcome. Our analyses also provided a rich set of concrete illustrations of the alliance phenomena identified by the theory. CONCLUSIONS: The findings suggest that an approach to the alliance based on interpersonal defense theory holds promise. However, although the qualitative method we employed has advantages, it also has limitations. We offer suggestions about how future qualitative and quantitative investigations could build on this study.

What's in a Transition? An Integrative Perspective on Transitions in Medical Education.

This Conversation Starters article presents a selected research abstract from the 2016 Association of American Medical Colleges Western Region Group on Educational Affairs annual spring meeting. The abstract is paired with the integrative commentary of three experts who shared their thoughts stimulated by the needs assessment study. These thoughts explore how the general theoretical mechanisms of transition may be integrated with cognitive load theory in order to design interventions and environments that foster transition.

Vicariant speciation resulting from biogeographic barriers in the Australian tropics: The case of the red-cheeked dunnart (Sminthopsis virginiae).

Global biodiversity loss continues unabated, and in Australia, the rate of recent mammal extinctions is among the worst in the world. Meanwhile, the diversity among and within many endemic mammal species remains undescribed. This information is crucial to delineate species boundaries and thus inform decision-making for conservation. Sminthopsis virginiae (the red-cheeked dunnart) is a small, dasyurid marsupial found in four disjunct populations around the northern coast of Australia and New Guinea. There are three currently recognized subspecies, each occupying a distinct geographic location. Sminthopsis v. virginiae occurs in Queensland, S. v. rufigenis is distributed across New Guinea and the Aru Islands, and S. v. nitela has populations in the Top End of the Northern Territory and the Kimberley region of Western Australia. Previous molecular work has suggested the current subspecies definitions are not aligned with DNA sequence data, though the sampling was limited. We undertook a comprehensive genetic and morphological review of S. virginiae to clarify relationships within the species. This included mitochondrial (CR, 12S, and cytb) and nuclear (omega-globin, IRBP, and bfib7) loci, and morphometric analysis of skulls and whole wet-preserved specimens held in museums. Maximum Likelihood and Bayesian phylogenetic analyses resolved samples into two distinct clades, demarcated by the Gulf of Carpentaria in Australia's north. Sminthopsis. v. nitela was consistently separated from S. v. virginiae and S. v. rufigenis, based on the overall body and skull size and craniodental features, while S. v. virginiae and S. v. rufigenis were more difficult to distinguish from each other. Thus, we redescribed S. virginiae, recognizing two species, S. nitela (raised from subspecies) and S. virginiae (now comprising the subspecies S. v. virginiae and S. v. rufigenis). This study highlights the importance of recognizing cryptic mammal fauna to help address the gap in our knowledge about diagnosing diversity during a time of conservation crisis.

Mitogenome of the extinct Desert 'rat-kangaroo' times the adaptation to aridity in macropodoids.

The evolution of Australia's distinctive marsupial fauna has long been linked to the onset of continent-wide aridity. However, how this profound climate change event affected the diversification of extant lineages is still hotly debated. Here, we assemble a DNA sequence dataset of Macropodoidea-the clade comprising kangaroos and their relatives-that incorporates a complete mitogenome for the Desert 'rat-kangaroo', Caloprymnus campestris. This enigmatic species went extinct nearly 90 years ago and is known from a handful of museum specimens. Caloprymnus is significant because it was the only macropodoid restricted to extreme desert environments, and therefore calibrates the group's specialisation for increasingly arid conditions. Our robustly supported phylogenies nest Caloprymnus amongst the bettongs Aepyprymnus and Bettongia. Dated ancestral range estimations further reveal that the Caloprymnus-Bettongia lineage originated in nascent xeric settings during the middle to late Miocene, ~ 12 million years ago (Ma), but subsequently radiated into fragmenting mesic habitats after the Pliocene to mid-Pleistocene. This timeframe parallels the ancestral divergences of kangaroos in woodlands and forests, but predates their adaptive dispersal into proliferating dry shrublands and grasslands from the late Miocene to mid-Pleistocene, after ~ 7 Ma. We thus demonstrate that protracted changes in both climate and vegetation likely staged the emergence of modern arid zone macropodoids.

Cytogenetics meets phylogenetics: a review of karyotype evolution in diprotodontian marsupials.

We have used a combined approach of phylogenetics and cytogenetics to describe karyotype evolution in Diprotodontia. Molecular relationships of diprotodontian marsupials have been clarified using a concatenation of 5 nuclear gene sequences from multiple exemplars of all extant genera. Our well-resolved phylogenetic tree has been used as a basis for understanding chromosome evolution both within this Order, as well as in marsupials in general. It is clear that the ancestral marsupial karyotype comprised 14 relatively large chromosomes of the form retained relatively unchanged in caenolestids, microbiotherians, peramelemorphians, vombatids, and pygmy possums. Four pericentric inversions occurred in the ancestral dasyuromorphian (chromosomes 1, 2, 4, and 6) and a different 4 in the ancestral didelphimorphian (chromosomes 1, 3, 5 and 6). Within Diprotodontia, although the ancestral marsupial karyotype has been retained in some families such as the extant wombats and pygmy possums, there have been major karytoypic repatternings early in the evolution of others. Chromosome rearrangements in diprotodontia include centric fissions and fusions, translocations, and centromere shifts. Karyotypic changes are discussed in the context of current hypotheses concerning centromeres, chromosomal fragile sites, and mobile elements in marsupials and the probable repeated involvement of these elements in karyotypic restructuring.

A phylogeny of Diprotodontia (Marsupialia) based on sequences for five nuclear genes.

Even though the marsupial order Diprotodontia is one of the most heavily studied groups of Australasian marsupials, phylogenetic relationships within this group remain contentious. The more than 125 living species of Diprotodontia can be divided into two main groups: Vombatiformes (wombats and koalas) and Phalangerida. Phalangerida is composed of the kangaroos (Macropodidae, Potoroidae, and Hypsiprymnodontidae) and possums (Phalangeridae, Burramyidae, Petauridae, Pseudocheiridae, Tarsipedidae, and Acrobatidae). Much of the debate has focused on relationships among the families of possums and whether possums are monophyletic or paraphyletic. A limitation of previous investigations is that no study to date has investigated diprotodontian relationships using all genera. Here, we examine diprotodontian interrelationships using a nuclear multigene molecular data set representing all recognized extant diprotodontian genera. Maximum parsimony, maximum likelihood, and Bayesian methods were used to analyze sequence data obtained from protein-coding portions of ApoB, BRCA1, IRBP, Rag1, and vWF. We also applied a Bayesian relaxed molecular clock method to estimate times of divergence. Diprotodontia was rooted between Vombatiformes and Phalangerida. Within Phalangerida, the model-based methods strongly support possum paraphyly with Phalangeroidea (Burramyidae+Phalangeridae) grouping with the kangaroos (Macropodiformes) to the exclusion of Petauroidea (Tarsipedidae, Acrobatidae, Pseudocheiridae, and Petauridae). Within Petauroidea, Tarsipedidae grouped with both Petauridae and Pseudocheiridae to the exclusion of Acrobatidae. Our analyses also suggest that the diprotodontian genera Pseudochirops and Strigocuscus are paraphyletic and diphyletic, respectively, as currently recognized. Dating analyses suggest Diprotodontia diverged from other australidelphians in the late Paleocene to early Eocene with all interfamilial divergences occurring prior to the early Miocene except for the split between the Potoroidae and Macropodidae, which occurred sometime in the mid-Miocene. Ancestral state reconstructions using a Bayesian method suggest that the patagium evolved independently in the Acrobatidae, Petauridae, and Pseudocheiridae. Ancestral state reconstructions of ecological venue suggest that the ancestor of Diprotodontia was arboreal. Within Diprotodontia, the common ancestor of Macropodidae was reconstructed as terrestrial, suggesting that tree kangaroos (Dendrolagus) are secondarily arboreal.

Marsupial genetics and genomics.

Marsupials, the 'other' mammals, are found only in Australasia and the Americas. They are quite different from eutherian ('placental') mammals, as well they might be after 130 million years of separate evolution. They display a unique pattern of mammalian organization and development that is reflected by differences in their genomes. Here, we introduce marsupials as alternative (but not inferior!) mammals and summarize the state of knowledge of marsupial relationships, marsupial chromosomes, maps, genes and genetic regulatory systems. We shamelessly present the case for a Kangaroo Genome Project.

Bandicoot fossils and DNA elucidate lineage antiquity amongst xeric-adapted Australasian marsupials.

Bandicoots (Peramelemorphia) are a unique order of Australasian marsupials whose sparse fossil record has been used as prima facie evidence for climate change coincident faunal turnover. In particular, the hypothesized replacement of ancient rainforest-dwelling extinct lineages by antecedents of xeric-tolerant extant taxa during the late Miocene (~10 Ma) has been advocated as a broader pattern evident amongst other marsupial clades. Problematically, however, this is in persistent conflict with DNA phylogenies. We therefore determine the pattern and timing of bandicoot evolution using the first combined morphological + DNA sequence dataset of Peramelemorphia. In addition, we document a remarkably archaic new fossil peramelemorphian taxon that inhabited a latest Quaternary mosaic savannah-riparian forest ecosystem on the Aru Islands of Eastern Indonesia. Our phylogenetic analyses reveal that unsuspected dental homoplasy and the detrimental effects of missing data collectively obscure stem bandicoot relationships. Nevertheless, recalibrated molecular clocks and multiple ancestral area optimizations unanimously infer an early diversification of modern xeric-adapted forms. These probably originated during the late Palaeogene (30-40 Ma) alongside progenitors of other desert marsupials, and thus occupied seasonally dry heterogenous habitats long before the onset of late Neogene aridity.

Germline viral "fossils" guide in silico reconstruction of a mid-Cenozoic era marsupial adeno-associated virus.

Germline endogenous viral elements (EVEs) genetically preserve viral nucleotide sequences useful to the study of viral evolution, gene mutation, and the phylogenetic relationships among host organisms. Here, we describe a lineage-specific, adeno-associated virus (AAV)-derived endogenous viral element (mAAV-EVE1) found within the germline of numerous closely related marsupial species. Molecular screening of a marsupial DNA panel indicated that mAAV-EVE1 occurs specifically within the marsupial suborder Macropodiformes (present-day kangaroos, wallabies, and related macropodoids), to the exclusion of other Diprotodontian lineages. Orthologous mAAV-EVE1 locus sequences from sixteen macropodoid species, representing a speciation history spanning an estimated 30 million years, facilitated compilation of an inferred ancestral sequence that recapitulates the genome of an ancient marsupial AAV that circulated among Australian metatherian fauna sometime during the late Eocene to early Oligocene. In silico gene reconstruction and molecular modelling indicate remarkable conservation of viral structure over a geologic timescale. Characterisation of AAV-EVE loci among disparate species affords insight into AAV evolution and, in the case of macropodoid species, may offer an additional genetic basis for assignment of phylogenetic relationships among the Macropodoidea. From an applied perspective, the identified AAV "fossils" provide novel capsid sequences for use in translational research and clinical applications.

Impacts of the Cretaceous Terrestrial Revolution and KPg extinction on mammal diversification.

Previous analyses of relations, divergence times, and diversification patterns among extant mammalian families have relied on supertree methods and local molecular clocks. We constructed a molecular supermatrix for mammalian families and analyzed these data with likelihood-based methods and relaxed molecular clocks. Phylogenetic analyses resulted in a robust phylogeny with better resolution than phylogenies from supertree methods. Relaxed clock analyses support the long-fuse model of diversification and highlight the importance of including multiple fossil calibrations that are spread across the tree. Molecular time trees and diversification analyses suggest important roles for the Cretaceous Terrestrial Revolution and Cretaceous-Paleogene (KPg) mass extinction in opening up ecospace that promoted interordinal and intraordinal diversification, respectively. By contrast, diversification analyses provide no support for the hypothesis concerning the delayed rise of present-day mammals during the Eocene Period.

A Phylogeny and Timescale for the Evolution of Pseudocheiridae (Marsupialia: Diprotodontia) in Australia and New Guinea.

PSEUDOCHEIRIDAE (MARSUPIALIA: Diprotodontia) is a family of endemic Australasian arboreal folivores, more commonly known as ringtail possums. Seventeen extant species are grouped into six genera (Pseudocheirus, Pseudochirulus, Hemibelideus, Petauroides, Pseudochirops, Petropseudes). Pseudochirops and Pseudochirulus are the only genera with representatives on New Guinea and surrounding western islands. Here, we examine phylogenetic relationships among 13 of the 17 extant pseudocheirid species based on protein-coding portions of the ApoB, BRCA1, ENAM, IRBP, Rag1, and vWF genes. Maximum parsimony, maximum likelihood, and Bayesian methods were used to estimate phylogenetic relationships. Two different relaxed molecular clock methods were used to estimate divergence times. Bayesian and maximum parsimony methods were used to reconstruct ancestral character states for geographic provenance and maximum elevation occupied. We find robust support for the monophyly of Pseudocheirinae (Pseudochirulus + Pseudocheirus), Hemibelidinae (Hemibelideus + Petauroides), and Pseudochiropsinae (Pseudochirops + Petropseudes), respectively, and for an association of Pseudocheirinae and Hemibelidinae to the exclusion of Pseudochiropsinae. Within Pseudochiropsinae, Petropseudes grouped more closely with the New Guinean Pseudochirops spp. than with the Australian Pseudochirops archeri, rendering Pseudochirops paraphyletic. New Guinean species belonging to Pseudochirops are monophyletic, as are New Guinean species belonging to Pseudochirulus. Molecular dates and ancestral reconstructions of geographic provenance combine to suggest that the ancestors of extant New Guinean Pseudochirops spp. and Pseudochirulus spp. dispersed from Australia to New Guinea ∼12.1-6.5 Ma (Pseudochirops) and ∼6.0-2.4 Ma (Pseudochirulus). Ancestral state reconstructions support the hypothesis that occupation of high elevations (>3000 m) is a derived feature that evolved on the terminal branch leading to Pseudochirops cupreus, and either evolved in the ancestor of Pseudochirulus forbesi, Pseudochirulus mayeri, and Pseudochirulus caroli, with subsequent loss in P. caroli, or evolved independently in P. mayeri and P. forbesi. Divergence times within the New Guinean Pseudochirops clade are generally coincident with the uplift of the central cordillera and other highlands. Diversification within New Guinean Pseudochirulus occurred in the Plio-Pleistocene after the establishment of the Central Range and other highlands. ELECTRONIC SUPPLEMENTARY MATERIAL: The online version of this article (doi:10.1007/s10914-010-9129-7) contains supplementary material, which is available to authorized users.

A timescale and phylogeny for "bandicoots" (Peramelemorphia: Marsupialia) based on sequences for five nuclear genes.

Relationships among the living and recently extinct genera of bandicoots (Marsupialia: Peramelemorphia) have proven difficult to discern. Previous phylogenetic studies have used only morphology or mitochondrial DNA and have reported conflicting results in regards to their relationships. Most phylogenetic reconstructions recognize a basal split between the bilby Macrotis (Thylacomyidae) and the Peramelidae. The Peramelidae is composed of the Peramelinae (Isoodon and Perameles), Echymiperinae (Echymipera and Microperoryctes), and Peroryctinae (Peroryctes). Within Peramelidae, Echymipera and Microperoryctes usually group together to the exclusion of Peroryctes. This clade is sister to the Peramelinae. Placement of the recently extinct pig-footed bandicoot (Chaeropus: Chaeropodidae) has been ambiguous. We address the interrelationships and estimate times of divergence for the living bandicoot genera using a 6 kilobase concatenation consisting of protein-coding regions of five nuclear genes (ApoB, BRCA1, IRBP, Rag1, and vWF). We analyzed this concatenation using maximum parsimony, maximum likelihood, and Bayesian methods and estimated times of divergence using two Bayesian relaxed molecular clock methods. In all concatenated analyses, all nodes associated with the Peramelemorphia were robustly supported (bootstrap support percentages=100; posterior probabilities=1.00). Macrotis was recovered as basal to the remaining living bandicoots. Within the Peramelidae, Echymipera and Microperoryctes grouped to the exclusion of Peroryctes and this clade was sister to the Peramelinae. Only Rag1 amplified for Chaeropus; analyses based on this gene provide moderate support for an association of Chaeropus plus Peramelidae to the exclusion of Macrotis. Both relaxed clock Bayesian methods suggest that the living bandicoots are a relatively recent radiation originating sometime in the late Oligocene or early Miocene with subsequent radiations in the late Miocene to early Pliocene.

Phylogenetic analysis of 18S rRNA and the mitochondrial genomes of the wombat, Vombatus ursinus, and the spiny anteater, Tachyglossus aculeatus: increased support for the Marsupionta hypothesis.

The monotremes, the duck-billed platypus and the echidnas, are characterized by a number of unique morphological characteristics, which have led to the common belief that they represent the living survivors of an ancestral stock of mammals. Analysis of new data from the complete mitochondrial (mt) genomes of a second monotreme, the spiny anteater, and another marsupial, the wombat, yielded clear support for the Marsupionta hypothesis. According to this hypothesis marsupials are more closely related to monotremes than to eutherians, consistent with a basal split between eutherians and marsupials/monotremes among extant mammals. This finding was also supported by analysis of new sequences from a nuclear gene--18S rRNA. The mt genome of the wombat shares some unique features with previously described marsupial mtDNAs (tRNA rearrangement, a missing tRNA(Lys), and evidence for RNA editing of the tRNA(Asp)). Molecular estimates of genetic divergence suggest that the divergence between the platypus and the spiny anteater took place approximately 34 million years before present (MYBP), and that between South American and Australian marsupials approximately 72 MYBP.

Nuclear gene sequences provide evidence for the monophyly of australidelphian marsupials.

Relationships among the seven extant orders of marsupials remain poorly understood. Most classifications recognize a fundamental split between Ameridelphia, which contains the American orders Didelphimorphia and Paucituberculata, and Australidelphia, which contains four Australasian orders (Dasyuromorphia, Diprotodontia, Notoryctemorphia, and Peramelina) and the South American order Microbiotheria, represented by Dromiciops gliroides. Ameridelphia and Australidelphia are each supported by key morphological characters with dichotomous character states. To date, molecular studies indexing all marsupial orders have reported inconclusive results. However, several studies have suggested that Dromiciops is nested within Australidelphia. This result has important implications for understanding the biogeographic history of living marsupials. To address questions in higher-level marsupial systematics, we sequenced portions of five nuclear genes (Apolipoprotein B gene; Breast and Ovarian cancer susceptibility gene 1; Recombination activating gene 1; Interphotoreceptor retinoid binding protein gene; and von Willebrand factor gene) for representatives of all orders of marsupials, as well as placental outgroups. The resulting 6.4kb concatenation was analyzed using maximum parsimony, distance methods, maximum likelihood, and Bayesian methods. tests were used to examine a priori hypotheses. All analyses provided robust support for the monophyly of Australidelphia (bootstrap support=99-100%; posterior probability=1.00). Ameridelphia received much lower support, although this clade was not rejected in statistical tests. Within Diprotodontia, both Vombatiformes and Phalangeriformes were supported at the 100% bootstrap level and with posterior probabilities of 1.00.