Search

Echimyidae is one of the most speciose and ecologically diverse rodent families in the world, occupying a wide range of habitats in the Neotropics. However, a resolved phylogeny at the genus-level is still lacking for these 22 genera of South American spiny rats, including the coypu (Myocastorinae), and 5 genera of West Indian hutias (Capromyidae) relatives. Here, we used Illumina shotgun sequencing to assemble 38 new complete mitogenomes, establishing Echimyidae, and Capromyidae as the first major rodent families to be completely sequenced at the genus-level for their mitochondrial DNA. Combining mitogenomes and nuclear exons, we inferred a robust phylogenetic framework that reveals several newly supported nodes as well as the tempo of the higher level diversification of these rodents. Incorporating the full generic diversity of extant echimyids leads us to propose a new higher level classification of two subfamilies: Euryzygomatomyinae and Echimyinae. Of note, the enigmatic Carterodon displays fast-evolving mitochondrial and nuclear sequences, with a long branch that destabilizes the deepest divergences of the echimyid tree, thereby challenging the sister-group relationship between Capromyidae and Euryzygomatomyinae. Biogeographical analyses involving higher level taxa show that several vicariant and dispersal events impacted the evolutionary history of echimyids. The diversification history of Echimyidae seems to have been influenced by two major historical factors, namely (1) recurrent connections between Atlantic and Amazonian Forests and (2) the Northern uplift of the Andes., (© The Author 2016. Published by Oxford University Press on behalf of the Society for Molecular Biology and Evolution. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.)

Background: Branch lengths are an important attribute of phylogenetic trees, providing essential information for many studies in evolutionary biology. Yet, part of the current methodology to reconstruct a phylogeny from genomic information - namely supertree methods - focuses on the topology or structure of the phylogenetic tree, rather than the evolutionary divergences associated to it. Moreover, accurate methods to estimate branch lengths - typically based on probabilistic analysis of a concatenated alignment - are limited by large demands in memory and computing time, and may become impractical when the data sets are too large., Results: Here, we present a novel phylogenomic distance-based method, named ERaBLE (Evolutionary Rates and Branch Length Estimation), to estimate the branch lengths of a given reference topology, and the relative evolutionary rates of the genes employed in the analysis. ERaBLE uses as input data a potentially very large collection of distance matrices, where each matrix is obtained from a different genomic region - either directly from its sequence alignment, or indirectly from a gene tree inferred from the alignment. Our experiments show that ERaBLE is very fast and fairly accurate when compared to other possible approaches for the same tasks. Specifically, it efficiently and accurately deals with large data sets, such as the OrthoMaM v8 database, composed of 6,953 exons from up to 40 mammals., Conclusions: ERaBLE may be used as a complement to supertree methods - or it may provide an efficient alternative to maximum likelihood analysis of concatenated alignments - to estimate branch lengths from phylogenomic data sets.

Mitogenomic data are increasingly used in evolutionary biology and ecology, stressing the importance for double checking the authenticity of DNA sequences. For example, Szcześniak et al. (2013) recently published the mitochondrial genome of a bat, the Leschenault's rousette (Rousettus leschenaultii). Here we show using straightforward phylogenetic analyses of available chiropteran sequence data that the taxonomic attribution of the reported mitogenome is erroneous. The purportedly-new complete mitochondrial genome likely belongs to the Egyptian fruit bat (R. aegyptiacus) for which a reference sequence already exists. We propose that future articles reporting complete mitochondrial genome sequences should mandatorily include maximum likelihood trees inferred from (i) the standard barcoding marker for the taxon under focus, which would benefit from the massive data available in public databases, and (ii) the available mitogenomes of closely related species. We also strongly advise these trees be presented as phylograms so that all pertinent phylogenetic information is displayed in the form of a topology and its associated branch lengths. Along with compulsory information on the geographical location and origin of the specimen, these new standards should help avoiding the publication of taxonomically misidentified mitogenomes that might end up as reference sequences in public databases and re-used in subsequent meta-analyses.

Natural selection is expected to cause convergence of life histories among taxa as well as correlated evolution of different life-history traits. Here, we quantify the extent of convergence of five key life-history traits (adult fire survival, seed storage, degree of sexual dimorphism, pollination mode, and seed-dispersal mode) and test hypotheses about their correlated evolution in the genus Leucadendron (Proteaceae) from the fire-prone South African fynbos. We reconstructed a new molecular phylogeny of this highly diverse genus that involves more taxa and molecular markers than previously. This reconstruction identifies new clades that were not detected by previous molecular study and morphological classifications. Using this new phylogeny and robust methods that account for phylogenetic uncertainty, we show that the five life-history traits studied were labile during the evolutionary history of the genus. This diversity allowed us to tackle major questions about the correlated evolution of life-history strategies. We found that species with longer seed-dispersal distances tended to evolve lower pollen-dispersal distance, that insect-pollinated species evolved decreased sexual dimorphism, and that species with a persistent soil seed-bank evolved toward reduced fire-survival ability of adults., (© 2014 The Author(s). Evolution © 2014 The Society for the Study of Evolution.)

The Capromyidae (hutias) are endemic rodents of the Caribbean and represent a model of dispersal for non-flying mammals in the Greater Antilles. This family has experienced severe extinctions during the Holocene and its phylogenetic affinities with respect to other caviomorph relatives are still debated as morphological and molecular data disagree. We used target enrichment and next-generation sequencing of mitochondrial and nuclear genes to infer the phylogenetic relationships of hutias, estimate their divergence ages, and understand their mode of dispersal in the Greater Antilles.We found that Capromyidae are nested within Echimyidae (spiny rats) and should be considered a subfamily thereof. We estimated that the split between hutias and Atlantic Forest spiny rats occurred 16.5 (14.8–18.2) million years ago (Ma), which is more recent than the GAARlandia land bridge hypothesis (34–35 Ma). This would suggest that during the Early Miocene, an echimyid-like ancestor colonized the Greater Antilles from an eastern South American source population via rafting. The basal divergence of the Hispaniolan Plagiodontia provides further support for a vicariant separation between Hispaniolan and western islands (Bahamas, Cuba, Jamaica) hutias. Recent divergences among these western hutias suggest Plio-Pleistocene dispersal waves associated with glacial cycles.

Comparative genomic studies extensively rely on alignments of orthologous sequences. Yet, selecting, gathering, and aligning orthologous exons and protein-coding sequences (CDS) that are relevant for a given evolutionary analysis can be a difficult and time-consuming task. In this context, we developed OrthoMaM, a database of ORTHOlogous MAmmalian Markers describing the evolutionary dynamics of orthologous genes in mammalian genomes using a phylogenetic framework. Since its first release in 2007, OrthoMaM has regularly evolved, not only to include newly available genomes but also to incorporate up-to-date software in its analytic pipeline. This eighth release integrates the 40 complete mammalian genomes available in Ensembl v73 and provides alignments, phylogenies, evolutionary descriptor information, and functional annotations for 13,404 single-copy orthologous CDS and 6,953 long exons. The graphical interface allows to easily explore OrthoMaM to identify markers with specific characteristics (e.g., taxa availability, alignment size, %G+C, evolutionary rate, chromosome location). It hence provides an efficient solution to sample preprocessed markers adapted to user-specific needs. OrthoMaM has proven to be a valuable resource for researchers interested in mammalian phylogenomics, evolutionary genomics, and has served as a source of benchmark empirical data sets in several methodological studies. OrthoMaM is available for browsing, query and complete or filtered downloads at http://www.orthomam.univ-montp2.fr/., (© The Author 2014. Published by Oxford University Press on behalf of the Society for Molecular Biology and Evolution. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.)

Ascidians are a fascinating group of filter-feeding marine chordates characterized by rapid evolution of both sequences and structure of their nuclear and mitochondrial genomes. Moreover, they include several model organisms used to investigate complex biological processes in chordates. To study the evolutionary dynamics of ascidians at short phylogenetic distances, we sequenced 13 new mitogenomes and analyzed them, together with 15 other available mitogenomes, using a novel approach involving detailed whole-mitogenome comparisons of conspecific and congeneric pairs. The evolutionary rate was quite homogeneous at both intraspecific and congeneric level, and the lowest congeneric rates were found in cryptic (morphologically undistinguishable) and in morphologically very similar species pairs. Moreover, congeneric nonsynonymous rates (dN) were up to two orders of magnitude higher than in intraspecies pairs. Overall, a clear-cut gap sets apart conspecific from congeneric pairs. These evolutionary peculiarities allowed easily identifying an extraordinary intraspecific variability in the model ascidian Botryllus schlosseri, where most pairs show a dN value between that observed at intraspecies and congeneric level, yet consistently lower than that of the Ciona intestinalis cryptic species pair. These data suggest ongoing speciation events producing genetically distinct B. schlosseri entities. Remarkably, these ongoing speciation events were undetectable by the cox1 barcode fragment, demonstrating that, at low phylogenetic distances, the whole mitogenome has a higher resolving power than cox1. Our study shows that whole-mitogenome comparative analyses, performed on a suitable sample of congeneric and intraspecies pairs, may allow detecting not only cryptic species but also ongoing speciation events.

Despite the recent advances in generating molecular data, reconstructing species-level phylogenies for non-models groups remains a challenge. The use of a number of independent genes is required to resolve phylogenetic relationships, especially for groups displaying low polymorphism. In such cases, low-copy nuclear exons and non-coding regions, such as 3' untranslated regions (3'-UTRs) or introns, constitute a potentially interesting source of nuclear DNA variation. Here, we present a methodology meant to identify new nuclear orthologous markers using both public-nucleotide databases and transcriptomic data generated for the group of interest by using next generation sequencing technology. To identify PCR primers for a non-model group, the genus Leucadendron (Proteaceae), we adopted a framework aimed at minimizing the probability of paralogy and maximizing polymorphism. We anchored when possible the right-hand primer into the 3'-UTR and the left-hand primer into the coding region. Seven new nuclear markers emerged from this search strategy, three of those included 3'-UTRs. We further compared the phylogenetic potential between our new markers and the ribosomal internal transcribed spacer region (ITS). The sequenced 3'-UTRs yielded higher polymorphism rates than the ITS region did. We did not find strong incongruences with the phylogenetic signal contained in the ITS region and the seven new designed markers but they strongly improved the phylogeny of the genus Leucadendron. Overall, this methodology is efficient in isolating orthologous loci and is valid for any non-model group given the availability of transcriptomic data., (Copyright © 2013 Elsevier Inc. All rights reserved.)

Leaf-nosed bats (Phyllostomidae) are one of the most studied groups within the order Chiroptera mainly because of their outstanding species richness and diversity in morphological and ecological traits. Rapid diversification and multiple homoplasies have made the phylogeny of the family difficult to solve using morphological characters. Molecular data have contributed to shed light on the evolutionary history of phyllostomid bats, yet several relationships remain unresolved at the intra-familial level. Complete mitochondrial genomes have proven useful to deal with this kind of situation in other groups of mammals by providing access to a large number of molecular characters. At present, there are only two mitogenomes available for phyllostomid bats hinting at the need for further exploration of the mitogenomic approach in this group. We used both standard Sanger sequencing of PCR products and next-generation sequencing (NGS) of shotgun genomic DNA to obtain new complete mitochondrial genomes from 10 species of phyllostomid bats, including representatives of major subfamilies, plus one outgroup belonging to the closely-related mormoopids. We then evaluated the contribution of mitogenomics to the resolution of the phylogeny of leaf-nosed bats and compared the results to those based on mitochondrial genes and the RAG2 and VWF nuclear makers. Our results demonstrate the advantages of the Illumina NGS approach to efficiently obtain mitogenomes of phyllostomid bats. The phylogenetic signal provided by entire mitogenomes is highly comparable to the one of a concatenation of individual mitochondrial and nuclear markers, and allows increasing both resolution and statistical support for several clades. This enhanced phylogenetic signal is the result of combining markers with heterogeneous evolutionary rates representing a large number of nucleotide sites. Our results illustrate the potential of the NGS mitogenomic approach for resolving the evolutionary history of phyllostomid bats based on a denser species sampling., (Copyright © 2013. Published by Elsevier Inc.)

Sequences of the mitochondrial cytochrome b (1140 bp) and nuclear Rag 2 (1148 bp) genes were used to assess the evolutionary history of the cosmopolitan bat genus Myotis, based on a worldwide sampling of over 88 named species plus 7 species with uncertain nomenclature. Phylogenetic reconstructions of this comprehensive taxon sampling show that most radiation of species occurred independently within each biogeographic region. Our molecular study supports an early divergence of species from the New World, where all Nearctic and Neotropical species plus a lineage from the Palaearctic constitute a monophyletic clade, sister to the remaining Old World taxa. The major Old World clade includes all remaining Eurasian taxa, most Oriental species, one Oceanian, and all Ethiopian species. Another lineage, including M. latirostris from Taiwan, appears at the base of these two major biogeographic clades and, because it bears nyctalodont molars, could be considered as a distinct genus. However, this molar configuration is also found in crown-group species, indicating that these dental characters are variable in the genus Myotis and may confound interpretation of the fossil record. Molecular datings suggest an origin of all recent Myotis in the early Miocene (about 21MYA with 95% highest posterior density interval 23-20MYA). This period was characterized by a global climatic cooling that reduced the availability of tropical habitats and favoured the development of more temperate vegetation. This sharp climatic change might have triggered the evolution of Myotis in the Northern continents, because Myotis ancestors seem to have been well adapted and successful in such temperate habitats. Ancestral area reconstructions based on the molecular phylogeny suggest that the eastern portion of the Asian continent was an important center of origin for the early diversification of all Myotis lineages, and involved relatively few subsequent transcontinental range expansions., (Copyright © 2013 Elsevier Inc. All rights reserved.)

An RNA-Seq strategy was used to obtain the complete set of protein-coding mitochondrial genes from two rodent taxa. Thanks to the next generation sequencing (NGS) 454 approach, we determined the complete mitochondrial DNA genome from Graphiurus kelleni (Mammalia: Rodentia: Gliridae) and partial mitogenome from Pedetes capensis (Pedetidae), and compared them with published rodent and outgroup mitogenomes. We finished the mitogenome sequencing by a series of amplicons using conserved PCR primers to fill the gaps corresponding to tRNA, rRNA and control regions. Phylogenetic analyses of the mitogenomes suggest a well-supported rodent phylogeny in agreement with nuclear gene trees. Pedetes groups with Anomalurus into the clade Anomaluromorpha, while Graphiurus branches within the squirrel-related clade. Moreover, Pedetes+Anomalurus branch with Castor into the mouse-related clade. Our study demonstrates the utility of NGS for obtaining new mitochondrial genomes as well as the importance of choosing adequate models of sequence evolution to infer the phylogeny of rodents., (© 2013 Elsevier B.V. All rights reserved.)

Despite the rapid increase of size in phylogenomic data sets, a number of important nodes on animal phylogeny are still unresolved. Among these, the rooting of the placental mammal tree is still a controversial issue. One difficulty lies in the pervasive phylogenetic conflicts among genes, with each one telling its own story, which may be reliable or not. Here, we identified a simple criterion, that is, the GC content, which substantially helps in determining which gene trees best reflect the species tree. We assessed the ability of 13,111 coding sequence alignments to correctly reconstruct the placental phylogeny. We found that GC-rich genes induced a higher amount of conflict among gene trees and performed worse than AT-rich genes in retrieving well-supported, consensual nodes on the placental tree. We interpret this GC effect mainly as a consequence of genome-wide variations in recombination rate. Indeed, recombination is known to drive GC-content evolution through GC-biased gene conversion and might be problematic for phylogenetic reconstruction, for instance, in an incomplete lineage sorting context. When we focused on the AT-richest fraction of the data set, the resolution level of the placental phylogeny was greatly increased, and a strong support was obtained in favor of an Afrotheria rooting, that is, Afrotheria as the sister group of all other placentals. We show that in mammals most conflicts among gene trees, which have so far hampered the resolution of the placental tree, are concentrated in the GC-rich regions of the genome. We argue that the GC content-because it is a reliable indicator of the long-term recombination rate-is an informative criterion that could help in identifying the most reliable molecular markers for species tree inference.

It is widely assumed that our mammalian ancestors, which lived in the Cretaceous era, were tiny animals that survived massive asteroid impacts in shelters and evolved into modern forms after dinosaurs went extinct, 65 Ma. The small size of most Mesozoic mammalian fossils essentially supports this view. Paleontology, however, is not conclusive regarding the ancestry of extant mammals, because Cretaceous and Paleocene fossils are not easily linked to modern lineages. Here, we use full-genome data to estimate the longevity and body mass of early placental mammals. Analyzing 36 fully sequenced mammalian genomes, we reconstruct two aspects of the ancestral genome dynamics, namely GC-content evolution and nonsynonymous over synonymous rate ratio. Linking these molecular evolutionary processes to life-history traits in modern species, we estimate that early placental mammals had a life span above 25 years and a body mass above 1 kg. This is similar to current primates, cetartiodactyls, or carnivores, but markedly different from mice or shrews, challenging the dominant view about mammalian origin and evolution. Our results imply that long-lived mammals existed in the Cretaceous era and were the most successful in evolution, opening new perspectives about the conditions for survival to the Cretaceous-Tertiary crisis.

Ascidians or sea squirts form a diverse group within chordates, which includes a few thousand members of marine sessile filter-feeding animals. Their mitochondrial genomes are characterized by particularly high evolutionary rates and rampant gene rearrangements. This extreme variability complicates standard polymerase chain reaction (PCR) based techniques for molecular characterization studies, and consequently only a few complete Ascidian mitochondrial genome sequences are available. Using the standard PCR and Sanger sequencing approach, we produced the mitochondrial genome of Ascidiella aspersa only after a great effort. In contrast, we produced five additional mitogenomes (Botrylloides aff. leachii, Halocynthia spinosa, Polycarpa mytiligera, Pyura gangelion, and Rhodosoma turcicum) with a novel strategy, consisting in sequencing the pooled total DNA samples of these five species using one Illumina HiSeq 2000 flow cell lane. Each mitogenome was efficiently assembled in a single contig using de novo transcriptome assembly, as de novo genome assembly generally performed poorly for this task. Each of the new six mitogenomes presents a different and novel gene order, showing that no syntenic block has been conserved at the ordinal level (in Stolidobranchia and in Phlebobranchia). Phylogenetic analyses support the paraphyly of both Ascidiacea and Phlebobranchia, with Thaliacea nested inside Phlebobranchia, although the deepest nodes of the Phlebobranchia-Thaliacea clade are not well resolved. The strategy described here thus provides a cost-effective approach to obtain complete mitogenomes characterized by a highly plastic gene order and a fast nucleotide/amino acid substitution rate.

Although environmental filtering has been observed to influence the biodiversity patterns of marine bacterial communities, it was restricted to the regional scale and to the species level, leaving the main drivers unknown at large biogeographic scales and higher taxonomic levels. Bacterial communities with different species compositions may nevertheless share phylogenetic lineages, and phylogenetic turnover (PT) among those communities may be surprisingly low along any biogeographic or environmental gradient. Here, we investigated the relative influence of environmental filtering and geographical distance on the PT between marine bacterial communities living more than 8000 km apart in contrasted abiotic conditions. PT was high between communities and was more structured by local environmental factors than by geographical distance, suggesting the predominance of a lineage filtering process. Strong phenotype-environment mismatches observed in the ocean may surpass high connectivity between marine microbial communities.

The analysis of extant sequences shows that molecular evolution has been heterogeneous through time and among lineages. However, for a given sequence alignment, it is often difficult to uncover what factors caused this heterogeneity. In fact, identifying and characterizing heterogeneous patterns of molecular evolution along a phylogenetic tree is very challenging, for lack of appropriate methods. Users either have to a priori define groups of branches along which they believe molecular evolution has been similar or have to allow each branch to have its own pattern of molecular evolution. The first approach assumes prior knowledge that is seldom available, and the second requires estimating an unreasonably large number of parameters. Here we propose a convenient and reliable approach where branches get clustered by their pattern of molecular evolution alone, with no need for prior knowledge about the data set under study. Model selection is achieved in a statistical framework and therefore avoids overparameterization. We rely on substitution mapping for efficiency and present two clustering approaches, depending on whether or not we expect neighbouring branches to share more similar patterns of sequence evolution than distant branches. We validate our method on simulations and test it on four previously published data sets. We find that our method correctly groups branches sharing similar equilibrium GC contents in a data set of ribosomal RNAs and recovers expected footprints of selection through dN/dS. Importantly, it also uncovers a new pattern of relaxed selection in a phylogeny of Mantellid frogs, which we are able to correlate to life-history traits. This shows that our programs should be very useful to study patterns of molecular evolution and reveal new correlations between sequence and species evolution. Our programs can run on DNA, RNA, codon, or amino acid sequences with a large set of possible models of substitutions and are available at http://biopp.univ-montp2.fr/forge/testnh.

Background: Development of phylogenetic methods that do not rely on fossils for the study of evolutionary processes through time have revolutionized the field of evolutionary biology and resulted in an unprecedented expansion of our knowledge about the tree of life. These methods have helped to shed light on the macroevolution of many taxonomic groups such as the placentals (Mammalia). However, despite the increase of studies addressing the diversification patterns of organisms, no synthesis has addressed the case of the most diversified mammalian clade: the Rodentia., Results: Here we present a rodent maximum likelihood phylogeny inferred from a molecular supermatrix. It is based on 11 mitochondrial and nuclear genes that covers 1,265 species, i.e., respectively 56% and 81% of the known specific and generic rodent diversity. The inferred topology recovered all Rodentia clades proposed by recent molecular works. A relaxed molecular clock dating approach provided a time framework for speciation events. We found that the Myomorpha clade shows a greater degree of variation in diversification rates than Sciuroidea, Caviomorpha, Castorimorpha and Anomaluromorpha. We identified a number of shifts in diversification rates within the major clades: two in Castorimorpha, three in Ctenohystrica, 6 within the squirrel-related clade and 24 in the Myomorpha clade. The majority of these shifts occurred within the most recent familial rodent radiations: the Cricetidae and Muridae clades. Using the topological imbalances and the time line we discuss the potential role of different diversification factors that might have shaped the rodents radiation., Conclusions: The present glimpse on the diversification pattern of rodents can be used for further comparative meta-analyses. Muroid lineages have a greater degree of variation in their diversification rates than any other rodent group. Different topological signatures suggest distinct diversification processes among rodent lineages. In particular, Muroidea and Sciuroidea display widespread distribution and have undergone evolutionary and adaptive radiation on most of the continents. Our results show that rodents experienced shifts in diversification rate regularly through the Tertiary, but at different periods for each clade. A comparison between the rodent fossil record and our results suggest that extinction led to the loss of diversification signal for most of the Paleogene nodes.

Fairy armadillos or pichiciegos (Xenarthra, Dasypodidae) are among the most elusive mammals. Due to their subterranean and nocturnal lifestyle, their basic biology and evolutionary history remain virtually unknown. Two distinct species with allopatric distributions are recognized: Chlamyphorus truncatus is restricted to central Argentina, while Calyptophractus retusus occurs in the Gran Chaco of Argentina, Paraguay, and Bolivia. To test their monophyly and resolve their phylogenetic affinities within armadillos, we obtained sequence data from modern and museum specimens for two mitochondrial genes (12S RNA [MT-RNR1] and NADH dehydrogenase 1 [MT-ND1]) and two nuclear exons (breast cancer 1 early onset exon 11 [BRCA1] and von Willebrand factor exon 28 [VWF]). Phylogenetic analyses provided a reference phylogeny and timescale for living xenarthran genera. Our results reveal monophyletic pichiciegos as members of a major armadillo subfamily (Chlamyphorinae). Their strictly fossorial lifestyle probably evolved as a response to the Oligocene aridification that occurred in South America after their divergence from Tolypeutinae around 32 million years ago (Mya). The ancient divergence date (∼17Mya) for separation between the two species supports their taxonomic classification into distinct genera. The synchronicity with Middle Miocene marine incursions along the Paraná river basin suggests a vicariant origin for pichiciegos by the disruption of their ancestral range. Their phylogenetic distinctiveness and rarity in the wild argue in favor of high conservation priority., (Copyright © 2011 Elsevier Inc. All rights reserved.)

Genes and genomes do not evolve similarly in all branches of the tree of life. Detecting and characterizing the heterogeneity in time, and between lineages, of the nucleotide (or amino acid) substitution process is an important goal of current molecular evolutionary research. This task is typically achieved through the use of non-homogeneous models of sequence evolution, which being highly parametrized and computationally-demanding are not appropriate for large-scale analyses. Here we investigate an alternative methodological option based on probabilistic substitution mapping. The idea is to first reconstruct the substitutional history of each site of an alignment under a homogeneous model of sequence evolution, then to characterize variations in the substitution process across lineages based on substitution counts. Using simulated and published datasets, we demonstrate that probabilistic substitution mapping is robust in that it typically provides accurate reconstruction of sequence ancestry even when the true process is heterogeneous, but a homogeneous model is adopted. Consequently, we show that the new approach is essentially as efficient as and extremely faster than (up to 25 000 times) existing methods, thus paving the way for a systematic survey of substitution process heterogeneity across genes and lineages.

The Mediterranean Sea is a highly diverse, highly studied, and highly impacted biogeographic region, yet no phylogenetic reconstruction of fish diversity in this area has been published to date. Here, we infer the timing and geographic origins of Mediterranean teleost species diversity using nucleotide sequences collected from GenBank. We assembled a DNA supermatrix composed of four mitochondrial genes (12S ribosomal DNA, 16S ribosomal DNA, cytochrome c oxidase subunit I and cytochrome b) and two nuclear genes (rhodopsin and recombination activating gene I), including 62% of Mediterranean teleost species plus 9 outgroups. Maximum likelihood and Bayesian phylogenetic and dating analyses were calibrated using 20 fossil constraints. An additional 124 species were grafted onto the chronogram according to their taxonomic affinity, checking for the effects of taxonomic coverage in subsequent diversification analyses. We then interpreted the time-line of teleost diversification in light of Mediterranean historical biogeography, distinguishing non-endemic natives, endemics and exotic species. Results show that the major Mediterranean orders are of Cretaceous origin, specifically ~100-80 Mya, and most Perciformes families originated 80-50 Mya. Two important clade origin events were detected. The first at 100-80 Mya, affected native and exotic species, and reflects a global diversification period at a time when the Mediterranean Sea did not yet exist. The second occurred during the last 50 Mya, and is noticeable among endemic and native species, but not among exotic species. This period corresponds to isolation of the Mediterranean from Indo-Pacific waters before the Messinian salinity crisis. The Mediterranean fish fauna illustrates well the assembly of regional faunas through origination and immigration, where dispersal and isolation have shaped the emergence of a biodiversity hotspot.

Background: Introgressive events (e.g., hybridization, gene flow, horizontal gene transfer) and incomplete lineage sorting of ancestral polymorphisms are a challenge for phylogenetic analyses since different genes may exhibit conflicting genealogical histories. Grasses of the Triticeae tribe provide a particularly striking example of incongruence among gene trees. Previous phylogenies, mostly inferred with one gene, are in conflict for several taxon positions. Therefore, obtaining a resolved picture of relationships among genera and species of this tribe has been a challenging task. Here, we obtain the most comprehensive molecular dataset to date in Triticeae, including one chloroplastic and 26 nuclear genes. We aim to test whether it is possible to infer phylogenetic relationships in the face of (potentially) large-scale introgressive events and/or incomplete lineage sorting; to identify parts of the evolutionary history that have not evolved in a tree-like manner; and to decipher the biological causes of gene-tree conflicts in this tribe., Results: We obtain resolved phylogenetic hypotheses using the supermatrix and Bayesian Concordance Factors (BCF) approaches despite numerous incongruences among gene trees. These phylogenies suggest the existence of 4-5 major clades within Triticeae, with Psathyrostachys and Hordeum being the deepest genera. In addition, we construct a multigenic network that highlights parts of the Triticeae history that have not evolved in a tree-like manner. Dasypyrum, Heteranthelium and genera of clade V, grouping Secale, Taeniatherum, Triticum and Aegilops, have evolved in a reticulated manner. Their relationships are thus better represented by the multigenic network than by the supermatrix or BCF trees. Noteworthy, we demonstrate that gene-tree incongruences increase with genetic distance and are greater in telomeric than centromeric genes. Together, our results suggest that recombination is the main factor decoupling gene trees from multigenic trees., Conclusions: Our study is the first to propose a comprehensive, multigenic phylogeny of Triticeae. It clarifies several aspects of the relationships among genera and species of this tribe, and pinpoints biological groups with likely reticulate evolution. Importantly, this study extends previous results obtained in Drosophila by demonstrating that recombination can exacerbate gene-tree conflicts in phylogenetic reconstructions.

The Mediterranean Sea (0.82% of the global oceanic surface) holds 4%-18% of all known marine species (~17,000), with a high proportion of endemism [1, 2]. This exceptional biodiversity is under severe threats [1] but benefits from a system of 100 marine protected areas (MPAs). Surprisingly, the spatial congruence of fish biodiversity hot spots with this MPA system and the areas of high fishing pressure has not been assessed. Moreover, evolutionary and functional breadth of species assemblages [3] has been largely overlooked in marine systems. Here we adopted a multifaceted approach to biodiversity by considering the species richness of total, endemic, and threatened coastal fish assemblages as well as their functional and phylogenetic diversity. We show that these fish biodiversity components are spatially mismatched. The MPA system covers a small surface of the Mediterranean (0.4%) and is spatially congruent with the hot spots of all taxonomic components of fish diversity. However, it misses hot spots of functional and phylogenetic diversity. In addition, hot spots of endemic species richness and phylogenetic diversity are spatially congruent with hot spots of fishery impact. Our results highlight that future conservation strategies and assessment efficiency of current reserve systems will need to be revisited after deconstructing the different components of biodiversity., (Copyright © 2011 Elsevier Ltd. All rights reserved.)

Background: Recent advances in comparative genomics have considerably improved our knowledge of the evolution of mammalian karyotype architecture. One of the breakthroughs was the preferential localization of evolutionary breakpoints in regions enriched in repetitive sequences (segmental duplications, telomeres and centromeres). In this context, we investigated the contribution of ribosomal genes to genome reshuffling since they are generally located in pericentromeric or subtelomeric regions, and form repeat clusters on different chromosomes. The target model was the genus Mus which exhibits a high rate of karyotypic change, a large fraction of which involves centromeres., Results: The chromosomal distribution of rDNA clusters was determined by in situ hybridization of mouse probes in 19 species. Using a molecular-based reference tree, the phylogenetic distribution of clusters within the genus was reconstructed, and the temporal association between rDNA clusters, breakpoints and centromeres was tested by maximum likelihood analyses. Our results highlighted the following features of rDNA cluster dynamics in the genus Mus: i) rDNA clusters showed extensive diversity in number between species and an almost exclusive pericentromeric location, ii) a strong association between rDNA sites and centromeres was retrieved which may be related to their shared constraint of concerted evolution, iii) 24% of the observed breakpoints mapped near an rDNA cluster, and iv) a substantial rate of rDNA cluster change (insertion, deletion) also occurred in the absence of chromosomal rearrangements., Conclusions: This study on the dynamics of rDNA clusters within the genus Mus has revealed a strong evolutionary relationship between rDNA clusters and centromeres. Both of these genomic structures coincide with breakpoints in the genus Mus, suggesting that the accumulation of a large number of repeats in the centromeric region may contribute to the high level of chromosome repatterning observed in this group. However, the elevated rate of rDNA change observed in the chromosomally invariant clade indicates that the presence of these sequences is insufficient to lead to genome instability. In agreement with recent studies, these results suggest that additional factors such as modifications of the epigenetic state of DNA may be required to trigger evolutionary plasticity.

Until now the most efficient solution to align nucleotide sequences containing open reading frames was to use indirect procedures that align amino acid translation before reporting the inferred gap positions at the codon level. There are two important pitfalls with this approach. Firstly, any premature stop codon impedes using such a strategy. Secondly, each sequence is translated with the same reading frame from beginning to end, so that the presence of a single additional nucleotide leads to both aberrant translation and alignment.We present an algorithm that has the same space and time complexity as the classical Needleman-Wunsch algorithm while accommodating sequencing errors and other biological deviations from the coding frame. The resulting pairwise coding sequence alignment method was extended to a multiple sequence alignment (MSA) algorithm implemented in a program called MACSE (Multiple Alignment of Coding SEquences accounting for frameshifts and stop codons). MACSE is the first automatic solution to align protein-coding gene datasets containing non-functional sequences (pseudogenes) without disrupting the underlying codon structure. It has also proved useful in detecting undocumented frameshifts in public database sequences and in aligning next-generation sequencing reads/contigs against a reference coding sequence.MACSE is distributed as an open-source java file executable with freely available source code and can be used via a web interface at: http://mbb.univ-montp2.fr/macse.

The origin, evolution, and functional relevance of genomic variations in GC content are a long-debated topic, especially in mammals. Most of the existing literature, however, has focused on a small number of model species and/or limited sequence data sets. We analyzed more than 1000 orthologous genes in 33 fully sequenced mammalian genomes, reconstructed their ancestral isochore organization in the maximum likelihood framework, and explored the evolution of third-codon position GC content in representatives of 16 orders and 27 families. We showed that the previously reported erosion of GC-rich isochores is not a general trend. Several species (e.g., shrew, microbat, tenrec, rabbit) have independently undergone a marked increase in GC content, with a widening gap between the GC-poorest and GC-richest classes of genes. The intensively studied apes and (especially) murids do not reflect the general placental pattern. We correlated GC-content evolution with species life-history traits and cytology. Significant effects of body mass and genome size were detected, with each being consistent with the GC-biased gene conversion model.

Phylogenomics has recently revealed that tunicates represent the sister-group of vertebrates in the newly defined clade Olfactores. However, phylogenomic and comparative genomic studies have also suggested that tunicates are characterized by an elevated rate of molecular evolution and a high degree of genomic divergence. Despite the recurrent interest in the group, the picture of tunicate peculiar evolutionary dynamics is still fragmentary, as it mainly lies in studies focusing on only a few model species. In order to expand the available genomic data for the group, we used the high-throughput 454 technology to sequence the partial transcriptome of a previously unsampled tunicate, Microcosmus squamiger. This allowed us to get further insights into tunicate-accelerated evolution through a comparative analysis based on pertinent phylogenetic markers, i.e., a core of 35 housekeeping genes conserved across bilaterians. Our results showed that tunicates evolved on average about two times faster than the other chordates, yet the degree of this acceleration varied extensively upon genes and upon lineages. Appendicularia and Aplousobranchia were detected as the most divergent groups which were also characterized by highly heterogeneous substitution rates across genes. Finally, an estimation of the d (N)/d (S) ratio in three pairs of closely related taxa within Olfactores did not reveal strong differences between the tunicate and vertebrate lineages suggesting that for this set of housekeeping genes, the accelerated evolution of tunicates is plausibly due to an elevated mutation rate rather than to particular selective effects.

Motivation: Phylogenetic tree-building methods use molecular data to represent the evolutionary history of genes and taxa. A recurrent problem is to reconcile the various phylogenies built from different genomic sequences into a single one. This task is generally conducted by a two-step approach whereby a binary representation of the initial trees is first inferred and then a maximum parsimony (MP) analysis is performed on it. This binary representation uses a decomposition of all source trees that is usually based on clades, but that can also be based on triplets or quartets. The relative performances of these representations have been discussed but are difficult to assess since both are limited to relatively small datasets., Results: This article focuses on the triplet-based representation of source trees. We first recall how, using this representation, the parsimony analysis is related to the median tree notion. We then introduce SuperTriplets, a new algorithm that is specially designed to optimize this alternative formulation of the MP criterion. The method avoids several practical limitations of the triplet-based binary matrix representation, making it useful to deal with large datasets. When the correct resolution of every triplet appears more often than the incorrect ones in source trees, SuperTriplets warrants to reconstruct the correct phylogeny. Both simulations and a case study on mammalian phylogenomics confirm the advantages of this approach. In both cases, SuperTriplets tends to propose less resolved but more reliable supertrees than those inferred using M(atrix) Representation with Parsimony., Availability: Online and JAVA standalone versions of SuperTriplets are available at http://www.supertriplets.univ-montp2.fr/.

Here, we present a new primate phylogeny inferred from molecular supermatrix analyses of size 42 kb containing 70% of missing data, and representing 75% of primate species diversity. The supermatrix was analysed using a gene-partitioned maximum likelihood approach to obtain an exhaustive molecular phylogenetic framework. All clades recovered from recent molecular works were upheld in our analyses demonstrating that the presence of missing data did not bias our supermatrix inference. The resulting phylogenetic tree was subsequently dated with a molecular dating method to provide a timescale for speciation events. Results obtained from our relaxed molecular clock analyses concurred with previous works based on the same fossil constraints. The resulting dated tree allowed to infer of macroevolutionary processes among the primates. Shifts in diversification rate and speciation rates were determined using the SymmeTREE method and a birthdeath process. No significant asymmetry was detected for the primate clade, but significant shifts in diversification rate were identified for seven clades: Anthropoidea, Lemuriformes, Lemuridae, Galagidae, Callithrix genus, the Cercopithecinae and Asian Macaca. Comparisons with previous primate supertree results reveal that (i) there was a diversification event at the root of the Lemuriformes, (ii) a higher diversification rate is detected for Cercopithecidae and Anthropoidea and (iii) a shift in diversification is always recovered for Macaca genus. Macroevolutionary inferences and primate divergence dates show that major primate diversification events occurred after the Paleogene, suggesting the extinction of ancient primate lineages.

Background: Tunicates represent a key metazoan group as the sister-group of vertebrates within chordates. The six complete mitochondrial genomes available so far for tunicates have revealed distinctive features. Extensive gene rearrangements and particularly high evolutionary rates have been evidenced with regard to other chordates. This peculiar evolutionary dynamics has hampered the reconstruction of tunicate phylogenetic relationships within chordates based on mitogenomic data., Results: In order to further understand the atypical evolutionary dynamics of the mitochondrial genome of tunicates, we determined the complete sequence of the solitary ascidian Herdmania momus. This genome from a stolidobranch ascidian presents the typical tunicate gene content with 13 protein-coding genes, 2 rRNAs and 24 tRNAs which are all encoded on the same strand. However, it also presents a novel gene arrangement, highlighting the extreme plasticity of gene order observed in tunicate mitochondrial genomes. Probabilistic phylogenetic inferences were conducted on the concatenation of the 13 mitochondrial protein-coding genes from representatives of major metazoan phyla. We show that whereas standard homogeneous amino acid models support an artefactual sister position of tunicates relative to all other bilaterians, the CAT and CAT+BP site- and time-heterogeneous mixture models place tunicates as the sister-group of vertebrates within monophyletic chordates. Moreover, the reference phylogeny indicates that tunicate mitochondrial genomes have experienced a drastic acceleration in their evolutionary rate that equally affects protein-coding and ribosomal-RNA genes., Conclusion: This is the first mitogenomic study supporting the new chordate phylogeny revealed by recent phylogenomic analyses. It illustrates the beneficial effects of an increased taxon sampling coupled with the use of more realistic amino acid substitution models for the reconstruction of animal phylogeny.

Background: Tunicates have been recently revealed to be the closest living relatives of vertebrates. Yet, with more than 2500 described species, details of their evolutionary history are still obscure. From a molecular point of view, tunicate phylogenetic relationships have been mostly studied based on analyses of 18S rRNA sequences, which indicate several major clades at odds with the traditional class-level arrangements. Nonetheless, substantial uncertainty remains about the phylogenetic relationships and taxonomic status of key groups such as the Aplousobranchia, Appendicularia, and Thaliacea., Results: Thirty new complete 18S rRNA sequences were acquired from previously unsampled tunicate species, with special focus on groups presenting high evolutionary rate. The updated 18S rRNA dataset has been aligned with respect to the constraint on homology imposed by the rRNA secondary structure. A probabilistic framework of phylogenetic reconstruction was adopted to accommodate the particular evolutionary dynamics of this ribosomal marker. Detailed Bayesian analyses were conducted under the non-parametric CAT mixture model accounting for site-specific heterogeneity of the evolutionary process, and under RNA-specific doublet models accommodating the occurrence of compensatory substitutions in stem regions. Our results support the division of tunicates into three major clades: 1) Phlebobranchia + Thaliacea + Aplousobranchia, 2) Appendicularia, and 3) Stolidobranchia, but the position of Appendicularia could not be firmly resolved. Our study additionally reveals that most Aplousobranchia evolve at extremely high rates involving changes in secondary structure of their 18S rRNA, with the exception of the family Clavelinidae, which appears to be slowly evolving. This extreme rate heterogeneity precluded resolving with certainty the exact phylogenetic placement of Aplousobranchia. Finally, the best fitting secondary-structure and CAT-mixture models suggest a sister-group relationship between Salpida and Pyrosomatida within Thaliacea., Conclusion: An updated phylogenetic framework for tunicates is provided based on phylogenetic analyses using the most realistic evolutionary models currently available for ribosomal molecules and an unprecedented taxonomic sampling. Detailed analyses of the 18S rRNA gene allowed a clear definition of the major tunicate groups and revealed contrasting evolutionary dynamics among major lineages. The resolving power of this gene nevertheless appears limited within the clades composed of Phlebobranchia + Thaliacea + Aplousobranchia and Pyuridae + Styelidae, which were delineated as spots of low resolution. These limitations underline the need to develop new nuclear markers in order to further resolve the phylogeny of this keystone group in chordate evolution.

Natural populations of Trypanosoma cruzi are structured into five genetic lineages, T. cruzi I and T. cruzi II a to e, as the result of clonal evolution with rare genetic recombination events. To explore more in depth these phenomenons, a multigene sequencing approach was used, for the first time in the case of T. cruzi. Three nuclear loci and a maxicircle locus were sequenced on 18 T. cruzi stocks. Sequences were used to build phylogenetic trees from each locus and from concatenated sequences of all loci. The data confirmed the hybrid origin of DTUs IId and IIe, as the result of an ancient genetic recombination between strains pertaining to IIb and IIc. The data confirmed also a hybrid origin of DTUs IIa and IIc. Contrary to previous reports, we failed to detect mosaic genes. The phylogenetic relationship between DTUs and the respective roles of recombination and selection were tested.

Programmed cell death (PCD) is a mechanism implicated in many physiological and pathological processes. Until recently, apoptosis (self-killing) was the most largely studied mechanism of PCD but a growing number of laboratories are now interested in autophagy (self-eating). In the past few years data showing a tight link between both pathways has accumulated. Until now our laboratory used Ciona intestinalis, a chordate model in which in vivo experiments are possible, to study apoptosis. Recently, we showed that autophagy also occurs in the development of Ciona intestinalis and that the specific markers of both types of death are found in the same tissues and/or in the same cells. These results drove us to postulate that Ciona intestinalis can be a good model to study the link between apoptosis and autophagy. In this article, we conducted an in silico study of autophagy genes. We explored the genomes of Ciona intestinalis, of the second ascidian Ciona savignyi, and those of the classical biological models (Saccharomyces cerevisiae, Drosophila melanogaster, Caenorhabditis elegans and Homo sapiens) to extract and compare autophagy gene sequences. This genomic study was completed by an analysis of: (i) mRNA profile expression during development and (ii) the localization of Beclin protein by immunofluorescent staining in the Ciona intestinalis larvae. Taken together, the results allowed us to conclude that a complex autophagic machinery is present in Ciona intestinalis. Actually, the number of autophagy genes in Ciona intestinalis is comparable to the number of autophagy genes in human.

Background: Supertree methods combine phylogenies with overlapping sets of taxa into a larger one. Topological conflicts frequently arise among source trees for methodological or biological reasons, such as long branch attraction, lateral gene transfers, gene duplication/loss or deep gene coalescence. When topological conflicts occur among source trees, liberal methods infer supertrees containing the most frequent alternative, while veto methods infer supertrees not contradicting any source tree, i.e. discard all conflicting resolutions. When the source trees host a significant number of topological conflicts or have a small taxon overlap, supertree methods of both kinds can propose poorly resolved, hence uninformative, supertrees., Results: To overcome this problem, we propose to infer non-plenary supertrees, i.e. supertrees that do not necessarily contain all the taxa present in the source trees, discarding those whose position greatly differs among source trees or for which insufficient information is provided. We detail a variant of the PhySIC veto method called PhySIC_IST that can infer non-plenary supertrees. PhySIC_IST aims at inferring supertrees that satisfy the same appealing theoretical properties as with PhySIC, while being as informative as possible under this constraint. The informativeness of a supertree is estimated using a variation of the CIC (Cladistic Information Content) criterion, that takes into account both the presence of multifurcations and the absence of some taxa. Additionally, we propose a statistical preprocessing step called STC (Source Trees Correction) to correct the source trees prior to the supertree inference. STC is a liberal step that removes the parts of each source tree that significantly conflict with other source trees. Combining STC with a veto method allows an explicit trade-off between veto and liberal approaches, tuned by a single parameter.Performing large-scale simulations, we observe that STC+PhySIC_IST infers much more informative supertrees than PhySIC, while preserving low type I error compared to the well-known MRP method. Two biological case studies on animals confirm that the STC preprocess successfully detects anomalies in the source trees while STC+PhySIC_IST provides well-resolved supertrees agreeing with current knowledge in systematics., Conclusion: The paper introduces and tests two new methodologies, PhySIC_IST and STC, that demonstrate the interest in inferring non-plenary supertrees as well as preprocessing the source trees. An implementation of the methods is available at: http://www.atgc-montpellier.fr/physic_ist/.

It is widely accepted that evolutionary changes in conserved developmental signaling pathways play an important role in morphological evolution. However, few in silico studies were interested in tracking such changes in a signaling pathway. The Ectodysplasin (EDA) pathway provides an opportunity to fill this gap because it is involved in vertebrate skin appendage development such as scales, teeth, hair, and feathers that take an obvious part in the adaptation of species to their environment. We benefited from the large amount of genomic data now available to explore the evolution of the upstream genes of the EDA pathway. In mammals, these genes are eda (encoding 2 ligands, EDA-A1 and EDA-A2), edar (EDA-A1 receptor), edaradd (EDA receptor [EDAR] adapter), xedar (EDA-A2 receptor), and troy (a XEDAR-related receptor). We show that the evolution of EDA pathway genes combines both strongly conserved features and evolutionary shifts. These shifts are found at different signaling levels (from the ligand to intracellular signaling) and at different taxonomic levels (class, suborder, and genera). Although conserved features likely participate to the similarities found in the early development of vertebrate skin appendages, these shifts might account for innovations and specializations. Moreover, our study demonstrates that we can now benefit from the large number of sequenced vertebrate genomes to explore the evolution of specific signaling pathways and thereby to open new perspectives for developmental biology and evolutionary developmental biology.

This study constitutes the first evaluation and application of quantitative taxonomy to the family Caulimoviridae and the first in-depth phylogenetic study of the family Caulimoviridae that integrates the common origin between LTR retrotransposons and caulimoviruses. The phylogenetic trees and PASC analyses derived from the full genome and from the corresponding partial RT concurred, providing strong support for the current genus classification based mainly on genome organisation and use of partial RT sequence as a molecular marker. The PASC distributions obtained are multimodal, making it possible to distinguish between genus, species and strain. The taxonomy of badnaviruses infecting banana (Musa spp.) was clarified, and the consequence of endogenous badnaviruses on the genetic diversity and evolution of caulimoviruses is discussed. The use of LTR retrotransposons as outgroups reveals a structured bipolar topology separating the genus Badnavirus from the other genera. Badnaviruses appear to be the most recent genus, with the genus Tungrovirus in an intermediary position. This structuring intersects the one established by genomic and biological properties and allows us to make a correlation between phylogeny and biogeography. The variability shown between members of the family Caulimoviridae is in a similar range to that reported within other DNA and RNA plant virus families.

Background: Molecular sequence data have become the standard in modern day phylogenetics. In particular, several long-standing questions of mammalian evolutionary history have been recently resolved thanks to the use of molecular characters. Yet, most studies have focused on only a handful of standard markers. The availability of an ever increasing number of whole genome sequences is a golden mine for modern systematics. Genomic data now provide the opportunity to select new markers that are potentially relevant for further resolving branches of the mammalian phylogenetic tree at various taxonomic levels., Description: The EnsEMBL database was used to determine a set of orthologous genes from 12 available complete mammalian genomes. As targets for possible amplification and sequencing in additional taxa, more than 3,000 exons of length > 400 bp have been selected, among which 118, 368, 608, and 674 are respectively retrieved for 12, 11, 10, and 9 species. A bioinformatic pipeline has been developed to provide evolutionary descriptors for these candidate markers in order to assess their potential phylogenetic utility. The resulting OrthoMaM (Orthologous Mammalian Markers) database can be queried and alignments can be downloaded through a dedicated web interface http://kimura.univ-montp2.fr/orthomam., Conclusion: The importance of marker choice in phylogenetic studies has long been stressed. Our database centered on complete genome information now makes possible to select promising markers to a given phylogenetic question or a systematic framework by querying a number of evolutionary descriptors. The usefulness of the database is illustrated with two biological examples. First, two potentially useful markers were identified for rodent systematics based on relevant evolutionary parameters and sequenced in additional species. Second, a complete, gapless 94 kb supermatrix of 118 orthologous exons was assembled for 12 mammals. Phylogenetic analyses using probabilistic methods unambiguously supported the new placental phylogeny by retrieving the monophyly of Glires, Euarchontoglires, Laurasiatheria, and Boreoeutheria. Muroid rodents thus do not represent a basal placental lineage as it was mistakenly reasserted in some recent phylogenomic analyses based on fewer taxa. We expect the OrthoMaM database to be useful for further resolving the phylogenetic tree of placental mammals and for better understanding the evolutionary dynamics of their genomes, i.e., the forces that shaped coding sequences in terms of selective constraints.

Armadillos, anteaters, and sloths (Order Xenarthra) comprise 1 of the 4 major clades of placental mammals. Isolated in South America from the other continental landmasses, xenarthrans diverged over a period of about 65 Myr, leaving more than 200 extinct genera and only 31 living species. The presence of both ancestral and highly derived anatomical features has made morphoanatomical analyses of the xenarthran evolutionary history difficult, and previous molecular analyses failed to resolve the relationships within armadillo subfamilies. We investigated the presence/absence patterns of retroposons from approximately 7,400 genomic loci, identifying 35 phylogenetically informative elements and an additional 39 informative rare genomic changes (RGCs). DAS-short interspersed elements (SINEs), previously described only in the Dasypus novemcinctus genome, were found in all living armadillo genera, including the previously unsampled Chlamyphorus, but were noticeably absent in sloths. The presence/absence patterns of the phylogenetically informative retroposed elements and other RGCs were then compared with data from the DNA sequences of the more than 12-kb flanking regions of these retroposons. Together, these data provide the first fully resolved genus tree of xenarthrans. Interestingly, multiple evidence supports the grouping of Chaetophractus and Zaedyus as a sister group to Euphractus within Euphractinae, an association that was not previously demonstrated. Also, flanking sequence analyses favor a close phylogenetic relationship between Cabassous and Tolypeutes within Tolypeutinae. Finally, the phylogenetic position of the subfamily Chlamyphorinae is resolved by the noncoding sequence data set as the sister group of Tolypeutinae. The data provide a stable phylogenetic framework for further evolutionary investigations of xenarthrans and important information for defining conservation priorities to save the diversity of one of the most curious groups of mammals.

This paper focuses on veto supertree methods; i.e., methods that aim at producing a conservative synthesis of the relationships agreed upon by all source trees. We propose desirable properties that a supertree should satisfy in this framework, namely the non-contradiction property (PC) and the induction property (PI). The former requires that the supertree does not contain relationships that contradict one or a combination of the source topologies, whereas the latter requires that all topological information contained in the supertree is present in a source tree or collectively induced by several source trees. We provide simple examples to illustrate their relevance and that allow a comparison with previously advocated properties. We show that these properties can be checked in polynomial time for any given rooted supertree. Moreover, we introduce the PhySIC method (PHYlogenetic Signal with Induction and non-Contradiction). For k input trees spanning a set of n taxa, this method produces a supertree that satisfies the above-mentioned properties in O(kn(3) + n(4)) computing time. The polytomies of the produced supertree are also tagged by labels indicating areas of conflict as well as those with insufficient overlap. As a whole, PhySIC enables the user to quickly summarize consensual information of a set of trees and localize groups of taxa for which the data require consolidation. Lastly, we illustrate the behaviour of PhySIC on primate data sets of various sizes, and propose a supertree covering 95% of all primate extant genera. The PhySIC algorithm is available at http://atgc.lirmm.fr/cgi-bin/PhySIC.

American trypanosomiasis or Chagas disease is endemic in Latin America and caused by the flagellate Trypanosoma cruzi, which exhibits broad genetic variation. In various areas, the transmission of Chagas disease is ensured by sylvatic vectors, mainly carrying the evolutionary lineage I of T. cruzi. Despite its epidemiological importance, this lineage is poorly studied. Here, we investigated the genetic variability and the phylogenetic relationships within T. cruzi I using sequences of the non-transcribed spacer of miniexon genes. The variability was firstly analysed between 10 repeats of spacer-miniexon genes in two strains of T. cruzi I and in the CL Brener strain, showing lower intra-strain variability than inter-strain. Furthermore, the phylogenetic analysis of 19 T. cruzi I strains (49 copies in total) clusters the copies into at least three groups. Two evolutionary phenomena can be proposed to explain the partition of the strains: (i) an association between strains and Didelphis sp. hosts and (ii) geographical clustering between the North American and South American strains. Thereby, the miniexon gene is an attractive marker to establish the phylogeny of lineage I and explore relationships between T. cruzi and mammal hosts.

It has recently been reported that the synaptic acetylcholinesterase (AChE) in mosquitoes is encoded by the ace-1 gene, distinct and divergent from the ace-2 gene, which performs this function in Drosophila. This is an unprecedented situation within the Diptera order because both ace genes derive from an old duplication and are present in most insects and arthropods. Nevertheless, Drosophila possesses only the ace-2 gene. Thus, a secondary loss occurred during the evolution of Diptera, implying a vital function switch from one gene (ace-1) to the other (ace-2). We sampled 78 species, representing 50 families (27% of the Dipteran families) spread over all major subdivisions of the Diptera, and looked for ace-1 and ace-2 by systematic PCR screening to determine which taxonomic groups within the Diptera have this gene change. We show that this loss probably extends to all true flies (or Cyclorrhapha), a large monophyletic group of the Diptera. We also show that ace-2 plays a non-detectable role in the synaptic AChE in a lower Diptera species, suggesting that it has non-synaptic functions. A relative molecular evolution rate test showed that the intensity of purifying selection on ace-2 sequences is constant across the Diptera, irrespective of the presence or absence of ace-1, confirming the evolutionary importance of non-synaptic functions for this gene. We discuss the evolutionary scenarios for the takeover of ace-2 and the loss of ace-1, taking into account our limited knowledge of non-synaptic functions of ace genes and some specific adaptations of true flies.

Background: Mitochondrial and nuclear genes have generally been employed for different purposes in molecular systematics, the former to resolve relationships within recently evolved groups and the latter to investigate phylogenies at a deeper level. In the case of rapid and recent evolutionary radiations, mitochondrial genes like cytochrome b (CYB) are often inefficient for resolving phylogenetic relationships. One of the best examples is illustrated by Arvicolinae rodents (Rodentia; Muridae), the most impressive mammalian radiation of the Northern Hemisphere which produced voles, lemmings and muskrats. Here, we compare the relative contribution of a nuclear marker--the exon 10 of the growth hormone receptor (GHR) gene--to the one of the mitochondrial CYB for inferring phylogenetic relationships among the major lineages of arvicoline rodents., Results: The analysis of GHR sequences improves the overall resolution of the Arvicolinae phylogeny. Our results show that the Caucasian long-clawed vole (Prometheomys schaposnikowi) is one of the basalmost arvicolines, and confirm that true lemmings (Lemmus) and collared lemmings (Dicrostonyx) are not closely related as suggested by morphology. Red-backed voles (Myodini) are found as the sister-group of a clade encompassing water vole (Arvicola), snow vole (Chionomys), and meadow voles (Microtus and allies). Within the latter, no support is recovered for the generic recognition of Blanfordimys, Lasiopodomys, Neodon, and Phaiomys as suggested by morphology. Comparisons of parameter estimates for branch lengths, base composition, among sites rate heterogeneity, and GTR relative substitution rates indicate that CYB sequences consistently exhibit more heterogeneity among codon positions than GHR. By analyzing the contribution of each codon position to node resolution, we show that the apparent higher efficiency of GHR is due to their third positions. Although we focus on speciation events spanning the last 10 million years (Myr), CYB sequences display highly saturated codon positions contrary to the nuclear exon. Lastly, variable length bootstrap predicts a significant increase in resolution of arvicoline phylogeny through the sequencing of nuclear data in an order of magnitude three to five times greater than the size of GHR exon 10., Conclusion: Our survey provides a first resolved gene tree for Arvicolinae. The comparison of CYB and GHR phylogenetic efficiency supports recent assertions that nuclear genes are useful for resolving relationships of recently evolved animals. The superiority of nuclear exons may reside both in (i) less heterogeneity among sites, and (ii) the presence of highly informative sites in third codon positions, that evolve rapidly enough to accumulate synapomorphies, but slow enough to avoid substitutional saturation.

Phylogenomic studies aim to build phylogenies from large sets of homologous genes. Such "genome-sized" data require fast methods, because of the typically large numbers of taxa examined. In this framework, distance-based methods are useful for exploratory studies and building a starting tree to be refined by a more powerful maximum likelihood (ML) approach. However, estimating evolutionary distances directly from concatenated genes gives poor topological signal as genes evolve at different rates. We propose a novel method, named super distance matrix (SDM), which follows the same line as average consensus supertree (ACS; Lapointe and Cucumel, 1997) and combines the evolutionary distances obtained from each gene into a single distance supermatrix to be analyzed using a standard distance-based algorithm. SDM deforms the source matrices, without modifying their topological message, to bring them as close as possible to each other; these deformed matrices are then averaged to obtain the distance supermatrix. We show that this problem is equivalent to the minimization of a least-squares criterion subject to linear constraints. This problem has a unique solution which is obtained by resolving a linear system. As this system is sparse, its practical resolution requires O(naka) time, where n is the number of taxa, k the number of matrices, and a < 2, which allows the distance supermatrix to be quickly obtained. Several uses of SDM are proposed, from fast exploratory studies to more accurate approaches requiring heavier computing time. Using simulations, we show that SDM is a relevant alternative to the standard matrix representation with parsimony (MRP) method, notably when the taxa sets of the different genes have low overlap. We also show that SDM can be used to build an excellent starting tree for an ML approach, which both reduces the computing time and increases the topogical accuracy. We use SDM to analyze the data set of Gatesy et al. (2002, Syst. Biol. 51: 652-664) that involves 48 genes of 75 placental mammals. The results indicate that these genes have strong rate heterogeneity and confirm the simulation conclusions.

The comparison of DNA and protein sequences of extant species might be informative for reconstructing the chronology of evolutionary events on Earth. A phylogenetic tree inferred from molecular data directly depicts the evolutionary affinities of species and indirectly allows estimating the age of their origin and diversification. Molecular dating is achieved by assuming the molecular clock hypothesis, i.e., that the rate of change of nucleotide and amino acid sequences is on average constant over geological time. If paleontological calibrations are available, then absolute divergence times of species can be estimated. However, three major difficulties potentially hamper molecular dating : (1) a limited sample of genes and organisms, (2) a limited number of fossil references, and (3) pervasive variations of molecular evolutionary rates among genomes and species. To circumvent these problems, different solutions have been recently proposed. Larger data sets are built with more genes and more species sampled through the mining of an increasing number of genomes. Moreover, independent key fossils are identified to calibrate molecular clocks, and the uncertainty on their age is integrated in subsequent analyses. Finally, models of molecular rate variations are constructed, and incorporated in the so-called relaxed molecular clock approaches. As an illustration of these improvements, we mention that the debated age of the animal (bilaterian metazoans) diversification may have occurred between 642-761 million years ago (Mya), roughly 100 Ma before the Cambrian explosion. Among mammals, the initial diversification of major placental groups may have taken place around 100 Mya, well before the Cretaceous/Tertiary boundary marking the extinction of dinosaurs.

Platyrrhine primates and caviomorph rodents are clades of mammals that colonized South America during its period of isolation from the other continents, between 100 and 3 million years ago (Mya). Until now, no molecular study investigated the timing of the South American colonization by these two lineages with the same molecular data set. Using sequences from three nuclear genes (ADRA2B, vWF, and IRBP, both separate and combined) from 60 species, and eight fossil calibration constraints, we estimated the times of origin and diversification of platyrrhines and caviomorphs via a Bayesian relaxed molecular clock approach. To account for the possible effect of an accelerated rate of evolution of the IRBP gene along the branch leading to the anthropoids, we performed the datings with and without IRBP (3768 sites and 2469 sites, respectively). The time window for the colonization of South America by primates and by rodents is demarcated by the dates of origin (upper bound) and radiation (lower bound) of platyrrhines and caviomorphs. According to this approach, platyrrhine primates colonized South America between 37.0 +/- 3.0 Mya (or 38.9 +/- 4.0 Mya without IRBP) and 16.8 +/- 2.3 (or 20.1 +/- 3.3) Mya, and caviomorph rodents between 45.4 +/- 4.1 (or 43.7 +/- 4.8) Mya and 36.7 +/- 3.7 (or 35.8 +/- 4.3) Mya. Considering both the fossil record and these molecular datings, the favored scenarios are a trans-Atlantic migration of primates from Africa at the end of the Eocene or beginning of the Oligocene, and a colonization of South America by rodents during the Middle or Late Eocene. Based on our nuclear DNA data, we cannot rule out the possibility of a concomitant arrival of primates and rodents in South America. The caviomorphs radiated soon after their arrival, before the Oligocene glaciations, and these early caviomorph lineages persisted until the present. By contrast, few platyrrhine fossils are known in the Oligocene, and the present-day taxa are the result of a quite recent, Early Miocene diversification.

Phylogenetic relationships of 19 species of didelphid marsupials were studied using two nuclear markers, the non-coding transthyretin intron 1 (TTR) and the coding interphotoreceptor retinoid binding protein exon 1 (IRBP), and two mitochondrial genes, the protein-coding cytochrome b (cyt-b) and the structural 12S ribosomal DNA (12S rDNA). Evolutionary dynamics of these four markers were compared to each other, revealing the appropriate properties presented by TTR intron 1 together with its well supported and resolved phylogenetic signal. Nuclear markers supported the monophyly of medium and large-sized opossums Metachirus+(Chironectes, Lutreolina, Didelphis, Philander), and the paraphyly of mouse-sized opossums, with the genera Gracilinanus, Thylamys, and Marmosops as a sister group to medium and large-sized didelphids. Conflicting branching patterns between mitochondrial and nuclear data involved the phylogenetic position of Marmosa-Micoureus-Monodelphis relative to other mouse-sized opossums. Nuclear phylogenetic inferences among genera were confirmed by the presence of synapomorphic indels observed in TTR intron 1. A Bayesian relaxed molecular clock dating of didelphid evolution using nuclear markers estimated their origin in the Middle Eocene (39.8 million years ago), with subsequent diversification during the Oligocene (Deseadan) and Miocene.

Although cassava (Manihot esculenta ssp. esculenta) is asexually propagated, farmers incorporate plants from seedlings into planting stocks. These products of sex are exposed to selection, which in agricultural environments should favour rapid growth. To examine whether seedling morphology has evolved under domestication, we compared domesticated cassava, its wild progenitor (M. esculenta ssp. flabellifolia) and their sister species (M. pruinosa) under controlled conditions. Field observations complemented laboratory study. In both wild taxa, the hypocotyl did not elongate (hypogeal germination) and cotyledons remained enclosed in the testa. In domesticated cassava, the hypocotyl elongated (epigeal germination), and cotyledons emerged and became foliaceous. The difference in hypocotyl elongation was fixed, whereas cotyledon morphology varied with environmental conditions in M. pruinosa. Comparative analysis suggests that epigeal germination is primitive in Manihot, that the lineage including wild ancestors of cassava evolved hypogeal germination--which confers greater tolerance to risks in their savanna environment--and that with domestication, there was a reversion to epigeal germination and photosynthetic cotyledons, traits conferring high initial growth rates in agricultural habitats., (Copyright New Phytologist (2005).)

The phylogeny of South American spiny rats (Rodentia; Echimyidae) was studied using the exon 28 of the von Willebrand Factor nuclear gene (vWF). Sequences were analysed separately and in combination with a mitochondrial dataset (cyt b, 12S and 16S rRNAs) used in previous publications. The basal polytomy of echimyids was partially resolved and unexpected intergeneric clades were recovered. Thus, the intimate nested position of Myocastor within echimyids is evidenced. A well-supported clade is identified, including all the arboreal genera, and a group formed by Myocastor, Thrichomys, and Proechimys+Hoplomys. The clustering of Euryzygomatomys+Clyomys with Trinomys is also suggested. On the opposite, the phylogenetic position of Capromys as well as the relationships among arboreal genera remain unclear. Molecular divergence times were estimated using a Bayesian relaxed molecular clock and suggest a Middle Miocene origin for most of modern genera. The ecomorphological diversification of echimyids is discussed in the light of these new results and past environmental modifications in South America.

Rodentia (e.g., mice, rats, dormice, squirrels, and guinea pigs) and Lagomorpha (e.g., rabbits, hares, and pikas) are usually grouped into the Glires. Status of this controversial superorder has been evaluated using morphology, paleontology, and mitochondrial plus nuclear DNA sequences. This growing corpus of data has been favoring the monophyly of Glires. Recently, Misawa and Janke [Mol. Phylogenet. Evol. 28 (2003) 320] analyzed the 6441 amino acids of 20 nuclear proteins for six placental mammals (rat, mouse, rabbit, human, cattle, and dog) and two outgroups (chicken and xenopus), and observed a basal position of the two murine rodents among the former. They concluded that "the Glires hypothesis was rejected." We here reanalyzed [loc. cit.] data set under maximum likelihood and Bayesian tree-building approaches, using phylogenetic models that take into account among-site variation in evolutionary rates and branch-length variation among proteins. Our observations support both the association of rodents and lagomorphs and the monophyly of Euarchontoglires (=Supraprimates) as the most likely explanation of the protein alignments. We conducted simulation studies to evaluate the appropriateness of lissamphibian and avian outgroups to root the placental tree. When the outgroup-to-ingroup evolutionary distance increases, maximum parsimony roots the topology along the long Mus-Rattus branch. Maximum likelihood, in contrast, roots the topology along different branches as a function of their length. Maximum likelihood appears less sensitive to the "long-branch attraction artifact" than is parsimony. Our phylogenetic conclusions were confirmed by the analysis of a different protein data set using a similar sample of species but different outgroups. We also tested the effect of the addition of afrotherian and xenarthran taxa. Using the linearized tree method, [loc. cit.] estimated that mice and rats diverged about 35 million years ago. Molecular dating based on the Bayesian relaxed molecular clock method suggests that the 95% credibility interval for the split between mice and rats is 7-17 Mya. We here emphasize the need for appropriate models of sequence evolution (matrices of amino acid replacement, taking into account among-site rate variation, and independent parameters across independent protein partitions) and for a taxonomically broad sample, and conclude on the likelihood that rodents and lagomorphs together constitute a monophyletic group (Glires).

The use of nucleotide and amino acid sequences allows improved understanding of the timing of evolutionary events of life on earth. Molecular estimates of divergence times are, however, controversial and are generally much more ancient than suggested by the fossil record. The limited number of genes and species explored and pervasive variations in evolutionary rates are the most likely sources of such discrepancies. Here we compared concatenated amino acid sequences of 129 proteins from 36 eukaryotes to determine the divergence times of several major clades, including animals, fungi, plants, and various protists. Due to significant variations in their evolutionary rates, and to handle the uncertainty of the fossil record, we used a Bayesian relaxed molecular clock simultaneously calibrated by six paleontological constraints. We show that, according to 95% credibility intervals, the eukaryotic kingdoms diversified 950-1,259 million years ago (Mya), animals diverged from choanoflagellates 761-957 Mya, and the debated age of the split between protostomes and deuterostomes occurred 642-761 Mya. The divergence times appeared to be robust with respect to prior assumptions and paleontological calibrations. Interestingly, these relaxed clock time estimates are much more recent than those obtained under the assumption of a global molecular clock, yet bilaterian diversification appears to be approximately 100 million years more ancient than the Cambrian boundary.