Phylogenomic data analyses provide evidence that Xenarthra and Afrotheria are sister groups

The phylogenetic positions of the 4 clades, Euarchontoglires, Laurasiatheria, Afrotheria, and Xenarthra, have been major issues in the recent discussion of basal relationships among placental mammals. However, despite considerable efforts these relationships, crucial to the understanding of eutherian evolution and biogeography, have remained essentially unresolved. Euarchontoglires and Laurasiatheria are generally joined into a common clade (Boreoeutheria), whereas the position of Afrotheria and Xenarthra relative to Boreoeutheria has been equivocal in spite of the use of comprehensive amounts of nuclear encoded sequences or the application of genome-level characters such as retroposons. The probable reason for this uncertainty is that the divergences took place long time ago and within a narrow temporal window, leaving only short common branches. With the aim of further examining basal eutherian relationships, we have collected conserved protein-coding sequences from 11 placental mammals, a marsupial and a bird, whose nuclear genomes have been largely sequenced. The length of the alignment of homologous sequences representing each individual species is 2,168,859 nt. This number of sites, representing 2840 protein-coding genes, exceeds by a considerable margin that of any previous study. The phylogenetic analysis joined Xenarthra and Afrotheria on a common branch, Atlantogenata. This topology was found to fit the data significantly better than the alternative trees.     

A new phylogenetic marker,apolipoprotein B,provides compelling evidence for eutherian relationships

Higher-level relationships within, and the root of Placentalia, remain contentious issues. Resolution of the placental tree is important to the choice of mammalian genome projects and model organisms, as well as for understanding the biogeography of the eutherian radiation. We present phylogenetic analyses of 63 species representing all extant eutherian mammal orders for a new molecular phylogenetic marker, a 1.3kb portion of exon 26 of the apolipoprotein B (APOB) gene. In addition, we analyzed a multigene concatenation that included APOB sequences and a previously published data set (Murphy et al., 2001b) of three mitochondrial and 19 nuclear genes, resulting in an alignment of over 17kb for 42 placentals and two marsupials. Due to computational difficulties, previous maximum likelihood analyses of large, multigene concatenations for placental mammals have used quartet puzzling, less complex models of sequence evolution, or phylogenetic constraints to approximate a full maximum likelihood bootstrap. Here, we utilize a Unix load sharing facility to perform maximum likelihood bootstrap analyses for both the APOB and concatenated data sets with a GTR+Gamma+I model of sequence evolution, tree-bisection and reconnection branch-swapping, and no phylogenetic constraints. Maximum likelihood and Bayesian analyses of both data sets provide support for the superordinal clades Boreoeutheria, Euarchontoglires, Laurasiatheria, Xenarthra, Afrotheria, and Ostentoria (pangolins+carnivores), as well as for the monophyly of the orders Eulipotyphla, Primates, and Rodentia, all of which have recently been questioned. Both data sets recovered an association of Hippopotamidae and Cetacea within Cetartiodactyla, as well as hedgehog and shrew within Eulipotyphla. APOB showed strong support for an association of tarsier and Anthropoidea within Primates. Parsimony, maximum likelihood and Bayesian analyses with both data sets placed Afrotheria at the base of the placental radiation. Statistical tests that employed APOB to examine a priori hypotheses for the root of the placental tree rejected rooting on myomorphs and hedgehog, but did not discriminate between rooting at the base of Afrotheria, at the base of Xenarthra, or between Atlantogenata (Xenarthra+Afrotheria) and Boreoeutheria. An orthologous deletion of 363bp in the aligned APOB sequences proved phylogenetically informative for the grouping of the order Carnivora with the order Pholidota into the superordinal clade Ostentoria. A smaller deletion of 237-246bp was diagnostic of the superordinal clade Afrotheria.     

Molecular phylogeny of living xenarthrans and the impact of character and taxon sampling on the placental tree rooting

Extant xenarthrans (armadillos, anteaters and sloths) are among the most derived placental mammals ever evolved. South America was the cradle of their evolutionary history. During the Tertiary, xenarthrans experienced an extraordinary radiation, whereas South America remained isolated from other continents. The 13 living genera are relics of this earlier diversification and represent one of the four major clades of placental mammals. Sequences of the three independent protein-coding nuclear markers alpha2B adrenergic receptor (ADRA2B), breast cancer susceptibility (BRCA1), and von Willebrand Factor (VWF) were determined for 12 of the 13 living xenarthran genera. Comparative evolutionary dynamics of these nuclear exons using a likelihood framework revealed contrasting patterns of molecular evolution. All codon positions of BRCA1 were shown to evolve in a strikingly similar manner, and third codon positions appeared less saturated within placentals than those of ADRA2B and VWF. Maximum likelihood and Bayesian phylogenetic analyses of a 47 placental taxa data set rooted by three marsupial outgroups resolved the phylogeny of Xenarthra with some evidence for two radiation events in armadillos and provided a strongly supported picture of placental interordinal relationships. This topology was fully compatible with recent studies, dividing placentals into the Southern Hemisphere clades Afrotheria and Xenarthra and a monophyletic Northern Hemisphere clade (Boreoeutheria) composed of Laurasiatheria and Euarchontoglires. Partitioned likelihood statistical tests of the position of the root, under different character partition schemes, identified three almost equally likely hypotheses for early placental divergences: a basal Afrotheria, an Afrotheria + Xenarthra clade, or a basal Xenarthra (Epitheria hypothesis). We took advantage of the extensive sampling realized within Xenarthra to assess its impact on the location of the root on the placental tree. By resampling taxa within Xenarthra, the conservative Shimodaira-Hasegawa likelihood-based test of alternative topologies was shown to be sensitive to both character and taxon sampling.     

The chromosomes of afrotheria and their bearing on Mammalian genome evolution

Afrotheria is the clade of placental mammals that, together with Xenarthra, Euarchontoglires and Laurasiatheria, represents 1 of the 4 main recognized supraordinal eutherian clades. It reunites 6 orders of African origin: Proboscidea, Sirenia, Hyracoidea, Macroscelidea, Afrosoricida and Tubulidentata. The apparently unlikely relationship among such disparate morphological taxa and their possible basal position at the base of the eutherian phylogenetic tree led to a great deal of attention and research on the group. The use of biomolecular data was pivotal in Afrotheria studies, as they were the basis for the recognition of this clade. Although morphological evidence is still scarce, a plethora of molecular data firmly attests to the phylogenetic relationship among these mammals of African origin. Modern cytogenetic techniques also gave a significant contribution to the study of Afrotheria, revealing chromosome signatures for the group as a whole, as well as for some of its internal relationships. The associations of human chromosomes HSA1/19 and 5/21 were found to be chromosome signatures for the group and provided further support for Afrotheria. Additional chromosome synapomorphies were also identified linking elephants and manatees in Tethytheria (the associations HSA2/3, 3/13, 8/22, 18/19 and the lack of HSA4/8) and elephant shrews with the aardvark (HSA2/8, 3/20 and 10/17). Herein, we review the current knowledge on Afrotheria chromosomes and genome evolution. The already available data on the group suggests that further work on this apparently bizarre assemblage of mammals will provide important data to a better understanding on mammalian genome evolution.     

Indels in protein-coding sequences of Euarchontoglires constrain the rooting of the eutherian tree

Despite the availability of large molecular data sets, the position of the root of the eutherian tree remains a controversial issue. Depending on source data, taxon sampling and analytical approach, the root can be placed at either Afrotheria, Xenarthra, Afrotheria+Xenarthra, or murid rodents. We explored the phylogenetic potential of indels in four nuclear protein-coding genes (SCA1, PRNP, TNFalpha, and HspB3) with regard to a possible rooting at the murid branch. According to parsimony principles, five indels were interpreted to contradict such a rooting, and one indel to support it. The results illustrate that indels, despite the occurrence of homoplasy, can be convincing sources of independent molecular evidence to distinguish between alternative phylogenetic hypotheses.     

Mitogenomic relationships of placental mammals and molecular estimates of their divergences

Molecular analyses of the relationships of placental mammals have shown a progressive congruence between mitogenomic and nuclear phylogenies. Some inconsistencies have nevertheless persisted, notably with respect to basal divergences. The current study has aimed to extend the representation of groups, whose position in the placental tree has been difficult to establish in mitogenomic studies. Both ML (maximum likelihood) and Bayesian analyses identified four basal monophyletic groups, Afroplacentalia (=Afrotheria: Hyracoidea, Proboscidea, Sirenia, Tenrecidea, Tubulidentata, Macroscelidea, Chrysochloridea), Xenarthra, Archontoglires (Primates, Dermoptera, Scandentia, Lagomorpha, Rodentia) and Laurasiaplacentalia (Lipotyphla, Chiroptera, Pholidota, Carnivora, Perissodactyla, Artiodactyla, Cetacea). All analyses joined Archontoglires and Laurasiaplacentalia on a common branch (Boreoplacentalia), but the relationship between Afroplacentalia, Xenarthra and Boreoplacentalia was not conclusively resolved. The phylogenomic hypothesis with a sister group relationship between Notoplacentalia (Afroplacentalia/Xenarthra) and Boreoplacentalia served as the basis for estimating the times of placental divergences using paleontologically well-supported mammalian calibration points. These estimates placed the basal placental divergence between Boreoplacentalia and Notoplacentalia at approximately 102 MYA (million years ago). The current estimates of ordinal placental divergences are congruent with recent estimates based on nuclear data, but inconsistent with paleontological notions that have placed the origin of essentially all placental orders within an interval of 5-10 MY in the early Tertiary. Among less deep divergences the estimates placed the split between Gorilla and Pan/Homo at approximately 11.5 MYA and that between Pan and Homo at approximately 8 MYA. As a consequence of these estimates, which are in accord with recent progress in primate paleontology, the earliest divergences among recent humans become placed approximately 270,000 years ago, i.e. approximately 100,000 years earlier than the traditional age of "Mitochondrial Eve". Comparison between the two new mt genomes of Hylomys suillus (short-tailed gymnure) patently demonstrates the inconsistency that may exist between taxonomic designations and molecular difference, as the distance between these two supposedly conspecific genomes exceeds that of the three elephantid genera Elephas, Mammuthus and Loxodonta. In accordance with the progressive use of the term Placentalia for extant orders and extinct taxa falling within this group we forward new proposals for the names of some superordinal clades of placental mammals.     

A molecular perspective on mammalian evolution from the gene encoding interphotoreceptor retinoid binding protein, with convincing evidence for bat monophyly.

The evolutionary relationships of the various orders of placental mammals remain an issue of uncertainty and controversy. Molecular studies of mammalian phylogeny at the DNA level that include more than just a few orders are still relatively meager. Here we report results on mammalian phylogeny deduced from the coding sequence of the single-copy nuclear gene for the interphotoreceptor retinoid binding protein (IRBP). Analysis of 13 species representing eight eutherian orders and one marsupial yielded results that falsify the hypothesis that megachiropteran bats are "flying primates," only convergently resembling microchiropteran bats. Instead, in agreement with more traditional views, as well as those from other recent molecular studies, the results strongly support a monophyletic Chiroptera (micro- and megabats grouped together). The IRBP results also offer some rare molecular support for the Glires concept, in which rodents and lagomorphs form a superordinal grouping. Also in congruence with other recent molecular evidence, IRBP sequences do not support the view of a superorder Archonta that includes Chiroptera along with Dermoptera (flying lemur), Scandentia (tree shrew), and Primates. IRBP was not however, without its shortcomings as a molecular phylogenetic system: high levels of homoplasy, evident in the marsupial outgroup, did not allow us to properly root the tree, and several of the higher level eutherian clades were only weakly supported (e.g., a Carnivora/Chiroptera clade and an Artiodactyla/Carnivora/Chiroptera clade). We suggest that these shortcomings may be diminished as the phylogenetic density of the data set is increased.     

Using novel phylogenetic methods to evaluate mammalian mtDNA,including amino acid-invariant sites-LogDet plus site stripping,to detect internal conflicts in the data,with special reference to the positions of hedgehog,armadillo, and elephant

We look at the higher-order phylogeny of mammals, analyzing in detail the complete mtDNA sequences of more than 40 species. We test the support for several proposed superordinal relationships. To this end, we apply a number of recently programmed methods and approaches, plus better-established methods. New pairwise tests show highly significant evidence that amino acid frequencies are changing among nearly all the genomes studied when unvaried sites are ignored. LogDet amino acid distances, with modifications to take into account invariant sites, are combined with bootstrapping and the Neighbor Joining algorithm to account for these violations of standard models. To weight the more slowly evolving sites, we exclude the more rapidly evolving sites from the data by using "site stripping". This leads to changing optimal trees with nearly all methods. The bootstrap support for many hypotheses varies widely between methods, and few hypotheses can claim unanimous support from these data. Rather, we uncover good evidence that many of the earlier branching patterns in the placental subtree could be incorrect, including the placement of the root. The tRNA genes, for example, favor a split between the group hedgehog, rodents, and primates versus all other sequenced placentals. Such a grouping is not ruled out by the amino acid sequence data. A grouping of all rodents plus rabbit, the old Glires hypothesis, is also feasible with stripped amino acid data, and rodent monophyly is also common. The elephant sequence allows confident rejection of the older taxon Ferungulata (Simpson, 1945). In its place, the new taxa Scrotifera and Fereuungulata are defined. A new likelihood ratio test is used to detect differences between the optimal tree for tRNA versus that for amino acids. While not clearly significant as made, some results indicate the test is tending towards significance with more general models of evolution. Individual placement tests suggest alternative positions for hedgehog and elephant. Congruence arguments to support elephant and armadillo together are striking, suggesting a superordinal group composed of Xenarthra and African endemic mammals, which in turn may be near the root of the placental subtree. Thus, while casting doubt on some recent conclusions, the analyses are also unveiling some interesting new possibilities.     

Chromosomal instability in Afrotheria: fragile sites,evolutionary breakpoints and phylogenetic inference from genome sequence assemblies

Background  

Extant placental mammals are divided into four major clades (Laurasiatheria, Supraprimates, Xenarthra and Afrotheria). Given that Afrotheria is generally thought to root the eutherian tree in phylogenetic analysis of large nuclear gene data sets, the study of the organization of the genomes of afrotherian species provides new insights into the dynamics of mammalian chromosomal evolution. Here we test if there are chromosomal bands with a high tendency to break and reorganize in Afrotheria, and by analyzing the expression of aphidicolin-induced common fragile sites in three afrotherian species, whether these are coincidental with recognized evolutionary breakpoints.     

Molecular Evidence for the Major Clades of Placental Mammals

Higher-level relationships among placental mammals, as well as the historical biogeography of this group against the backdrop of continental fragmentation and reassembly, remain poorly understood. Here, we analyze two independent molecular data sets that represent all placental orders. The first data set includes six genes (A2AB, IRBP, vWF, 12S rRNA, tRNA valine, 16S rRNA; total = 5.71 kb) for 26 placental taxa and two marsupials; the second data set includes 2.95 kb of exon 11 of the BRCA1 gene for 51 placental taxa and four marsupials. We also analyzed a concatenation of these data sets (8.66 kb) for 26 placentals and one marsupial. Unrooted and rooted analyses were performed with parsimony, distance methods, maximum likelihood, and a Bayesian approach. Unrooted analyses provide convincing support for a fundamental separation of placental orders into groups with southern and northern hemispheric origins according to the current fossil record. On rooted trees, one or both of these groups are monophyletic depending on the position of the root. Maximum likelihood and Bayesian analyses with the BRCA1 and combined 8.66 kb data sets provide strong support for the monophyly of the northern hemisphere group (Boreoeutheria). Boreoeutheria is divided into Laurasiatheria (Carnivora + Cetartiodactyla + Chiroptera + Eulipotyphla + Perissodactyla + Pholidota) and Euarchonta (Dermoptera + Primates + Scandentia) + Glires (Lagomorpha + Rodentia). The southern hemisphere group is either monophyletic or paraphyletic, depending on the method of analysis used. Within this group, Afrotheria (Proboscidea + Sirenia + Hyracoidea + Tubulidentata + Macroscelidea + Afrosoricida) is monophyletic. A unique nine base-pair deletion in exon 11 of the BRCA1 gene also supports Afrotheria monophyly. Given molecular dates that suggest that the southern hemisphere group and Boreoeutheria diverged in the Early Cretaceous, a single trans-hemispheric dispersal event may have been of fundamental importance in the early history of crown-group Eutheria. Parallel adaptive radiations have subsequently occurred in the four major groups: Laurasiatheria, Euarchonta + Glires, Afrotheria, and Xenarthra.     

Comparative aspects of trophoblast development and placentation

Based on the number of tissues separating maternal from fetal blood, placentas are classified as epitheliochorial, endotheliochorial or hemochorial. We review the occurrence of these placental types in the various orders of eutherian mammals within the framework of the four superorders identified by the techniques of molecular phylogenetics. The superorder Afrotheria diversified in ancient Africa and its living representatives include elephants, sea cows, hyraxes, aardvark, elephant shrews and tenrecs. Xenarthra, comprising armadillos, anteaters and sloths, diversified in South America. All placentas examined from members of these two oldest superorders are either endotheliochorial or hemochorial. The superorder Euarchontoglires includes two sister groups, Glires and Euarchonta. The former comprises rodents and lagomorphs, which typically have hemochorial placentas. The most primitive members of Euarchonta, the tree shrews, have endotheliochorial placentation. Flying lemurs and all higher primates have hemochorial placentas. However, the lemurs and lorises are exceptional among primates in having epitheliochorial placentation. Laurasiatheria, the last superorder to arise, includes several orders with epitheliochorial placentation. These comprise whales, camels, pigs, ruminants, horses and pangolins. In contrast, nearly all carnivores have endotheliochorial placentation, whilst bats have endotheliochorial or hemochorial placentas. Also included in Laurasiatheria are a number of insectivores that have many conserved morphological characters; none of these has epitheliochorial placentation. Consideration of placental type in relation to the findings of molecular phylogenetics suggests that the likely path of evolution in Afrotheria was from endotheliochorial to hemochorial placentation. This is also a likely scenario for Xenarthra and the bats. We argue that a definitive epitheliochorial placenta is a secondary specialization and that it evolved twice, once in the Laurasiatheria and once in the lemurs and lorises.     

Mammalian evolution and the interphotoreceptor retinoid binding protein (IRBP) gene: Convincing evidence for several superordinal clades

Phylogenetic relationships of 25 mammalian species representing 17 of the 18 eutherian orders were examined using DNA sequences from a 1.2-kb region of the 5′ end of exon 1 of the single-copy nuclear gene known as interphotoreceptor retinoid binding protein (IRBP). A wide variety of methods of analysis of the DNA sequence, and of the translated products, all supported a five-order clade consisting of elephant shrew (Macroscelidea)/aardvark (Tubulidentata)/and the paenungulates (hyracoids, sirenians, and elephants), with bootstrap support in all cases of 100%. The Paenungulata was also strongly supported by these IRBP data. In the majority of analyses this monophyletic five-order grouping was the first branch off the tree after the Edentata. These results are highly congruent with two other recent sources of molecular data. Another superordinal grouping, with similar 100% bootstrap support in all of the same wide-ranging types of analyses, was Artiodactyla/Cetacea. Other superordinal affinities, suggested by the analyses, but with less convincing support, included a Perissodactyla/Artiodactyla/Cetacea clade, an Insectivora/Chiroptera clade, and Glires (an association of rodents and lagomorphs). Correspondence to: M. J. Stanhope     

Interordinal relationships and timescale of eutherian evolution as inferred from mitochondrial genome data

Extensive phylogenetic analyses of the updated sequence data of mammalian mitochondrial genomes were carried out using the maximum likelihood method in order to resolve deep branchings in eutherian evolution. The divergence times in the mammalian tree were estimated by a relaxed molecular clock of the mitochondrial proteins calibrated with multiple references. A Chiroptera/Eulipotyphla (i.e. bat/mole) clade and a close relationship of this clade to Fereuungulata (Carnivora+Perissodactyla+Cetartiodactyla) were reconfirmed with high statistical significance. However, a support for a monophyly of Fereuungulata relative to the Chiroptera/Eulipotyphla clade was fragile, and we suggest that the three branchings among Carnivora, Perissodactyla, Cetartiodactyla and Chiroptera/Eulipotyphla occurred successively in a short time period, estimated to be approximately 77Myr BP. The Chiroptera/Eulipotyphla divergence was estimated to roughly coincide with the Cretaceous-Tertiary boundary (65Myr BP). The monophyly of Rodentia, the Lagomorpha/Rodentia clade (traditionally called Glires), and the Afrotheria/Xenarthra clade were preferred over alternative relationships, but the supports of these clades were not strong enough to exclude other possibilities. Although several super-order taxa of eutherians were strongly supported by the analyses of the mitochondrial genome data, the branching order in the deepest part of the eutherian tree remained ambiguous from the data presently available.     

Conflict Among Individual Mitochondrial Proteins in Resolving the Phylogeny of Eutherian Orders

The phylogenetic relationship among primates, ferungulates (artiodactyls + cetaceans + perissodactyls + carnivores), and rodents was examined using proteins encoded by the H strand of mtDNA, with marsupials and monotremes as the outgroup. Trees estimated from individual proteins were compared in detail with the tree estimated from all 12 proteins (either concatenated or summing up log-likelihood scores for each gene). Although the overall evidence strongly suggests ((primates, ferungulates), rodents), the ND1 data clearly support another tree, ((primates, rodents), ferungulates). To clarify whether this contradiction is due to (1) a stochastic (sampling) error; (2) minor model-based errors (e.g., ignoring site rate variability), or (3) convergent and parallel evolution (specifically between either primates and rodents or ferungulates and the outgroup), the ND1 genes from many additional species of primates, rodents, other eutherian orders, and the outgroup (marsupials + monotremes) were sequenced. The phylogenetic analyses were extensive and aimed to eliminate the following artifacts as possible causes of the aberrant result: base composition biases, unequal site substitution rates, or the cumulative effects of both. Neither more sophisticated evolutionary analyses nor the addition of species changed the previous conclusion. That is, the statistical support for grouping rodents and primates to the exclusion of all other taxa fluctuates upward or downward in quite a tight range centered near 95% confidence. These results and a site-by-site examination of the sequences clearly suggest that convergent or parallel evolution has occurred in ND1 between primates and rodents and/or between ferungulates and the outgroup. While the primate/rodent grouping is strange, ND1 also throws some interesting light on the relationships of some eutherian orders, marsupials, and montremes. In these parts of the tree, ND1 shows no apparent tendency for unexplained convergences. Received: 5 December 1997 / Accepted: 24 February 1998     

Four new mitochondrial genomes and the increased stability of evolutionary trees of mammals from improved taxon sampling

We have sequenced four new mitochondrial genomes to improve the stability of the tree for placental mammals; they are two insectivores (a gymnure, Echinosorex gymnurus and Formosan shrew Soriculus fumidus); a Formosan lesser horseshoe bat (Rhinolophus monoceros); and the New Zealand fur seal (Arctocephalus forsteri). A revision to the hedgehog sequence (Erinaceus europaeus) is also reported. All five are from the Laurasiatheria grouping of eutherian mammals. On this new data set there is a strong tendency for the hedgehog and its relative, the gymnure, to join with the other Laurasiatherian insectivores (mole and shrews). To quantify the stability of trees from this data we define, based on nuclear sequences, a major four-way split in Laurasiatherians. This ([Xenarthra, Afrotheria], [Laurasiatheria, Supraprimates]) split is also found from mitochondrial genomes using either protein-coding or RNA (rRNA and tRNA) data sets. The high similarity of the mitochondrial and nuclear-derived trees allows a quantitative estimate of the stability of trees from independent data sets, as detected from a triplet Markov analysis. There are significant changes in the mutational processes within placental mammals that are ignored by current tree programs. On the basis of our quantitative results, we expect the evolutionary tree for mammals to be resolved quickly, and this will allow other problems to be solved.     

Congruent mammalian trees from mitochondrial and nuclear genes using Bayesian methods

Analyses of mitochondrial and nuclear gene sequences have often produced different mammalian tree topologies, undermining confidence in the merit of molecular approaches with respect to "traditional" morphological classification. The recent sequencing of the complete mitochondrial genomes of two additional rodents (Spalax judaei and Jaculus jaculus) and one lagomorph (Ochotona princeps) has prompted us to reinvestigate the issue. Using Bayesian phylogenetics, we found phylogenetic relationships between mammalian species highly congruent with previous results based on nuclear genes. Our results show the existence of four primary lineages of placental mammals: Xenarthra, Afrotheria, Laurasiatheria, and Euarchontoglires. Relationships between and within these lineages strongly suggest that the gene trees may also be congruent with the underlying species phylogeny.     

Phylogeny of rodentia (Mammalia) inferred from the nuclear-encoded gene IRBP

The order Rodentia includes nearly half of all living mammalian species. Phylogenetic relationships among 22 species of rodents were investigated by use of a 1.2-kb region from exon 1 of the single-copy nuclear gene IRBP. IRBP has been extensively used for study of interordinal phylogeny in mammals, which allowed inclusion of 50 outgroup species, representing every eutherian order plus seven marsupials. Several clades were strongly supported, regardless of analytical method or inclusion/exclusion of data. These include a monophyletic Muroidea, with a clade including Spalax and Rhizomys as the first divergence; a clade uniting Zapus with Dipus, but excluding Sicista; a monophyletic Myodonta (Muroidea plus Dipodidae); and a clade including Aplodontidae as sister to Sciuridae. One bipartition, separating Hystricognathi and Geomyoidea from the remaining rodents, is strongly supported in all analyses that include third-position sites but almost completely absent from analyses that exclude third-position sites. A combination of nonstationary nucleotide composition and branch length effects may be causing all methods examined (including those using the LogDet distance) to support an incorrect conclusion when third-position sites are analyzed together with first- and second-position sites.     

Genomic inferences from Afrotheria and the evolution of elephants

Recent genetic studies have established that African forest and savanna elephants are distinct species with dissociated cytonuclear genomic patterns, and have identified Asian elephants from Borneo and Sumatra as conservation priorities. Representative of Afrotheria, a superordinal clade encompassing six eutherian orders, the African savanna elephant was among the first mammals chosen for whole-genome sequencing to provide a comparative understanding of the human genome. Elephants have large and complex brains and display advanced levels of social structure, communication, learning and intelligence. The elephant genome sequence might prove useful for comparative genomic studies of these advanced traits, which have appeared independently in only three mammalian orders: primates, cetaceans and proboscideans.     

The mitochondrial DNA molecule of the aardvark, Orycteropus afer, and the position of the Tubulidentata in the eutherian tree

An outstanding problem in mammal phylogeny is the relationship of the aardvark (Orycteropus afer), the only living species of the order Tubulidentata, to the extant eutherian lineages. In order to examine this problem the complete mitochondrial DNA (mtDNA) molecule of the aardvark was sequenced and analysed. The aardvark tRNA-Ser (UCN) differs from that of other mammalian mtDNAs reported and appears to have reversed to the ancestral secondary structure of non-mammalian vertebrates and mitochondrial tRNAs in general. Phylogenetic analysis of 12 concatenated protein-coding genes (3325 amino acids) included the aardvark and 15 additional eutherians, two marsupials and a monotreme. The most strongly supported tree identified the aardvark as a sister group of a clade including the armadillo (Xenarthra) and the Cetferungulata (carnivores, perissodactyls, artiodactyls and cetaceans). By applying three molecular calibration points the divergence between the aardvark and armadillo-cetferungulates was estimated at ca. 90 million years before present.