Phylogenetic relationships and divergence times among mustelids (Mammalia: Carnivora) based on nucleotide sequences of the nuclear interphotoreceptor retinoid binding protein and mitochondrial cytochrome b genes

Phylogenetic relationships among 20 species-group taxa of Mustelidae, representing Mustelinae (Mustela, Martes, Gulo), Lutrinae (Enhydra), and Melinae (Meles), were examined using nucleotide sequences of the nuclear interphotoreceptor retinoid binding protein (IRBP) and mitochondrial cytochrome b genes. Neighbor-joining and maximum-parsimony phylogenetic analyses on these genes separately and combined were conducted. While IRBP performed better than cytochrome b in recovering more-inclusive clades, cytochrome b demonstrated more resolving power in recovering less-inclusive clades. Strong support was found for a close affinity of Enhydra with Mustela to the exclusion of Martes and Gulo (causing Mustelinae to be paraphyletic); the most-basal position of Mustela vison within Mustela, followed by Mustela erminea; an association of Mustela lutreola, Mustela itatsi, Mustela sibirica, and the subgenus Putorius (including Mustela putorius and Mustela eversmanii), to the exclusion of Mustela nivalis and Mustela altaica; and a basal position of Mustela itatsi to a clade containing Mustela sibirica and Putorius. Whereas cytochrome b strongly supported Mustela lutreola as the sister species to Putorius, IRBP strongly supported its basal placement to the Mustela itatsi-Mustela sibirica-Putorius clade. The low level of sequence divergence in cytochrome b between Mustela lutreola and Putorius is therefore a result of interspecific mitochondrial introgression between these taxa, rather than a recent origin of Mustela lutreola in a close relationship to Putorius. Time estimates inferred from IRBP and cytochrome b for mustelid divergence events are mostly in agreement with the fossil record.     

New insights into the evolution of intronic sequences of the beta-fibrinogen gene and their application in reconstructing mustelid phylogeny

Mustelidae is the largest and most diverse family in the order Carnivora. The phylogenetic relationships among the subfamilies have especially long been a focus of study. Herein we are among the first to employ two new introns (4 and 7) of the nuclear beta-fibrinogen gene to clarify these enigmatic problems. In addition, two previously available nuclear (IRBP exon 1 and TTR intron 1) and one mt (ND2) data sets were also combined and analyzed simultaneously with the newly obtained sequence data in this study. Detailed characterizations of the two intronic regions not only reveal the remarkable occurrences of short interspersed element (SINE) insertion events, providing a new example supporting the attractive hypothesis that attrition of an earlier retroposition may offer a proper environment for successive retropositions by forming a "dimer-like" structure, but also demonstrate their utility in the resolution of mustelid phylogeny. All of our analyses confirm the assemblage of Mustelinae, Lutrinae, and Melinae with confidence; moreover, two clades within Mustelinae were clearly recognized, i.e., genera Mustela and Martes. Notably, genus Martes of Mustelinae was found to branch off first, followed by Melinae and then a clade containing Lutrinae and genus Mustela of Mustelinae, indicating paraphyly of Mustelinae. In addition, Mephitinae diverges before the other mustelids and the monophyletic Procyonidae in all cases, supporting its elevation to a separate family. Additional independent genetic markers are still in need to resolve the trichotomy among Mephitinae and the other two carnivoran clades, Ailuridae and Procyonidae/non-mephitine Mustelidae.     

Molecular phylogenetic study on the origin and evolution of Mustelidae

The family Mustelidae, which consists of Mustelinae, Lutrinae, Melinae, and Taxidiinae, is the largest family among Carnivora and is a highly diverse group. Recent molecular phylogenetic studies have clarified the phylogenetic relations among Mustelidae, but there remain several unresolved problems, particularly concerning the deep branchings. Whereas many studies support the monophyly of Mustelidae+Procyonidae among Musteloidea, the relations between Mustelidae+Procyonidae, Ailuridae, and Miphitidae are still unclear. To address these problems, we inferred a tree on the basis of the sequences of mitochondrial genomes and of multiple nuclear genes using the maximum likelihood method. Our results strongly support the hypothesis that the Taxidiinae branched at first, followed by the branching of the Melinae. After that, Mustelinae diversified, and Lutrinae evolved within Mustelinae. With respect to the deep branchings in Musteloidea, the Ailuridae/Mephitidae monophyly tree and the Mephitidae-basal tree are indistinguishable in log-likelihood score, and this problem remains unresolved.     

Evolutionary and biogeographic history of weasel-like carnivorans (Musteloidea)

We analyzed a concatenated (8492 bp) nuclear-mitochondrial DNA data set from 44 musteloids (including the first genetic data for Lyncodon patagonicus) with parsimony, maximum likelihood, and Bayesian methods of phylogenetic and biogeographic inference and two Bayesian methods of chronological inference. Here we show that Musteloidea emerged approximately 32.4-30.9 million years ago (MYA) in Asia, shortly after the greenhouse-icehouse global climate shift at the Eocene-Oligocene transition. During their Oligocene radiation, which proceeded wholly or mostly in Asia, musteloids diversified into four primary divisions: the Mephitidae lineage separated first, succeeded by Ailuridae and the divergence of the Procyonidae and Mustelidae lineages. Mustelidae arose approximately 16.1 MYA within the Mid-Miocene Climatic Optimum, and extensively diversified in the Miocene, mostly in Asia. The early offshoots of this radiation largely evolved into badger and marten ecological niches (Taxidiinae, Melinae, Mellivorinae, Guloninae, and Helictidinae), whereas the later divergences have adapted to other niches including those of weasels, polecats, minks, and otters (Mustelinae, Ictonychinae, and Lutrinae). Notably, and contrary to traditional beliefs, the morphological adaptations of badgers, martens, weasels, polecats, and minks each evolved independently more than once within Mustelidae. Ictonychinae (which is most closely related to Lutrinae) arose approximately 9.5-8.9 MYA, most likely in Asia, where it diverged into the Old World Ictonychini (Vormela, Poecilictis, Ictonyx, and Poecilogale) and New World Lyncodontini (Lyncodon and Galictis) lineages. Ictonychini presumably entered Africa during the Messinian Salinity Crisis (at the Miocene-Pliocene transition), which interposed the origins of this clade (approximately 6.5-6.0 MYA) and its African Poecilictis-Ictonyx-Poecilogale subclade (approximately 4.8-4.5 MYA). Lyncodontini originated approximately 2.9-2.6 MYA at the Pliocene-Pleistocene transition in South America, slightly after the emergence of the Panamanian land bridge that provided for the Great American Biotic Interchange. As the genera Martes and Ictonyx (as currently circumscribed) are paraphyletic with respect to the genera Gulo and Poecilogale, respectively, we propose that Pekaniaand Poecilictis be treated as valid genera and that "Martes"pennanti and "Ictonyx"libyca, respectively, be assigned to these genera.     

Evidence from nuclear DNA sequences sheds light on the phylogenetic relationships of Pinnipedia: single origin with affinity to Musteloidea

Considerable long-standing controversy and confusion surround the phylogenetic affinities of pinnipeds, the largely marine group of "fin-footed" members of the placental mammalian order Carnivora. Until most recently, the two major competing hypotheses were that the pinnipeds have a single (monophyletic) origin from a bear-like ancestor, or that they have a dual (diphyletic) origin, with sea lions (Otariidae) derived from a bear-like ancestor, and seals (Phocidae) derived from an otter-, mustelid-, or musteloid-like ancestor. We examined phylogenetic relationships among 29 species of arctoid carnivorans using a concatenated sequence of 3228 bp from three nuclear loci (apolipoprotein B, APOB; interphotoreceptor retinoid-binding protein, IRBP; recombination-activating gene 1, RAG1). The species represented Pinnipedia (Otariidae: Callorhinus, Eumetopias; Phocidae: Phoca), bears (Ursidae: Ursus, Melursus), and Musteloidea (Mustelidae: Mustela, Enhydra, Melogale, Martes, Gulo, Meles; Procyonidae: Procyon; Ailuridae: Ailurus; Mephitidae: Mephitis). Maximum parsimony, maximum likelihood, and Bayesian inference phylogenetic analyses of separate and combined datasets produced trees with largely congruent topologies. The analyses of the combined dataset resulted in well-resolved and well-supported phylogeny reconstructions. Evidence from nuclear DNA evolution presented here contradicts the two major hypotheses of pinniped relationships and strongly suggests a single origin of the pinnipeds from an arctoid ancestor shared with Musteloidea to the exclusion of Ursidae.     

鼬科动物线粒体DNA控制区结构分析

利用PCR技术获得紫貂(Martes zibellina)和黄喉貂(Martes flavigula)线粒体DNA控制区全序列,并结合从GenBank中下载的9种鼬科动物相应序列,用ClustalX排序后对控制区结构进行分析,识别出延长终止序列区、中央区和保守序列区3个区域,指出了一个终止相关序列ETAS1及8个保守序列(CSB-F、E、D、C、B、1、2和3),并给出了序列通式,在CSB1和CSB2之间发现不同形式的短重复序列.此外,以狼为外类群,应用邻接法构建鼬科线粒体控制区全序列的系统进化树,结果表明:臭鼬亚科最先从鼬科中分化出来,随后剩余类群分为两大支系,即貂属种类与貂熊聚为一支,并与獾亚科的狗獾形成姐妹群;另一支为水獭亚科的物种与鼬属的林鼬形成姐妹群,再与虎鼬聚在一起,狗獾与貂属的紫貂亲缘关系最近,水獭亚科与鼬属亲缘关系最近.     

Phylogeny, evolutionary history and taxonomy of the Mustelidae based on sequences of the cytochrome b gene and a complex repetitive flanking region

The Mustelidae is a diverse family of carnivores which includes weasels, polecats, mink, tayra, martens, otters, badgers and, according to some authors, skunks. Evolutionary relationships within the family are under debate at a number of different taxonomic levels, and incongruencies between molecular and morphological results are important. We analysed a total of 241 cytochrome b (cyt b) gene sequences and 33 sequences of a complex repetitive flanking region from 33 different species to compile an extensive molecular phylogeny for the Mustelidae. We analysed these sequences and constructed phylogenetic trees using Bayesian and neighbor‐joining methods that are evaluated to propose changes to the taxonomy of the family. The peripheral position of skunks in phylogenetic trees based on both loci suggests that they should be considered a separate family, Mephitidae. The subfamily Melinae is the basal group within the Mustelidae and trees based on the cyt b gene suggest that the American badger, Taxidea taxus, should be considered a separate monotypic subfamily, Taxidiinae. Otters classified within the genera Lutra, Amblonyx and Aonyx are grouped within the same clade in cyt b and combined partial cyt b and flanking region trees and show reduced levels of inter specific divergence, suggesting that they could be classified together under a single genus, Lutra. The Bayesian tree based on combined data from both loci supports the idea that subfamily Mustelinae is paraphyletic, as otters (subfamily Lutrinae) are included in this subfamily. Low levels of genetic divergence among European polecat, Mustela putorius, steppe polecat, Mustela eversmannii, and European mink, Mustela lutreola, suggest that these species could be considered subspecies within a single species, Mustela putorius. Our results are consistent with a rapid diversification of mustelid lineages in six different radiation episodes identified since the Early Eocene, the oldest events being the separation of subfamilies and the split of marten (Martes, Gulo) and weasel (Mustela) lineages in the Early Middle Miocene. The separation of New World from Old World lineages and the split of the remaining genera are estimated to have occurred in Late Miocene. The most recent events have been the differentiation of species within genera and this probably occurred in four radiation episodes at the end of Late Miocene, Early Pliocene, Late Pliocene and Pleistocene epochs.     

Phylogenetic relationships within the extant Mustelidae (Garnivora): appraisal of the cladistic status of the Simpsonian subfamilies

Cladistic analysis of extant mustelids based on a data matrix consisting of 30 characters derived from morphological study of the head skeleton in all 23 genera, and 16 characters gleaned from the literature, yielded 75 most parsimonious trees with a consistency index of 0.487 using PAUP 3.0s. The analysis strongly supports the monophyly of both the Mephitinae (skunks) and Lutrinae (otters). The Mustelinae (weasels, martens, wolverine, etc) is probably paraphyletic, but there is some support for a monophyletic group consisting of the mustelines and the honey badger Mellivora. The Melinae (badgers) is a polyphyletic group; the badger ecomorph probably evolved independently at least three times within the Mustelidae. On consensus trees, there is a lack of resolution concerning the relationships within the Mephitinae and Lutrinae and the positions of Galictis and Mustela. These results are broadly congruent with a cladistic interpretation of Simpson's intent in his influential 1945 classification. Thus, a paraphyletic Mustelinae is consistent with Simpson's conception of a horizontal grouping, whereas the Mephitinae and Lutrinae represent vertical divergent lineages. In contrast, the cladistic status of the Melinae is not consistent with Simpsonian classificatory principles. The status of the monotypic Mellivorinae cannot be evaluated without considering its putative fossil members; however, the clustering of Mellivora with the badger Taxidea is not consistent with Simpson's scheme. Some of the more inclusive clades in the most parsimonious trees are vulnerable to minor changes to the data matrix. The fossil record is consistent with the basic outlines of the cladistic pattern and explains various ostensible biogeographic anomalies. Bootstrap analysis identified marked variation in the support for the components of the cladistic pattern.     

Independent nonframeshift deletions in the MC1R gene are not associated with melanistic coat coloration in three mustelid lineages

Sequence variation within the 5' flanking (about 240 bp) and exon regions (426 bp) of the melanocortin-1 receptor (MC1R) gene was examined to determine the potential role of this protein in the melanistic coat coloration of 17 mustelid species in four genera: Gulo (wolverines), Martes (martens), Mustela (weasels), and Meles (badgers). Members of the genera Mustela and Meles, together with Martes flavigula and Martes pennanti, were shown to have intact gene sequences. However, several "in frame" deletions of the MC1R gene region implicated in melanism of other species were detected within members of the genera Martes and Gulo. For instance, Gulo gulo possessed a 15 bp deletion in the second transmembrane domain coding region, while Martes americana, Martes melampus, Martes zibellina, and Martes martes shared a 45 bp deletion overlapping this area. In addition, Martes foina was found to possess a 10 bp insertion followed closely by a 28 bp deletion immediately downstream of the deletion found in other martens. Notably, none of these indels was associated with a melanistic phenotype. Phylogenetic analysis revealed that each of these nonrandomly distributed deletions arose independently during the evolution of this family. Specific indel-neighboring motifs appear to largely account for the biased and repeated occurrence of deletion events in the Martes/Gulo clade.     

Molecular phylogeny of the carnivora (mammalia): assessing the impact of increased sampling on resolving enigmatic relationships

This study analyzed 76 species of Carnivora using a concatenated sequence of 6243 bp from six genes (nuclear TR-i-I, TBG, and IRBP; mitochondrial ND2, CYTB, and 12S rRNA), representing the most comprehensive sampling yet undertaken for reconstructing the phylogeny of this clade. Maximum parsimony and Bayesian methods were remarkably congruent in topologies observed and in nodal support measures. We recovered all of the higher level carnivoran clades that had been robustly supported in previous analyses (by analyses of morphological and molecular data), including the monophyly of Caniformia, Feliformia, Arctoidea, Pinnipedia, Musteloidea, Procyonidae + Mustelidae sensu stricto, and a clade of (Hyaenidae + (Herpestidae + Malagasy carnivorans)). All of the traditional "families," with the exception of Viverridae and Mustelidae, were robustly supported as monophyletic groups. We further have determined the relative positions of the major lineages within the Caniformia, which previous studies could not resolve, including the first robust support for the phylogenetic position of marine carnivorans (Pinnipedia) within the Arctoidea (as the sister-group to musteloids [sensu lato], with ursids as their sister group). Within the pinnipeds, Odobenidae (walrus) was more closely allied with otariids (sea lions/fur seals) than with phocids ("true" seals). In addition, we recovered a monophyletic clade of skunks and stink badgers (Mephitidae) and resolved the topology of musteloid interrelationships as: Ailurus (Mephitidae (Procyonidae, Mustelidae [sensu stricto])). This pattern of interrelationships of living caniforms suggests a novel inference that large body size may have been the primitive condition for Arctoidea, with secondary size reduction evolving later in some musteloids. Within Mustelidae, Bayesian analyses are unambiguous in supporting otter monophyly (Lutrinae), and in both MP and Bayesian analyses Martes is paraphyletic with respect to Gulo and Eira, as has been observed in some previous molecular studies. Within Feliformia, we have confirmed that Nandinia is the outgroup to all other extant feliforms, and that the Malagasy Carnivora are a monophyletic clade closely allied with the mongooses (Herpestidae [sensu stricto]). Although the monophyly of each of the three major feliform clades (Viverridae sensu stricto, Felidae, and the clade of Hyaenidae + (Herpestidae + Malagasy carnivorans)) is robust in all of our analyses, the relative phylogenetic positions of these three lineages is not resolvable at present. Our analyses document the monophyly of the "social mongooses," strengthening evidence for a single origin of eusociality within the Herpestidae. For a single caniform node, the position of pinnipeds relative to Ursidae and Musteloidea, parsimony analyses of data for the entire Carnivora did not replicate the robust support observed for both parsimony and Bayesian analyses of the caniform ingroup alone. More detailed analyses and these results demonstrate that outgroup choice can have a considerable effect on the strength of support for a particular topology. Therefore, the use of exemplar taxa as proxies for entire clades with diverse evolutionary histories should be approached with caution.The Bayesian analysis likelihood functions generally were better able to reconstruct phylogenetic relationships (increased resolution and more robust support for various nodes) than parsimony analyses when incompletely sampled taxa were included. Bayesian analyses were not immune, however, to the effects of missing data; lower resolution and support in those analyses likely arise from non-overlap of gene sequence data among less well-sampled taxa. These issues are a concern for similar studies, in which different gene sequences are concatenated in an effort to increase resolving power.     

A comparative analysis of the complete mitochondrial genome of the Eurasian otter <Emphasis Type="Italic">Lutra lutra</Emphasis> (Carnivora; Mustelidae)

Otter populations are declining throughout the world and most otter species are considered endangered. Molecular methods are suitable tools for population genetic research on endangered species. In the present study, we analyzed the complete mitochondrial genome (mitogenome) sequence of the Eurasian otter Lutra lutra. The mitochondrial DNA sequence of the Eurasian otter is 16,505 bp in length and consists of 13 protein-coding genes, 22 tRNAs, 2 rRNAs, and a control region (CR). The CR sequence of otters from Europe and Asia showed nearly identical numbers and nucleotide sequences of minisatellites. Phylogenetic analysis of Mustelidae mitogenomes, including individual genes, revealed that Lutrinae and Mustelinae form a clade, and that L. lutra and Enhydra lutris are sister taxa within the Lutrinae. Phylogenetic analyses revealed that of the 13 mitochondrial protein-coding genes, ND5 is the most reliable marker for analysis of phylogenetic relationships within the Mustelidae.     

Endogenous lentiviral elements in the weasel family (mustelidae)

Endogenous retroviruses provide molecular fossils for studying the ancient evolutionary history of retroviruses. Here, we report our independent discovery and analysis of endogenous lentiviral insertions (Mustelidae endogenous lentivirus [MELV]) within the genomes of weasel family (Mustelidae). Genome-scale screening identified MELV elements in the domestic ferret (Mustela putorius furo) genome (MELVmpf). MELVmpf exhibits a typical lentiviral genomic organization. Phylogenetic analyses position MELVmpf basal to either primate lentiviruses or feline immunodeficiency virus. Moreover, we verified the presence of MELV insertions in the genomes of several species of the Lutrinae and Mustelinae subfamilies but not the Martinae subfamily, suggesting that the invasion of MELV into the Mustelidae genomes likely took place between 8.8 and 11.8 Ma. The discovery of MELV in weasel genomes extends the host range of lentiviruses to the Caniformia (order Carnivora) and provides important insights into the prehistoric diversity of lentiviruses.     

Molecular phylogeny of the Arctoidea (Carnivora): effect of missing data on supertree and supermatrix analyses of multiple gene data sets

Phylogenetic relationships of 79 caniform carnivores were addressed based on four nuclear sequence-tagged sites (STS) and one nuclear exon, IRBP, using both supertree and supermatrix analyses. We recovered the three major arctoid lineages, Ursidae, Pinnipedia, and Musteloidea, as monophyletic, with Ursidae (bears) strongly supported as the basal arctoid lineage. Within Pinnipedia, Phocidae (true seals) were sister to the Otaroidea [Otariidae (fur seals and sea lions) and Odobenidae (walrus)]. Phocid subfamily and tribal designations were supported, but the otariid subfamily split between fur seals and sea lions was not. All family designations within Musteloidea were strongly supported: Mephitidae (skunks), Ailuridae (monotypic red panda), Mustelidae (weasels, badgers, otters), and Procyonidae (raccoons). A novel hypothesis for the position of the red panda was recovered, placing it as branching after Mephitidae and before Mustelidae+Procyonidae. Within Mustelidae, subfamily taxonomic changes are considered. This study represents the most comprehensive sampling to date of the Caniformia in a molecular study and contains the most complete molecular phylogeny for the Procyonidae. Our data set was also used in an empirical examination of the effect of missing data on both supertree and supermatrix analyses. Sequence for all genes in all taxa could not be obtained, so two variants of the data set with differing amounts of missing data were examined. The amount of missing data did not have a strong effect; instead, phylogenetic resolution was more dependent on the presence of sufficient informative characters. Supertree and supermatrix methods performed equivalently with incomplete data and were highly congruent; conflicts arose only in weakly supported areas, indicating that more informative characters are required to confidently resolve close species relationships.     

Evolutionary trends of the mitochondrial lineage differentiation in species of genera Martes and Mustela

We compared partial sequences (402 bp) of the mitochondrial cytochrome b gene in 68 individuals of martens (Martes), weasels (Mustela) and their relatives from the Northern Hemisphere to identify the modes of geographic differentiation in each species. We then compared complete sequences (1140 bp) of the gene in 17 species of the family Mustelidae to know the spatial and temporal modes of speciation, constructing linearized trees with transversional substitutions for deeper lineage divergences and with transversions and transitions for younger lineages. Our data suggested that these lineages of Martes and Mustela differentiated in a stepwise fashion with five radiation stages from the generic divergences (stage I) to the intraspecific divergences (stage V), during the last 10 or 20 million years as the fossil evidence suggests. In the lineage of Martes, the first offshoots are of Martes flavigula, M. pennanti, and Gulo gulo (stage II), the second is M. foina (stage III), and the third are M. americana, M. martes, M. melampus, and M. zibellina (stage IV). The divergence of the lineages of Mustela is likely to have taken place concurrently with the radiations of the Martes. These divergence processes are attributable in part to the geographic allocation along the two continents, North America and Eurasia, as well as among peripheral insular domains, such as Taiwan and the Japanese Islands. In addition, the Eurasian continent itself was shown to have been involved in the species diversification in the martens and weasels.     

Phylogenetic inference and comparative evolution of a complex microsatellite and its flanking regions in carnivores

We sequenced locus Mel 08, with complex short repetitive motifs, in 24 carnivore species belonging to five different families in order to explore mutational changes in the region in the context of locus and species evolution. This non-coding locus includes up to four different parts or repetitive motifs showing size variability. The variability consists of repeat additions and deletions; substitutions, insertions and/or deletions creating interruptions in the repeat; and substitutions, insertions and deletions in the flanking regions. The locus has different repeat expansions in different carnivore subfamilies. We hypothesize that the complexity of this locus is due to a high mutation rate at an ancestral DNA sequence and, thus, prompts the emergence of repeats at mutational hotspots. High levels of homoplasy were evident, with nine electromorphs representing 28 haplotypes never shared across species. The variability in flanking regions was informative for phylogenetic inference and their evolutionary content. Tree topologies were congruent with relevant hypotheses on current conflicts in carnivore phylogenies, such as: (i) the monophyly of Lutrinae, (ii) the paraphyly of Mustelinae, (iii) the basal position of the Eurasian badger, Meles meles , in the Mustelidae, (iv) the classification of skunks as a separate family, Mephitidae, and (v) the placement of the red panda, Ailurus fulgens , as a monotypic family, Ailuridae, at a basal position in the Musteloidea.     

Phylogenetic relationships within caniform carnivores based on analyses of the mitochondrial 12S rRNA gene

The complete 12S rRNA gene of 32 carnivore species, including four feliforms and 28 caniforms, was sequenced. The sequences were aligned on the basis of their secondary structures and used in phylogenetic analyses that addressed several evolutionary relationships within the Caniformia. The analyses showed an unresolved polytomy of the basic caniform clades; pinnipeds, mustelids, procyonids, skunks,Ailurus (lesser panda), ursids, and canids. The polytomy indicates a major diversification of caniforms during a relatively short period of time. The lesser panda was distinct from other caniforms, suggesting its inclusion in a monotypic family, Ailuridae. The giant panda and the bears were joined on the same branch. The skunks are traditionally included in the family Mustelidae. The present analysis, however, showed a less close molecular relationship between the skunks and the remaining Mustelidae (sensu stricto) than between Mustelidae (sensu stricto) and Procyonidae, making Mustelidae (sensu lato) paraphyletic. The results suggest that the skunks should be included in a separate family, Mephitidae. Within the Pinnipedia, the grouping of walrus, sea lions, and fur seals was strongly supported. Analyses of a combined set of 12S rRNA and cytochromeb data were generally consistent with the findings based on each gene.     

Late- and Post-Glacial history of the Mustelidae in Europe

1. Analyses of the subfossil records of mustelid species in Europe indicate specific differences in the pattern of temporal and spatial recolonization of central Europe after the maximum glaciation of the last glacial period. 2. For Meles meles, Martes martes and (with some reservations) Mustela putorius it can be seen that the populations were separated in several glacial refugia during the maximum glaciation of the Weichselian. In contrast, the European population of Lutra lutra was restricted to a single glacial refuge, which had not been clearly localized until now. 3. Besides the known glacial refugia of the Iberian Peninsula, Italian Peninsula and the Balkans, there is evidence of possible additional glacial refugia for mustelids near the Carpathians, in western Moldova and in the northern Pontic region. 4. Gulo gulo, Mustela nivalis, and Mustela erminea show adaptations for survival in Pleistocene conditions, but they were historically also distributed in the warmer areas of southern Europe. 5. Among the more thermophilic mustelid species, Mustela putorius is likely to have been the earliest immigrant following the maximum glaciation. Meles meles has been recorded in comparably early times and also seems to be relatively tolerant of climatic extremes. It is clear that Martes martes had already arrived in central Europe during the Allerød, in connection with the recolonization by birch and pine woods. Lutra lutra, by contrast, seems to have been an absolute Holocene immigrant.     

Mitochondrial genome of the eurasian otterLutra lutra (Mammalia, Carnivora, Mustelidae)

We determined the complete mitochondrial genome of the Eurasian otterLutra lutra, which is an endangered species in Korea. The circle genome (16,536 bp in size) consists of 13 protein-coding, 22 tRNA, and 2 rRNA genes, and a control region, as found in other metazoan animals. Out of the 37 genes, 28 are encoded on the H-strand, and the nine (ND6 and 8 tRNA genes) on the L-strand. Three overlaps among the 13 protein-coding genes were found: ATP8-ATP6, ND4L-ND4, and ND5-ND6. A control region (1090 bp) including the origin of H-strand replication (OH), TAS (a conserved motif TACAT-16bp-ATGTA) and CSB (CSB-1, CSB-2. and CSB-3) was observed between tRNA-Pro and tRNA-Phe genes, and O_L, with 36 highly conserved nucleotides between tRNA-Asn (N) and tRNA-Cys (C) within a cluster of five tRNA genes (WANCY), as typically found in vertebrates. The other important characteristics of theL. lutra mitochondrial genome were described in detail. In addition, a maximum likelihood and Bayesian trees of 9 mustelid species and 1 outgroup were reconstructed based on the nucleotide sequences of 11 protein-coding genes excluding ATP8 and ND6. It showed that Lutrinae formed a monophyletic group with Mustelinae that is not monophyletic. Within the subfamily Lutrinae,L. lutra andEnhydra lutris were grouped together and thenLontra canadentis placed as a sister of the clade. The present result is the first complete mitochondrial genome sequence reported from the genusLutra, and is applicable to molecular phylogenetic, phylogeographic, conservation biological studies for mustelid members. In particular, exploration of sequence variations of the control region may be helpful for analyzing inter-and intra-species variations in the genusLutra.     

Mitogenomic analyses of caniform relationships

Extant members of the order Carnivora split into two basal groups, Caniformia (dog-like carnivorans) and Feliformia (cat-like carnivorans). In this study we address phylogenetic relationships within Caniformia applying various methodological approaches to analyses of complete mitochondrial genomes. Pinnipeds are currently well represented with respect to mitogenomic data and here we add seven mt genomes to the non-pinniped caniform collection. The analyses identified a basal caniform divergence between Cynoidea and Arctoidea. Arctoidea split into three primary groups, Ursidae (including the giant panda), Pinnipedia, and a branch, Musteloidea, which encompassed Ailuridae (red panda), Mephitidae (skunks), Procyonidae (raccoons) and Mustelidae (mustelids). The analyses favored a basal arctoid split between Ursidae and a branch containing Pinnipedia and Musteloidea. Within the Musteloidea there was a preference for a basal divergence between Ailuridae and remaining families. Among the latter, the analyses identified a sister group relationship between Mephitidae and a branch that contained Procyonidae and Mustelidae. The mitogenomic distance between the wolf and the dog was shown to be at the same level as that of basal human divergences. The wolf and the dog are commonly considered as separate species in the popular literature. The mitogenomic result is inconsistent with that understanding at the same time as it provides insight into the time of the domestication of the dog relative to basal human mitogenomic divergences.     

Tandemly repeated satellite DNA ofDolichopoda schiavazzii: A test for models on the evolution of highly repetitive DNA

The evolutionary relationships among the Carnivora were studied in a phylogenetic analysis based on the complete mitochondrial cytochromeb gene. The study, which addressed primarily the relationships among the Caniformia, included 4 feliform and 26 caniform species, with 9 pinnipeds. The analysis identified five caniform clades: Canidae, Ailuridae (with the monotypic lesser panda), Musteloidea (Mustelidae+Procyonidae), Ursidae (including the giant panda), and Pinnipedia. The closest relatives of the Pinnipedia among terrestrial caniforms were not identified conclusively. Our analysis shows that the skunks are only distantly related to remaining mustelids (Mustelidae sensu stricto) and that the family Mustelidae, including the skunks, is paraphyletic. The relationship among the five caniform clades was unresolved, suggesting an evolutionary separation within a relatively short period of time. Based on distance values, we propose that this primary diversification took place 45 million years ago.