Phylogenetic relationships among the colobine monkeys revisited: new insights from analyses of complete mt genomes and 44 nuclear non-coding markers

Background

Phylogenetic relationships among Asian and African colobine genera have been disputed and are not yet well established. In the present study, we revisit the contentious relationships within the Asian and African Colobinae by analyzing 44 nuclear non-coding genes (>23 kb) and mitochondrial (mt) genome sequences from 14 colobine and 4 non-colobine primates.

Principal Findings

The combined nuclear gene and the mt genome as well as the combined nuclear and mt gene analyses yielded different phylogenetic relationships among colobine genera with the exception of a monophyletic ‘odd-nosed’ group consisting of Rhinopithecus, Pygathrix and Nasalis, and a monophyletic African group consisting of Colobus and Piliocolobus. The combined nuclear data analyses supported a sister-grouping between Semnopithecus and Trachypithecus, and between Presbytis and the odd-nosed monkey group, as well as a sister-taxon association of Pygathrix and Rhinopithecus within the odd-nosed monkey group. In contrast, mt genome data analyses revealed that Semnopithecus diverged earliest among the Asian colobines and that the odd-nosed monkey group is sister to a Presbytis and Trachypithecus clade, as well as a close association of Pygathrix with Nasalis. The relationships among these genera inferred from the analyses of combined nuclear and mt genes, however, varied with the tree-building methods used. Another remarkable finding of the present study is that all of our analyses rejected the recently proposed African colobine paraphyly and hybridization hypothesis and supported reciprocal monophyly of the African and Asian groups.

Significance

The phylogenetic utility of large-scale new non-coding genes was assessed using the Colobinae as a model, We found that these markers were useful for distinguishing nodes resulting from rapid radiation episodes such as the Asian colobine radiation. None of these markers here have previously been used for colobine phylogenetic reconstruction, increasing the spectrum of molecular markers available to mammalian systematics.     

Analysis of DNA sequences of six chloroplast and nuclear genes suggests incongruence, introgression, and incomplete lineage sorting in the evolution of Lespedeza (Fabaceae)

The genus Lespedeza (Fabaceae) consists of 40 species disjunctively distributed in East Asia and eastern North America. Phylogenetic relationships of all Lespedeza species and closely related genera were reconstructed using maximum parsimony, maximum likelihood, and Bayesian analyses of sequence data from five chloroplast (rpl16, rpl32-trnL, rps16-trnQ, trnL-F, and trnK/matK) and one nuclear (ITS) DNA regions. All analyses yielded consistent relationships among major lineages. Our results suggested that Campylotropis, Kummerowia, and Lespedeza are monophyletic, respectively. Lespedeza is resolved as sister to Kummerowia and these two together are further sister to Campylotropis. Neither of the two subgenera, subgen. Lespedeza and subgen. Macrolespedeza, in Lespedeza based on morphological characters, is recovered as monophyletic. Within Lespedeza, the North American clade is retrieved as sister to the Asian clade. The nuclear and chloroplast markers showed incongruent phylogenetic signals at shallow-level phylogeny, which may point to either introgression or incomplete lineage sorting in Lespedeza. The divergence times within Lespedeza and among related genera were estimated using Bayesian approach with BEAST. It is assumed that following the divergence between Kummerowia and Lespedeza in Asia in the late Miocene, the ancestor of Lespedeza diverged into the North American and the Asian lineages. The North American ancestor quickly migrated to North America through the Bering land bridge in the late Miocene. The North American and Asian lineages started to diversify almost simultaneously in the late Miocene but resulted in biased numbers of species in two continents.     

Evolutionary history of woodpeckers and allies (Aves: Picidae): placing key taxa on the phylogenetic tree

We analyzed 2995 base pairs of nucleotide sequence data (nuclear beta-fibrinogen intron 7 and mitochondrial cytochrome b and ND2 genes), using parsimony and model-based approaches to infer phylogenetic relationships of the woodpeckers and allies, yielding novel hypotheses for several critical gaps in the knowledge of picid phylogeny. We tested the monophyly of sub-families within the Picidae, and sampled from widely distributed and diverse genera (Celeus, Colaptes, Dryocopus, Melanerpes, Picoides, Picumnus, Sasia, Piculus, and Picus). Relationships of three poorly known Southeast Asian genera (Dinopium, Reinwardtipicus, and Blythipicus) were also examined, revealing unexpected sister relationships. All phylogenetic approaches recovered largely congruent topologies, supporting a monophyletic Picinae and paraphyletic Picumninae, with the monotypic piculet, Nesoctites micromegas, as sister to the Picinae. We report paraphyly for Celeus and Piculus, whereas the broadly distributed genera Picumnus and Dryocopus were supported as monophyletic. Our phylogenetic results indicate a complex geographic history for the Picidae, with multiple disjunct sister lineages distributed between the New World and Asia. The relationships and geographic distribution of basal picid lineages indicates an Old World origin of the Picidae; however, the geographic origin of the Picinae remains equivocal, as the sister relationship between the Caribbean N. micromegas and the true woodpeckers presents the possibility of a New World origin for the Picinae.     

Phylogeny of firs (genus <Emphasis Type="Italic">Abies</Emphasis>, Pinaceae) based on multilocus nuclear markers (AFLP)

To study the phylogenetic relationships, evolutionary history, and molecular systematics of firs (genus Abies), the phylogenetic reconstruction, based on nuclear multilocus markers—amplified fragment length polymorphism (AFLP)—was conducted. Using seven combinations of selective primers, 84 samples of 39 taxa were genotyped for 553 polymorphic AFLP loci. A comparison with our earlier chloroplast and mitochondrial phylogenies of the genus (in 2014) shows that the nuclear phylogeny generally is more congruent to the chloroplast tree. Most of the clades resolved by the chloroplast phylogeny were supported also in the AFLP tree. Employing the nuclear DNA-based tree, we revealed the presence of new groups and the differences in the topology of several clades. AFLP confirmed the monophyly of Asian species of section Balsamea and their sister position in relation to the American group of species of this section. As shown by the tree of chloroplast DNA, Asian species of section Balsamea do not form a monophyletic group, but belong to the clade comprising the majority of Asian species. Phylogenetically mitochondrial DNA data to a large extent are not congruent to the nuclear and chloroplast DNA trees, and are more in line with geographical distribution of species. Conflicts between nuclear and cytoplasmic phylogeny were analyzed. Taking them into account, we consider the hypothesis of a hybrid origin of particular groups of firs, including ancient hybridization in section Balsamea. A comparison of molecular data with traditional taxonomy of the genus is discussed.     

Monophyly and phylogenetic relationships of the catfish genus Glyptothorax (Teleostei: Sisoridae) inferred from nuclear and mitochondrial gene sequences

GlyptothoraxBlyth (1860) is the most species-diverse and widely-distributed genus in the Sisoridae, but few studies have examined monophyly of the genus and phylogenetic relations within it. We used the nuclear RAG2 gene and mitochondrial COI and Cyt b genes from 50 of the approximately 70 species to examine monophyly of Glyptothorax and phylogenetic relationships within the genus. Molecular phylogenetic trees were constructed using maximum parsimony, maximum likelihood and Bayesian inference methods. All methods strongly supported monophyly of Glyptothorax, with Bagarius as its sister group. Both analyses of two- and three-gene datasets recovered nine major subclades of Glyptothorax, but some internal nodes remained poorly resolved. The phylogenetic relationships within the genus and existing taxonomic problems are discussed.     

Phylogeny and reclassification of species groups in Aquarius Schellenberg, Limnoporus Stål and Gerris Fabricius (Insecta: Hemiptera-Heteroptera, Gerridae)

Abstract. We investigated phylogenetic relationships of water striders (Hemiptera‐Heteroptera: Gerridae) from the three principal Holarctic genera, Aquarius Schellenberg, Limnoporus Stål and Gerris Fabricius with parsimony analyses of sixty‐six morphological characters and DNA sequences from mitochondrial (cytochrome c oxidase subunit I + II; large mitochondrial ribosomal subunit) and nuclear (elongation factor 1‐alpha) genes. The taxon sampling included all species of Aquarius and Limnoporus, and a dense, near complete, sample of Gerris species with representatives from all subgenera and species groups, and Gigantometra gigas (China) was selected as an outgroup species. A simultaneous analysis of all data sets gave eight equally parsimonious trees, and a strict consensus tree left only a few relationships within Gerris unresolved. While Limnoporus and Gerris each were resolved as monophyletic entities, Aquarius was found to be polyphyletic, because the Nearctic Aquarius remigis‐group, comprising A. remigis (Say), A. amplus (Drake and Harris), A. nyctalis (Drake and Hottes) and A. remigoides Gallant and Fairbairn, was placed as sister group to Gerris, while the Andean Aquarius chilensis (Berg) was sister group to all three genera. Remaining species of Aquarius comprised a sister group to the Gerris + the A. remigis‐group clade. Based on our phylogenetic reconstruction we discuss relationships within and among the three genera, reassess and diagnose species groups, and discuss zoogeographical relationships among all taxa.     

Phylogenetic position of the genus Tetraophasis (Aves, Galliformes, Phasianidae) as inferred from mitochondrial and nuclear sequences

The genus Tetraophasis is endemic to China and the systematic relationships of the genus in Phasianidae based solely on their morphology are not clear. The phylogenetic position of this genus and other genera in Phasianidae was investigated in this study using the mitochondrial genes cytochrome b (CYT B), NADH dehydrogenase subunit 2 (ND2) and nuclear ovomucoid intron G (OVO-G). Maximum parsimony, Bayesian and maximum likelihood analyses were employed. Comparison between the mitochondrial and nuclear findings suggested that our final combined data had higher resolving power than the separate data. Both separate data and combined data yielded a sister relationship between Tetraophasis and Lophophorus with strong support, which provided new evidence that pheasants and partridges were not monophyletic. In addition, our results indicated a basal position of Ithaginis, but the position of Perdix and Pucrasia were still unresolved. The result of this study provided a phylogenetic background for a revised classification of Galliformes.     

Phylogenetic Analysis of the Spider Mite Sub-Family Tetranychinae (Acari: Tetranychidae) Based on the Mitochondrial COI Gene and the 18S and the 5′ End of the 28S rRNA Genes Indicates That Several Genera Are Polyphyletic

The spider mite sub-family Tetranychinae includes many agricultural pests. The internal transcribed spacer (ITS) region of nuclear ribosomal RNA genes and the cytochrome c oxidase subunit I (COI) gene of mitochondrial DNA have been used for species identification and phylogenetic reconstruction within the sub-family Tetranychinae, although they have not always been successful. The 18S and 28S rRNA genes should be more suitable for resolving higher levels of phylogeny, such as tribes or genera of Tetranychinae because these genes evolve more slowly and are made up of conserved regions and divergent domains. Therefore, we used both the 18S (1,825–1,901 bp) and 28S (the 5′ end of 646–743 bp) rRNA genes to infer phylogenetic relationships within the sub-family Tetranychinae with a focus on the tribe Tetranychini. Then, we compared the phylogenetic tree of the 18S and 28S genes with that of the mitochondrial COI gene (618 bp). As observed in previous studies, our phylogeny based on the COI gene was not resolved because of the low bootstrap values for most nodes of the tree. On the other hand, our phylogenetic tree of the 18S and 28S genes revealed several well-supported clades within the sub-family Tetranychinae. The 18S and 28S phylogenetic trees suggest that the tribes Bryobiini, Petrobiini and Eurytetranychini are monophyletic and that the tribe Tetranychini is polyphyletic. At the genus level, six genera for which more than two species were sampled appear to be monophyletic, while four genera (Oligonychus, Tetranychus, Schizotetranychus and Eotetranychus) appear to be polyphyletic. The topology presented here does not fully agree with the current morphology-based taxonomy, so that the diagnostic morphological characters of Tetranychinae need to be reconsidered.     

Toward a Better Understanding of the Phylogeny of the Asplanchnidae (Rotifera)

We investigated the phylogenetic relationships of Family Asplanchnidae using both morphological and molecular data. The morphological database, comprising 23 characters from 19 taxa (15 Asplanchnidae and 4 outgroups), was compiled from a survey of the literature and our own observations; the molecular data (ITS and V4 region nuclear regions and mitochondrial cox1) was sequenced from specimens that we collected. Our analysis of the morphological data set (maximum parsimony) yielded 12 most-parsimonious trees with a tree length of 27 steps. From this analysis we conclude (1) Asplanchnidae is a monophyletic group as are the three genera comprising it, (2) there is no compelling support for the argument that Asplanchna should be separated into two discrete genera, and (3) there is some support for the proposal that Asplanchnidae and Synchaetidae are sister groups. Our analysis of the molecular data set supports the first two of these conclusions while the sister group of the family varied depending on the gene region analyzed and families and genera included. Current understanding of the phylogeny of Asplanchnidae is hampered by the need for additional informative morphological characters and a lack of molecular data for the genus Harringia and several other members of the Asplanchnidae.     

Molecular Phylogeny of Tribe Theeae (Theaceae s.s.) and Its Implications for Generic Delimitation

Tribe Theeae, which includes some economically important and widely grown plants, such as beverage tea and a number of woody ornamentals, is the largest member of the Theaceae family. Using five genomic regions (chloroplast: atpI-H, matK, psbA5''R-ALS-11F, rbcL; nuclear: LEAFY) and 30 species representing four of the five genera in this tribe (Apterosperma, Camellia, Polyspora, and Pyrenaria s.l.), we investigated the phylogeny of Theeae and assessed the delimitation of genera in the tribe. Our results showed that Polyspora was monophyletic and the sister of the three other genera of Theeae investigated, Camellia was paraphyletic and Pyrenaria was polyphyletic. The inconsistent phylogenetic placement of some species of Theeae between the nuclear and chloroplast trees suggested widespread hybridization between Camellia and Pyrenaria, Polyspora and Parapyrenaria. These results indicate that hybridization, rather than morphological homoplasy, has confused the current classification of Theeae. In addition, the phylogenetic placement and possible allies of Laplacea are also discussed.     

Molecular phylogeny of the subfamily Schizothoracinae (Teleostei: Cypriniformes: Cyprinidae) inferred from complete mitochondrial genomes

The schizothoracine fishes, members of the Teleost order Cypriniformes, are one of the most diverse group of cyprinids in the Qinghai–Tibetan Plateau and surrounding regions. However, taxonomy and phylogeny of these species remain unclear. In this study, we determined the complete mitochondrial genome of Schizopygopsis malacanthus. We also used the newly obtained sequence, together with 31 published schizothoracine mitochondrial genomes that represent eight schizothoracine genera and six outgroup taxa to reconstruct the phylogenetic relationships of the subfamily Schizothoracinae by different partitioned maximum likelihood and partitioned Bayesian inference at nucleotide and amino acid levels. The schizothoracine fishes sampled form a strongly supported monophyletic group that is the sister taxon to Barbus barbus. A sister group relationship between the primitive schizothoracine group and the specialized schizothoracine group + the highly specialized schizothoracine group was supported. Moreover, members of the specialized schizothoracine group and the genera Schizothorax, Schizopygopsis, and Gymnocypris were found to be paraphyletic.     

Phylogenetic analysis of eastern Asian and eastern North American disjunct Lespedeza (Fabaceae) inferred from nuclear ribosomal ITS and plastid region sequences

Lespedeza (tribe Desmodieae, Fabaceae) follows a disjunct distribution in eastern Asia and eastern North America. Phylogenetic relationships among its species and related taxa were inferred from nuclear ribosomal internal transcribed spacer (ITS) and plastid sequences (trnH‐psbA, psbK‐psbI, trnK‐matK and rpoC1). We examined 35 species of Lespedeza, two of Kummerowia and one of Campylotropis, the sole constituents of the Lespedeza group. An analysis of these data revealed that the genus Campylotropis is sister to the other two genera. However, we were unable to resolve the relationships between Kummerowia and Lespedeza in the strict consensus trees of parsimony analyses based on plastid and combined DNA data. In the genus Lespedeza, the Old World subgenus Macrolespedeza is monophyletic, whereas the transcontinental subgenus Lespedeza is paraphyletic. Monophyly of eastern Asian species and of North American species is strongly supported. Although inconsistent with the traditional classification, this phylogenetic finding is consistent with seedling morphology. Three subgroups recognized in subgenus Macrolespedeza were unresolved in our phylogenetic trees. An incongruence length difference (ILD) test indicated that the two partitions (nuclear ITS and plastid sequences) were significantly incongruent, perhaps because of hybridization between species in Lespedeza. Most of the primary clades of tribe Desmodieae are Asian, implying that the relatively few New World ones, such as those in Lespedeza, are more recently derived from Asia. © 2010 The Linnean Society of London, Botanical Journal of the Linnean Society, 2010, 164 , 221–235.     

Mitogenomic analysis of decapod crustacean phylogeny corroborates traditional views on their relationships

Phylogenetic relationships within decapod crustaceans are highly controversial. Even recent analyses based on molecular datasets have shown largely contradictory results. Previous studies using mitochondrial genomes are promising but suffer from a poor and unbalanced taxon sampling. To fill these gaps we sequenced the (nearly) complete mitochondrial genomes of 13 decapod species: Stenopus hispidus, Polycheles typhlops, Panulirus versicolor, Scyllarides latus, Enoplometopus occidentalis, Homarus gammarus, Procambarus fallax f. virginalis, Upogebia major, Neaxius acanthus, Calocaris macandreae, Corallianassa coutierei, Cryptolithodes sitchensis, Neopetrolisthes maculatus, and add that of Dromia personata. Our new data allow for comprehensive analyses of decapod phylogeny using the mitochondrial genomes of 50 species covering all major taxa of the Decapoda. Five species of Stomatopoda and one species of Euphausiacea serve as outgroups. Most of our analyses using Maximum Likelihood (ML) and Bayesian inference (BI) of nucleotide and amino acid datasets revealed congruent topologies for higher level decapod relationships: (((((((Anomala, Brachyura), Thalassinida: Gebiidea), Thalassinida: Axiidea), (Astacidea, Polychelida), Achelata), Stenopodidea), Caridea), Dendrobranchiata). This result corroborates several traditional morphological views and adds new perspectives. In particular, the position of Polychelida is surprising. Nevertheless, some problems can be identified. In a minority of analyses the basal branching of Reptantia is not fully resolved, Thalassinida are monophyletic; Polychelida are the sister group to Achelata, and Stenopodidea are resolved as sister group to Caridea. Despite this and although some nodal supports are low in our phylogenetic trees, we think that the largely stable topology of the trees regardless of different types of analyses suggests that mitochondrial genomes show good potential to resolve the relationship within Decapoda.     

Phylogeny of the East Asian botiine loaches (Cypriniformes,Botiidae) inferred from mitochondrial cytochrome <Emphasis Type="Italic">b</Emphasis> gene sequences

The Botiinae have traditionally represented a subfamily of the Cobitidae. At present, the classification and phylogenetic relationships of the Botiinae are controversial. To address systematic and phylogenetic questions concerning this group, we sequenced the complete cytochrome b gene from 34 samples, of which 24 represented 13 species of the East Asian botiine fishes, while the other 10 were non-botiine loach species. For the 1140 bp sequences determined, 494 sites were variable ones, of which 424 were parsimony informative. With Myxocyprinus asiaticus as an outgroup, molecular phylogenetic trees were constructed using the neighbor-joining, maximum parsimony, maximum likelihood and Bayesian methods. All molecular phylogenetic trees revealed that botiine fishes form a monophyletic group and are distantly related to other loaches, suggesting that the Botiinae should be placed in their own family. Within the Botiinae, there are three genera; Botia, Parabotia, andLeptobotia, each genus forming a monophyletic group, with the genus Botia as the most ancestral split. Our molecular results are in agreement with morphological analyses of botiines, suggesting that Botia is the ancestral genus, while Leptobotia and Parabotia were resolved as more derived sister groups.     

Phylogenetic relationships of elapid snakes based on cytochrome b mtDNA sequences

Published molecular phylogenetic studies of elapid snakes agree that the marine and Australo-Melanesian forms are collectively monophyletic. Recent studies, however, disagree on the relationships of the African, American, and Asian forms. To resolve the relationships of the African, American, and Asian species to each other and to the marine/Australo-Melanesian clade, we sequenced the entire cytochrome b gene for 28 elapids; 2 additional elapid sequences from GenBank were also included. This sample includes all African, American, and Asian genera (except for the rare African Pseudohaje), as well as a representative sample of marine/Australo-Melanesian genera. The data were analyzed by the methods of maximum-parsimony and maximum-likelihood. Both types of analyses yielded similar trees, from which the following conclusions can be drawn: (1) Homoroselaps falls outside a clade formed by the remaining elapids; (2) the remaining elapids are divisible into two broad sister clades, the marine/Australo-Melanesian species vs the African, American, and Asian species; (3) American coral snakes cluster with Asian coral snakes; and (4) the "true" cobra genus Naja is probably not monophyletic as the result of excluding such genera as Boulengerina and Paranaja.     

Evolution of blindness in scolopendromorph centipedes (Chilopoda: Scolopendromorpha): insight from an expanded sampling of molecular data

Relative to its diversity (34 genera, 700 species), Scolopendromorpha has been undersampled in molecular phylogenetic analyses compared with the other chilopod orders. Previous analyses based on morphology have not resolved several key controversies in systematics and evolutionary morphology unambiguously. Here we apply new molecular and morphological data to scolopendromorph phylogenetics, with a focus on the evolution of blindness. The taxonomic sample includes 19 genera, many lacking previous molecular data, and diverse, cosmopolitan genera of Scolopendridae are sampled by multiple species. Phylogenetic analysis with Direct Optimization used 94 morphological characters and ca. 4.5 kb of sequence data from two nuclear (18S and 28S rRNA) and two mitochondrial (16S rRNA and COI) loci. A single most‐parsimonious cladogram selected after sensitivity analyses resolves Scolopendromorpha as monophyletic, and divides it into a blind clade of three families (Plutoniumidae, Cryptopidae, Scolopocryptopidae) and its ocellate sister group, Scolopendridae. Some species‐rich, cosmopolitan genera (Cormocephalus, Otostigmus, Scolopendra) in Scolopendridae are non‐monophyletic, and in several instances (e.g. New and Old World Scolopendra) relationships are more congruent with geographical distributions than with traditional classifications. The tribe Asanadini is particularly subject to parameter‐sensitivity, nesting in the combined analysis within Scolopendrini but as sister to all other Scolopendrinae for molecular data alone. The total‐evidence tree unambiguously optimizes trunk segmentation: a 23‐segmented trunk has a single origin in the blind clade. © The Willi Hennig Society 2011.     

A Phylogeny of the Family Poritidae (Cnidaria,Scleractinia) Based on Molecular and Morphological Analyses

The family Poritidae formerly included 6 genera: Alveopora, Goniopora, Machadoporites, Porites, Poritipora, and Stylaraea. Morphologically, the genera can be differentiated based on the number of tentacles, the number of septa and their arrangement, the length of the polyp column, and the diameter of the corallites. However, the phylogenetic relationships within and between the genera are unknown or contentious. On the one hand, Alveopora has been transferred to the Acroporidae recently because it was shown to be more closely related to this family than to the Poritidae by previous molecular studies. On the other hand, Goniopora is morphologically similar to 2 recently described genera, Machadoporites and Poritipora, particularly with regard to the number of septa (approximately 24), but they have not yet been investigated at the molecular level. In this study, we analyzed 93 samples from all 5 poritid genera and Alveopora using 2 genetic markers (the barcoding region of the mitochondrial COI and the ITS region of the nuclear rDNA) to investigate their phylogenetic relationships and to revise their taxonomy. The reconstructed molecular trees confirmed that Alveopora is genetically distant from all poritid genera but closely related to the family Acroporidae, whereas the other genera are genetically closely related. The molecular trees also revealed that Machadoporites and Poritipora were indistinguishable from Goniopora. However, Goniopora stutchburyi was genetically isolated from the other congeneric species and formed a sister group to Goniopora together with Porites and Stylaraea, thus suggesting that 24 septa could be an ancestral feature in the Poritidae. Based on these data, we move G. stutchburyi into a new genus, Bernardpora gen. nov., whereas Machadoporites and Poritipora are merged with Goniopora.     

Molecular phylogeny of Pamphagidae (Acridoidea,Orthoptera) from China based on mitochondrial cytochrome oxidase II sequences

Abstract Phylogenetic relationships of Pamphagidae were examined using cytochrome oxidase subunit II (COII) mtDNA sequences (684 bp). Twenty‐seven species of Acridoidea from 20 genera were sequenced to obtain mtDNA data, along with four species from the GenBank nucleotide database. The purpose of this study was analyzing the phylogenetic relationships among subfamilies within Pamphagidae and interpreting the phylogenetic position of this family within the Acridoidea superfamily. Phylogenetic trees were reconstructed using neighbor‐joining (NJ), maximum parsimony (MP) and Bayesian inference (BI) methods. The 684 bp analyzed fragment included 126 parsimony informative sites. Sequences diverged 1.0%–11.1% between genera within subfamilies, and 8.8%–12.3% between subfamilies. Amino acid sequence diverged 0–6.1% between genera within subfamilies, and 0.4%–7.5% between subfamilies. Our phylogenetic trees revealed the monophyly of Pamphagidae and three distinct major groups within this family. Moreover, several well supported and stable clades were found in Pamphagidae. The global clustering results were similar to that obtained through classical morphological classification: Prionotropisinae, Thrinchinae and Pamphaginae were monophyletic groups. However, the current genus Filchnerella (Prionotropisinae) was not a monophyletic group and the genus Asiotmethis (Prionotropisinae) was a sister group of the genus Thrinchus (Thrinchinae). Further molecular and morphological studies are required to clarify the phylogenetic relationships of the genera Filchnerella and Asiotmethis.     

Molecular Evidence for Multiple Origins of the European Spined Loaches (Teleostei,Cobitidae)

We present a phylogenetic investigation of the Northern Clade, the major monophyletic clade within the freshwater fish family Cobitidae, one of the most prominent families of freshwater fishes found in Asian and European waters. Phylogenetic reconstructions based on the cytochrome b and RAG-1 genes show the genera Microcobitis, Sabanejewia, Koreocobitis and Kichulchoia as monophyletic groups. These reconstructions also show a Cobitis sensu lato and a Misgurnus sensu lato group. The Cobitis sensu lato group includes all species of Cobitis, Iksookimia, Niwaella and Kichulchoia, while the Misgurnus sensu lato group includes Misgurnus, Paramisgurnus and Koreocobitis. Although the monophyly of both the Cobitis sensu lato and Misgurnus sensu lato groups is supported, relationships within the groups are incongruent with current generic definitions. The absence of monophyly of most genera included in the Cobitis sensu lato group (Cobitis, Iksookimia and Niwaella) or their low genetic differentiation (Kichuchoia) supports their consideration as synonyms of Cobitis. Molecular phylogenies indicate that the Asian species of Misgurnus experienced a mitochondrial introgression from a lineage of Cobitis. We also find two nuclear haplotypes in the same Cobitis species from the Adriatic area that, in the absence of morphological differentiation, may indicate molecular introgression. Most lineages within the Northern Clade consist of species found in East Asia. However, some lineages also contain species from Europe and Asia Minor. The phylogenetic relationships presented here are consistent with previous studies suggesting an East Asian origin of the Northern Clade. According to the current distributions and phylogenetic relationships of the Misgurnus sensu lato and Cobitis clade lineages, particularly of M. fossilis and C. melanoleuca, the range expansion of East Asian species into Europe was most likely via Siberia into Northern and Central Europe. Phylogenetic analyses also show that the Cobitis sensu lato group consists of two clear subgroups (I and II), each presenting geographical differences. Subgroup I is distributed exclusively in East Asian drainages with an Eastern European offshoot (C. melanoleuca), whereas Subgroup II includes species widespread throughout Europe (including the Mediterranean), Asia Minor, the Black Sea and the Caucasus, with some lineages related to species restricted to East Asia.