A molecular phylogeny of the wild onions (Allium; Alliaceae) with a focus on the western North American center of diversity

Nuclear ribosomal DNA (ITS and ETS) sequences from 39 native Californian (USA) Allium species and congeners were combined with 154 ITS sequences available on GenBank to develop a global Allium phylogeny with the simultaneous goals of investigating the evolutionary history (monophyly) of Allium in the Californian center of diversity and exploring patterns of adaptation to serpentine soils. Phylogenies constructed with ITS alone or ITS in combination with ETS provided sufficient resolution for investigating evolutionary relationships among species. The ITS region alone was sufficient to resolve the deeper relationships in North American species. Addition of a second marker (ETS) further supports the phylogenetic placements of the North American species and adds resolution within subgenus Amerallium, a clade containing many Californian endemics. Within the global phylogeny, the native North American species were found to be monophyletic, with the exception of Allium tricoccum and Allium schoenoprasum. All native Californian species included in the analysis fell into a monophyletic subgenus Amerallium section Lophioprason, although endemic Californian species were not monophyletic due to the inclusion of species with ranges extending beyond the California Floristic Province. The molecular phylogeny strongly supports previous morphology-based taxonomic groupings. Based on our results, serpentine adaptation appears to have occurred multiple times within section Lophioprason, while the ancestor of the Californian center of diversity may not have been serpentine-adapted.     

Phylogenetic relationships in Nuphar (Nymphaeaceae): evidence from morphology,chloroplast DNA,and nuclear ribosomal DNA

The genus Nuphar consists of yellow-flowered waterlilies and is widely distributed in north-temperate bodies of water. Despite regular taxonomic evaluation of these plants, no explicit phylogenetic hypotheses have been proposed for the genus. We investigated phylogenetic relationships in Nuphar using morphology and sequences of the chloroplast gene matK and of the internal transcribed spacer (ITS) regions of nuclear ribosomal DNA. Two major lineages within Nuphar are consistently resolved with the morphological and molecular data sets. One lineage comprises New World taxa and the other represents a primarily Old World lineage. Relationships within the major lineages were poorly resolved by morphology and ITS, yet certain relationships were elucidated by all analyses. Most notable is the strong support for a monophyletic lineage of dwarf taxa and the alliance of the North American N. microphylla with the Eurasian taxa. Minor discordance between the independent cladograms is accounted for by hybridization. The common taxonomic practice of uniting all North American and Eurasian taxa under one species is not supported phylogenetically.     

Phylogeny and biogeography of the flowering plant genus Styrax (Styracaceae) based on chloroplast DNA restriction sites and DNA sequences of the internal transcribed spacer region

Phylogenetic relationships within the flowering plant genus Styrax were investigated with DNA sequence data from the internal transcribed spacer (ITS) region of nuclear ribosomal DNA (nrDNA) and with chloroplast DNA restriction site data from the genes trnK, rpoC1, and rpoC2. The data sets from each genome were analyzed separately and in combination with parsimony methods. The results strongly support the monophyly of each of the four series of the genus but provide little phylogenetic resolution among them. Reticulate evolution may at least partly explain discordance between the molecular phylogenetic estimates and a prior morphological estimate within series Cyrta. The historical biogeography of the genus was inferred with unweighted parsimony character optimization of trees recovered from a combined ITS and morphological data set, after a series of combinability tests for data set congruence was conducted. The results are consistent with the fossil record in supporting a Eurasian origin of Styrax. The nested phylogenetic position of the South American members of the genus within those from southern North America and Eurasia suggests that the boreotropics hypothesis best explains the amphi-Pacific tropical disjunct distribution occurring within section Valvatae. The pattern of relationship recovered among the species of section Styrax ((western North America + western Eurasia) (eastern North America + eastern Eurasia)) is rare among north-temperate Tertiary forest relicts. The monophyly of the group of species from western North America and western Eurasia provides qualified support for the Madrean-Tethyan hypothesis, which posits a Tertiary floristic connection among the semiarid regions in which these taxa occur. A single vicariance event between eastern Asia and eastern North America accounts for the pattern of relationship among intercontinental disjuncts in series Cyrta.     

Biogeographical and evolutionary insights onErigeron and allies (Asteraceae) from ITS sequence data

Complex geographical distribution patterns and agamospermy have united to obscure systematic relationships in the genusErigeron (400+ species). Sequence data were analyzed from the internal transcribed spacer region of nrDNA (ITS) and 5.8S cistron for 77 taxa including 63Erigeron species. Results support the North American origin ofErigeron and document thatAphanostephus, Conyza, and three small genera restricted to South America (Apopyros, Hysterionica, Neja) are all derived from withinErigeron. Phylogenetic data indicate that agamospermy has arisen at least three times inErigeron and that autogamy has evolved independently inConyza and inE. sect.Trimorpha.     

The position ofCastrilanthemum Vogt & Oberprieler and the phylogeny of Mediterranean Anthemideae (Compositae) as inferred from nrDNA ITS and cpDNAtrnL/trnF IGS sequence variation

Sequences of the nrDNA internal transcribed spacer (ITS) region and the cpDNAtrnL/trnF intergenic spacer (IGS) region were analysed for 41 Mediterranean and Eurasian representatives of the Anthemideae (Compositae) to ascertain the systematic position of the unispecific genusCastrilanthemum Vogt & Oberprieler and to study the phylogeny of the Anthemideae in the Mediterranean region. Both markers consistently point to the close relationship ofCastrilanthemum with the W Mediterranean generaLeucanthemopsis (Giroux) Heywood andProlongoa Boiss., forming a strongly supported monophyletic group (theLeucanthemopsis-group which also comprises the unispecific genusHymenostemma (Kunze) Willk.). Results also demonstrate that subtribes Achilleinae, Leucantheminae, and Matricariinae sensu Bremer & Humphries are non-monophyletic. Besides results from sequence variation, a marked 5bp-deletion intrnL/trnF IGS divides all these subtribes into more basal subgroups which are related to each other and to the large Eurasian generaTanacetum L. andAnthemis L., and a monophyletic group of closely related and more advanced subgroups which also contain the monophyletic Chrysantheminae sensu Bremer & Humphries. For this second group a W Mediterranean centre of diversification is suggested, however, its sister-group relationships within the basal grade of generic groups remain unclear.     

四照花亚属的nrDNA ITS致同进化不完全

四照花亚属(Cornus subg.Syncarpea)隶属于山茱萸科山茱萸属(Cornus),我国该亚属共有5种8亚种。为探讨四照花亚属nrDNA ITS序列的致同进化不完全现象及假基因产生的可能原因,分析了该亚属4种(每种1~2个居群)共21个个体的nrDNA ITS序列。结果表明,这些类群的nrDNA ITS存在多态性,通过分析这些nrDNA ITS克隆序列的G+C含量、5.8S保守基序和二级结构最小自由能,推测其可能存在假基因。系统发育研究结果显示所有nrDNA ITS序列分成5个分支,同一个体的不同拷贝被分别置于两个甚至多个分支中,且不同分支显示了不同种间关系。四照花亚属物种个体内部存在nrDNA ITS不完全致同进化,可能归咎于不完全的世系分选(incomplete lineage sorting)、种间杂交或多倍化等进化事件,从而导致基因组内nrITS区序列出现多态性,同时也导致难以通过外部形态来划分亚属内种间界限。     

Molecular phylogeny of the Acre clade (Crassulaceae): Dealing with the lack of definitions for Echeveria and Sedum

The phylogenetic relationships within many clades of the Crassulaceae are still uncertain, therefore in this study attention was focused on the “Acre clade”, a group comprised of approximately 526 species in eight genera that include many Asian and Mediterranean species of Sedum and the majority of the American genera (Echeveria, Graptopetalum, Lenophyllum, Pachyphytum, Villadia, and Thompsonella). Parsimony and Bayesian analyses were conducted with 133 species based on nuclear (ETS, ITS) and chloroplast DNA regions (rpS16, matK). Our analyses retrieved four major clades within the Acre clade. Two of these were in a grade and corresponded to Asian species of Sedum, the rest corresponded to a European–Macaronesian group and to an American group. The American group included all taxa that were formerly placed in the Echeverioideae and the majority of the American Sedoideae. Our analyses support the monophyly of three genera – Lenophyllum, Thompsonella, and Pachyphytum; however, the relationships among Echeveria, Sedum and the various segregates of Sedum are largely unresolved. Our analyses represents the first broad phylogenetic framework for Acre clade, but further studies are necessary on the groups poorly represented here, such as the European and Asian species of Sedum and the Central and South American species of Echeveria.     

Molecular phylogenetics of the tribeInuleae s. str.(Asteraceae), based on ITS sequences of nuclear ribosomal DNA

The sequences of the internal transcribed spacer (ITS) region of 18S–26S nrDNA for a sample of 16 taxa from theInuleae s. str. and two outgroup taxa are analysed cladistically with PAUP. A consensus tree of the four most parsimonious cladograms is presented. Three different tests of cladogram stability are conducted (Bremer support, parsimony jackknifing and bootstrapping); all tests indicate a high degree of support for the basal nodes of the tree. The ITS phylogeny of the tribe is compared with previous hypotheses based on morphological data. The position ofAnisopappus as sister group to the rest of the tribe is supported by the molecular data, but the proposed subdivision ofInuleae s. str. into a paleate grade group and an epaleate clade is not. The interpretation of the character evolution of, e.g. receptacular paleae and pappus features within the tribe is discussed.     

Internal transcribed spacer region evolution in Larix and Pseudotsuga (Pinaceae)

The nuclear ribosomal DNA (nrDNA) internal transcribed spacer (ITS) region has been characterized in the sister genera Larix and Pseudotsuga (Pinaceae). Complete sequences were obtained for seven species of Larix from North America and Eurasia and five species of Pseudotsuga from western North America and eastern Asia. ITS region lengths ranged from 1759 to 1770 bp in Larix and from 1564 to 1571 bp in Pseudotsuga. In both genera, ITS1 is three times as long as the 5.8S plus ITS2 and contains subrepeats as observed in other genera of Pinaceae. Secondary structure models predicted that the subrepeats fold into terminal stem and loop domains. ITS polymorphism detected within individuals of Larix and Pseudotsuga suggests a slow rate of concerted evolution among nrDNA loci. Except for the placement of L. sibirica, phylogenetic analyses of the ITS region agreed with previously reported restriction site analyses of Larix and Pseudotsuga. The data were not consistent with phylogenetic hypotheses for Larix based primarily upon ovulate cone characters, failing to support a derivation of the North American L. laricina from a short-bracted Eurasian lineage. The phylogenetic hypothesis did not conflict with a stepping stone model of evolution for Pseudotsuga, but a basal lineage could not be inferred for either genus.     

A phylogenetic analysis of the Illiciaceae based on sequences of internal transcribed spacers (ITS) of nuclear ribosomal DNA

Sequences of the internal transcribed spacers (ITS) of nuclear ribosomal DNA were determined for 15 species ofIllicium (Illiciaceae) to examine phylogenetic relationships. The ITS trees show a major dichotomy between the two North American species (I. floridanum andI. parviflorum) and the remaining east Asian species. This suggests that the existing division between two sections (sect.Illicium and sect.Cymbostemon) ofIllicium based on tepal characters in unnatural. The ITS phylogeny shows congruence with palynology: of the species examined, the three species (I. angustisepalum, I. anisatum andI. fargesii) from sect.Illicium that possess trizonocolpate pollen consistently form a clade, although nesting within a clade consisting of the species of sect.Cymbostemon, which generally have trisyncolpate pollen. The low ITS sequence divergence and the close relationship among east Asian species suggest a recent diversification of this group of species or an unusual slowdown of sequence mutations.     

Phylogenetic relationship of China Dendrobium species based on the sequence of the internal transcribed spacer of ribosomal DNA

The genetic relationship of 36 Dendrobium species in China was determined based on sequence analysis of the internal transcribed spacer (ITS) region of ribosomal DNA. Aligned sequences of the complete ITS region obtained from the 36 Dendrobium species and 2 outgroup species (Epigeneium amplum and Epigeneium nakaharaei) by using PCR amplification and direct DNA sequencing. The nrDNA ITS1 of Dendrobium was 225–234 bp and ITS2 was 239–248 bp. Phylogenetic tree was constructed, and seven main clusters were generated among the 36 Dendrobium species. From the results, D. moulmeinense was not grouped in the classification of Dendrobium and E. amplum and E. nakaharaei were shown to be divergent from Dendrobium species. The phylogenetic relationships revealed by ITS DNA analysis partially supported previously published morphological data.     

Phylogeny of Bonatea (Orchidaceae: Habenariinae) based on molecular and morphological data

The genus Bonatea is widely distributed throughout southern and eastern Africa. Considerable debate surrounds the generic status of Bonatea, but there have been neither previous studies of evolutionary relationships among Bonatea species, nor any tests of the monophyly of the genus in relation to its close relative Habenaria. We investigated phylogenetic relationships between Bonatea and selected Habenaria species using morphology, as well as sequences of the chloroplast gene matK and the internal transcribed spacer (ITS) regions of nuclear ribosomal DNA. A fully resolved cladogram was obtained using morphological data, but neither the ITS, matK, nor combined data sets yielded well-resolved and well-supported phylogenetic structure for Bonatea. There is poor congruence between ITS and matK data for interspecific relationships in Bonatea, whilst the morphological results are largely congruent with the ITS analysis. Relative to Habenaria, there is little sequence variation between Bonatea species, which could indicate a recent and rapid radiation of Bonatea. Although the sampled Bonatea species form a distinct clade, more extensive sampling of Habenaria would be required to establish unambiguously whether or not Bonatea is monophyletic.     

Molecular phylogeny of selected Old World Astragalus (Fabaceae): incongruence among chloroplast trnL-F, ndhF and nuclear ribosomal DNA ITS sequences

This study reports maximum parsimony and Bayesian phylogenetic analyses of selected Old World Astragalus using two chloroplast fragments including trnL-F and ndhF and the nuclear ribosomal internal transcribed spacer (nrDNA ITS). A total of 52 taxa including 34 euploid Old World and New World Astragalus , one aneuploid species from the Neo-Astragalus clade as a representative and 14 other Astragalean taxa, plus Cheseneya astragalina and two species of Caragana as outgroups were analyzed for both trnL-F and nrDNA ITS regions. ndhF was analyzed in 30 taxa and the same number for the combination of these three datasets were examined. In general, the trnL-F dataset and the ndhF and nrDNA ITS datasets generated more or less the same clades within Astragalus . However, in the trnL-F and ndhF phylogenies, Astragalus species are not gathered in a single clade, the so-called Astragalus s.s., as indicated by the nrDNA ITS tree. Visual inspection of these three phylogenies revealed that they were inconsistent regarding the position and relationships of Astragalus hemsleyi , A. ophiocarpus , A. annularis–A. epiglottis / Astragalus pelecinus, A. echinatus and A. arizonicus . Incongruence length difference test suggested that the trnL-F , ndhF and nrDNA ITS datasets were incongruent. In spite of this, phylogenetic analyses of the combined datasets as one unit or as three partitions generated trees that were topologically similar as a mix of the cpDNA and the nrDNA ITS trees. However, the combined dataset provided more resolved and statistically supported clades. The recently described A. memoriosus appeared closely related to A. stocksii (both from sect. Caraganella ) based on both trnL-F and nrDNA ITS sequences.     

Phylogenetic placement of <Emphasis Type="Italic">Marasmiellus juniperinus</Emphasis>

Recent collections and the type specimen of Marasmiellus juniperinus, the type species of the genus, were examined. Phylogenetic placement, based on ribosomal large subunit (LSU) and internally transcribed spacer (ITS) sequences, is within the lentinuloid clade, nested among Gymnopus taxa. This placement dictates genus name usage and phylogenetic position of other putative species of Marasmiellus. The mating system is tetrapolar.     

ITS phylogeny of West Asian Heracleum species and related taxa of Umbelliferae–Tordylieae W.D.J.Koch, with notes on evolution of their psbA-trnH sequences

Summary Heracleum is a large and taxonomically complex genus of the Umbelliferae–Tordylieae. The phylogenetic relationships of West Asian Heracleum species and related taxa were explored using data from sequences of the internal transcribed spacer (ITS1 and ITS2) region of nuclear ribosomal DNA. The data set consists of 56 species, of which 47 were analyzed for the first time; it represents all subdivisions of the genus Heracleum, as well as some representatives of Pastinaca complex. Heracleum was shown to be a polyphyletic genus, as its species fall into two different clades, one of which comprises also Symphyoloma and Mandenovia. Section Pubescentia was confirmed, in contrast to the sections Villosa and Heracleum being polyphyletic. A separate position of the section Wendia was supported. H. marashicum was shown to be a member of a clade comprising Pastinaca and related genera. The sequences of chloroplast psbA-trnH intergenic spacer, the region recently proposed for DNA barcoding in plants, were also analyzed for 33 species, representing all principal clades within Heracleum and its relatives. They have been proven to be very similar and not suitable for DNA barcoding in this group. However, some sequence variation was revealed. This variation could be explained by the combination of such evolutionary events as inversion and duplication. It was shown that these events are rather common in Tordylieae and can occur independently in different lineages. The evolution patterns of psbA-trnH spacer are hypothesized.     

Nuclear ribosomal ITS sequences and phylogeny in East AsianAconitum subgenusAconitum (Ranunculaceae), with special reference to extensive polymorphism in individual plants

Internal transcribed spacer (ITS) sequences of nuclear ribosomal DNA (nrDNA) were used to examine the phylogeny of East Asian aconites. Individual aconites were discovered to contain as many as eight different ITS sequences after cloning and PCR-SSCP (single-stranded conformational polymorphisms) analysis. We identified eight putative ITS pseudogenes from four taxa with low predicted secondary structure stability and high substitution rates. Maximum likelihood (ML) and neighbor-joining (NJ) methods were used for phylogenetic reconstruction. The ITS trees agree with the previous chloroplast DNA (cpDNA) tree for the vast majority of the taxa. We found two East Asian clades in the ITS trees: 1) a clade with the Chinese diploid,Aconitum volubile and East Asian tetraploids, and 2) a clade of East Asian diploids and Siberian tetraploids. In the former clade, most tetraploid taxa appear to be polyphyletic; sequences from individual plants did not correspond to recognized taxonomic units. This indicates a recent divergence of the East Asian tetraploids.     

Taxonomic and phytogeographic implications from ITS phylogeny in<Emphasis Type="Italic"> Berberis</Emphasis> (Berberidaceae)

A phylogeny based on the internal transcribed spacer (ITS) sequences from 79 taxa representing much of the diversity of Berberis L. (four major groups and 22 sections) was constructed for the first time. The phylogeny was basically congruent with the previous classification schemes at higher taxonomic levels, such as groups and subgroups. A notable exception is the non-monophyly of the group Occidentales of compound-leaved Berberis (previously separated as Mahonia). At lower levels, however, most of previous sections and subsections were not evident especially in simple-leaved Berberis. Possible relationship between section Horridae (group Occidentales) and the simple-leaved Berberis clade implies paraphyly of the compound-leaved Berberis. A well-known South America-Old World (mainly Asia) disjunctive distribution pattern of the simple-leaved Berberis is explained by a vicariance event occurring in the Cretaceous period. The ITS phylogeny also suggests that a possible connection between the Asian and South American groups through the North American species (Berberis canadensis or B. fendleri) is highly unlikely.     

The evolution of Dactylorhiza (Orchidaceae) allotetraploid complex: insights from nrDNA sequences and cpDNA PCR-RFLP data

Sequence data from a portion of the external transcribed spacer (ETS) and from the internal transcribed spacers (ITS1 and ITS2) of 18S-26S nuclear ribosomal DNA were used together with chloroplast DNA PCR-RFLP data to unravel patterns of allotetraploid speciation within the Western European Dactylorhiza polyploid complex. A maximum likelihood tree based on combined ETS and ITS sequences suggests that the Western European Dactylorhiza allotetraploids have evolved by hybridization between four main diploid lineages. Cloned sequences and the topology of the ITS plus ETS tree indicate that the allotetraploid species D. elata, D. brennensis, and D. sphagnicola have originated from the autotetraploid D. maculata together with the diploid D. incarnata, while D. majalis, D. traunsteineri, and D. angustata seem to have evolved by hybridization between the D. fuchsii s.str and D. incarnata lineages. Finally, the diploid D. saccifera lineage seems to have been involved together with the D. incarnata lineage in the formation of the allotetraploid D. praetermissa. The observed congruence between the chloroplast tree and the ITS/ETS tree suggests a directional evolution of the nrDNA after polyploidization in favor of the maternal genome. Considered together with morphological, biogeographical, and ecological evidence, the molecular analysis leads us to recognize four species within the investigated allotetraploid complex, namely D. majalis, D. praetermissa, D. elata, and D. sphagnicola.     

Phylogenetic relationships of Scaphyglottis and related genera (Laeliinae: Orchidaceae) based on nrDNA ITS sequence data

Sequences of the nuclear ribosomal internal transcribed spacer regions 1 & 2 (nrDNA ITS) including the intervening 5.8S region were analyzed cladistically for 43 individuals of 35 species ofScaphyglottis s.l. plus two outgroup taxa. Low levels of sequence divergence do not allow estimation of relationships among most clades, but the analyses indicate that four segregate genera (Hexisea Lindl.,Reichenbachanthus Barb. Rodr.,Hexadesmia Brogn., andPlatyglottis coriacea L.O. Williams) are embedded within a broad paraphyleticScaphyglottis. This broadly definedScaphyglottis sensu Dressler is characterized within Laeliinae by the usual presence of superposed growth habit and the presence of a column foot. In order to accommodate species formerly placed inPlatyglottis andReichenbachanthus, three new combinations are made inScaphyglottis:Scaphyglottis brasiliensis (Schltr.) Dressler,S. coriacea (L. O. Williams) Dressier, andS. emarginata (Garay) Dressler.     

An expanded phylogeny of Cuphea (Lythraceae) and a North American monophyly

A phylogenetic analysis of the New World genus Cuphea was conducted employing sequences from the nuclear rDNA internal transcribed spacer (ITS) and chloroplast trnL-trnF spacer and rpl16 intron. The analysis expands the number of Cuphea species from 53 in an earlier ITS study to 70 and adds two chloroplast data sets in order to generate a more complete and robust phylogeny and to test a previous result that suggested the presence of a large North American clade. Results reaffirm the monophyly of Cuphea with Pleurophora as the sister genus and recover a basal divergence event that mirrors the two subgenera of the current classification. Phylogenies of the two chloroplast regions are largely unresolved beyond the initial dichotomy and some resolution at the terminal and subterminal nodes. Based on the ITS phylogeny, five major clades are recognized. Subgenus Cuphea (Clade 1), defined morphologically by the synapomorphic loss of bracteoles, is sister to the much larger subg. Bracteolatae (Clades 2–5). Clades 2–4, comprising the South American and Caribbean species, grade successively to Clade 5, an exclusively North American lineage of 29 species. Among the 12 sections included in the study, only section Trispermum, a subclade of Clade 4, is monophyletic. Section Pseudocircaea is nested within Clade 3, which is largely equivalent to section Euandra. The North American endemic clade includes four sections, of which none are recovered as monophyletic in this study.