Phylogenetic relationships within Chamaecrista sect. Xerocalyx (Leguminosae, Caesalpinioideae) inferred from the cpDNA trnE-trnT intergenic spacer and nrDNA ITS sequences

Chamaecrista belongs to subtribe Cassiinae (Caesalpinioideae), and it comprises over 330 species, divided into six sections. The section Xerocalyx has been subjected to a profound taxonomic shuffling over the years. Therefore, we conducted a phylogenetic analysis using a cpDNA trnE-trnT intergenic spacer and nrDNA ITS/5.8S sequences from Cassiinae taxa, in an attempt to elucidate the relationships within this section from Chamaecrista. The tree topology was congruent between the two data sets studied in which the monophyly of the genus Chamaecrista was strongly supported. Our analyses reinforce that new sectional boundaries must be defined in the Chamaecrista genus, especially the inclusion of sections Caliciopsis and Xerocalyx in sect. Chamaecrista, considered here paraphyletic. The section Xerocalyx was strongly supported as monophyletic; however, the current data did not show C. ramosa (microphyllous) and C. desvauxii (macrophyllous) and their respective varieties in distinct clades, suggesting that speciation events are still ongoing in these specimens.     

Phylogenetic relationships and biogeographic patterns in North American members of Carex section Acrocystis (Cyperaceae) using nrDNA ITS and ETS sequence data

Carex section Acrocystis currently includes 27 taxa in North America. Recent phylogenetic studies have suggested that the North American and some but not all of the Eurasian species form a clade. Relationships and biogeographic patterns among species in this core-Acrocystis group are explored here using nuclear ribosomal (nrDNA) internal transcribed spacer region (ITS) and nrDNA external transcribed spacer region (ETS) sequence data. While maximum parsimony analysis of the ITS and ETS data provides only a moderately resolved branching structure for species relationships within the core-Acrocystis clade, maximum likelihood analysis provides a more resolved hypothesis of relationships in the section. The core-Acrocystis clade consists of a grade of Eurasian and primarily western North American species, with a well-supported clade of only eastern North American species nested within this grade. ITS and ETS types do not coalesce within many species or species complexes. Possible explanations for the non-coalescent nature of ITS and ETS copies in Acrocystis are explored, including lineage sorting, hybridization, and cryptic species.     

Molecular phylogeny of Phalaenopsis Blume (Orchidaceae) based on the internal transcribed spacer of the nuclear ribosomal DNA

The internal transcribed spacer (ITS1, 5.8S rDNA, and ITS2) region of nuclear ribosomal DNA (nrDNA) was sequenced from 53 species, which represent most of the living species diversity in the genus Phalaenopsis (Orchidaceae). A phylogeny was developed for the genus based on the neighbor-joining and maximum parsimony analyses of molecular data. Results of these analyses provided support for the monophyly of the genus Phalaenopsis and concurred in that the genera Doritis and Kingidium should be treated as being parts of the genus Phalaenopsis as suggested by Christenson (2001). Within the genus Phalaenopsis, neither subgenera Aphyllae nor Parishianae were monophyletic, and they were highly clustered with subgenus Proboscidioides plus sections Esmeralda and Deliciosae of subgenus Phalaenopsis based on ITS data. Those species also have the same characters of morphology of four pollinia and similar biogeographies. Furthermore, neither subgenus Phalaenopsis nor Polychilos was monophyletic. Within the subgenus Phalaenopsis, only section Phalaenopsis was highly supported as being monophyletic. As for the subgenus Polychilos, only section Polychilos was moderately supported as being monophyletic. In conclusion, the present molecular data obtained from the ITS sequence of nrDNA of the genus Phalaenopsis provide valuable information for elucidating the phylogeny of this genus.     

Unexpected high divergence in nrDNA ITS and extensive parallelism in floral morphology of Pedicularis (Orobanchaceae)

Internal transcribed spacer (ITS) region sequences of nuclear ribosomal DNA (nrDNA) were used in the phylogenetic reconstruction of Pedicularis, a genus with strong adaptive radiation. Forty-two species representing 12 greges of the genus were all, except P. resupinata, sampled from the Hengduan Mountain region, China. A high level of ITS sequence variation was found among the species distributed in such a small geographical area, which had been rarely reported in other groups. The great ITS divergence in Pedicularis could be explained by a relatively ancient origin and diversification of the genus followed by migration of different floristic components into the Hengduan Mountains, or accelerated rates of molecular evolution in parasitic lineages of Orobanchaceae. In the present ITS phylogeny, almost all the main clades are not consistent with the high hierarchical taxa within Pedicularis, which implies that significant parallel evolution occurred in floral morphology of the genus, and that undue attention has been paid to corolla characters in the intragenus classifications.We thank the two anonymous reviewers for valuable comments and suggestions; Professor Guo You-Hao, Drs. Tan Dun-Yan, Luo Yi-Bo, Bao Ying, Luo Yan, Mr. Mao Jian-Feng and Mr. Qin Wei for their great help in the field collection; Drs. Song Bao-Hua, Wei Xiao-Xin and Miss Sun Ying-Xue for their help in the laboratory work. This study was supported by State Key Basic Research and Development Plan (Grant No. G2000046804) and the National Natural Science Foundation of China (Grant No. 30121003).     

Phylogenetic relationships among subgenera,species, and varieties of Japanese <Emphasis Type="Italic">Salvia</Emphasis> L. (Lamiaceae)

To determine evolutionary relationships among all Japanese members of the genus Salvia (Lamiaceae), we conducted molecular phylogenetic analyses of two chloroplast DNA (cpDNA) regions (rbcL and the intergenic spacer region of trnL−trnF:trnL−trnF) and one nuclear DNA (nrDNA) region (internal transcribed spacer, ITS). In cpDNA, nrDNA, and cpDNA+nrDNA trees, we found evidence that all Japanese and two Taiwanese Salvia species are included in a clade with other Asian Salvia, and Japanese Salvia species were distributed among three subclades: (1) S. plebeia (subgenus Sclarea), (2) species belonging to subg. Salvia, and (3) species belonging to subg. Allagospadonopsis. At the specific level our findings suggest: a close relationship between S. nipponica and S. glabrescens, no support for monophyly of S. lutescens and its varieties in cpDNA, nrDNA and cpDNA+nrDNA trees, and that S. pygmaea var. simplicior may be more closely related to S. japonica than to other varieties of S. pygmaea.     

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.     

Species-specific molecular signature of Commiphora species of Saudi Arabia inferred from internal transcribed spacer sequences of nuclear ribosomal DNA

The deciduous habit and tendency to produce flowers prior to developing leaves, and a predominantly dioecious system of breeding in the genus Commiphora leads to difficulties in its taxonomic identification at species level. The characteristics of easy amplification by universal primer, shorter length and higher discrimination power at the species level makes the internal transcribed spacer (ITS) sequence of nuclear ribosomal DNA (nrDNA) to a smart gene for generating species-specific phylogenetic inferences in most of the plants groups. The present study deals the ITS sequence of nrDNA based molecular genotyping of seven species of the genus Commiphora of Saudi Arabia. The molecular phylogenetic analysis of ITS sequences of nrDNA of Commiphora species distributed in Saudi Arabia reveals the the occurrence of C. madagascariens in Saudi Arabia.     

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.     

Testing and using complete plastomes and ribosomal DNA sequences as the next generation DNA barcodes in Panax (Araliaceae)

Complete plastid genome (plastome) sequences and nuclear ribosomal DNA (nrDNA) regions have been proposed as candidates for the next generation of DNA barcodes for plant species discrimination. However, the efficacy of this approach still lacks comprehensive evaluation. We carried out a case study in the economically important but phylogenetically and taxonomically difficult genus Panax (Araliaceae). We generated a large data set of plastomes and nrDNA sequences from multiple accessions per species. Our data improved the phylogenetic resolution and levels of species discrimination in Panax, compared to any previous studies using standard DNA barcodes. This provides new insights into the speciation, lineage diversification and biogeography of the genus. However, both plastome and nrDNA failed to completely resolve the phylogenetic relationships in the Panax bipinnatifidus species complex, and only half of the species within it were recovered as monophyletic units. The results suggest that complete plastome and ribosomal DNA sequences can substantially increase species discriminatory power in plants, but they are not powerful enough to fully resolve phylogenetic relationships and discriminate all species, particularly in evolutionarily young and complex plant groups. To gain further resolving power for closely related species, the addition of substantial numbers of nuclear markers is likely to be required.     

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.     

Molecular phylogenetics of Hippophae L. (Elaeagnaceae) based on the internal transcribed spacer (ITS) sequences of nrDNA

 The genus Hippophae comprises 7 species and 8 subspecies according to the latest classification, and has shown enormous ecological, nutrient and medicinal values. Here we analyzed the phylogenetic relationships among 15 taxa of the genus by comparing sequences of the internal transcribed spacer (ITS) region of nuclear ribosomal DNA (nrDNA). ITS sequences in Hippophae varied in length from 651 bp to 666 bp. The aligned sequences were 690 bp in length and 269 (39.0%) were variable sites with 150 being parsimony-informative. The amount of polymorphism observed within a taxon was extremely low in most taxa except for two putative hybrid species. The aligned sequences were analyzed by maximum parsimony (MP) and neighbor-joining (NJ) methods. In the strict consensus trees of parsimony analysis, the monophyly of Hippophae was supported by 100% bootstrap value. H. tibetana was at the basal position of the genus, and the remaining taxa formed two clades with high bootstrap support. The first clade included subspecies of H.␣rhamnoides and the other one consisted of remaining species. Parsimony analysis also suggested that the species H. tibetana, H. neurocarpa and H.␣salicifolia were all distinct. Although the sequence divergence among subspecies of H. rhamnoides was also remarkably high, the molecular data supported the monophyly of H. rhamnoides when H. rhamnoides subsp. gyantsensis Rousi was excxluded. The NJ trees showed essentially the same topology. The taxonomical arrangement that divided the genus into two sections was not supported based on the ITS sequences. However, the hybrid origin of H. goniocarpa and H. litangensis proposed previously was supported by the present ITS data. Received January 7, 2002; accepted May 10, 2002 Published online: November 22, 2002 Addresses of the authors: Kun Sun, Xuelin Chen, Ruijun Ma, Qin Wang, Institute of Botany, Northwest Normal University, Lanzhou 730070, China. Changbao Li, Song Ge (e-mail: gesong@ns.ibcas.ac.cn or song_ge@hotmail.com), Laboratory of Systematic and Evolutionary Botany, Institute of Botany, Chinese Academy of Sciences, Beijing 100093, China.     

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.     

Phylogenetic analysis of the internal transcribed spacer region of Japanese Lilium species

 The DNA from 16 Lilium species and one variety endemic to or naturalized in Japan were obtained and their internal transcribed spacer regions of nuclear ribosomal DNA (nrDNA) were amplified by PCR and sequenced by cycle sequencing. Phylogenetic analysis of the ITS sequences supported the validity of Comber’s classification system. It has also provided molecular evidence for the transfer of Lilium dauricum to sect. Sinomartagon. The phylogenetic relationships revealed by ITS DNA analysis were supported by previously published crossability data. The molecular phylogeny of Japanese Lilium species was discussed with reference to the putative migration routes of these species. Received: 30 June 1998 / Accepted: 19 October 1998     

MONOPHYLY OF ANEUPLOID ASTRAGALUS (FABACEAE): EVIDENCE FROM NUCLEAR RIBOSOMAL DNA INTERNAL TRANSCRIBED SPACER SEQUENCES

Evolutionary relationships within Astragalus L. (Fabaceae) were inferred from nucleotide sequence variation in nuclear ribosomal DNA of both New World and Old World species. The internal transcribed spacer regions (ITS) of 18S–26S nuclear ribosomal DNA from representatives of 26 species of Astragalus, three species of Oxytropis DC., and two outgroup taxa were analyzed by polymerase chain reaction amplification and direct DNA sequencing. The length of the ITS 1 region within these taxa varied from 221 to 231 bp, while ITS 2 varied in length from 207 to 217 bp. Of the aligned, unambiguous positions, approximately 34% were variable in each spacer region. In pairwise comparisons among Astragalus species and outgroup taxa, sequence divergence at these sites ranged from 0 to 18.8% in ITS 1 and from 0 to 21.7% in ITS 2. Parsimony analyses of these sequences resulted in a well-resolved phylogeny that is highly concordant with previous cytogenetic and chloroplast DNA evidence for a major phylogenetic division in the genus. These data suggest that the New World aneuploid species of Astragalus form a monophyletic but morphologically cryptic group derived from euploid species of Old World (Eurasian) origin, which are consequently paraphyletic.     

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.     

Molecular phylogenetics of the sexually deceptive orchid genus Ophrys (Orchidaceae) based on nuclear and chloroplast DNA sequences.

We present a phylogenetic analysis of the major lineages of the sexually deceptive orchid genus Ophrys based on nuclear ribosomal (nr) DNA (internal transcribed spacer region) and noncoding chloroplast (cp) DNA (trnL-trnF region) sequences. Sequence divergence within and among major Ophrys lineages was low for both nrDNA and cpDNA sequences. Separate analyses resulted in similar but poorly resolved trees. An incongruence length difference test revealed that nrDNA and cpDNA data sets were not incongruent. A combined analysis resulted in a better-resolved phylogenetic hypothesis of relationships among the major Ophrys lineages. Our data strongly support a division of Ophrys into two groups. These groups do not correspond to the earlier proposed sections Euophrys and Pseudophrys and are thus in conflict with traditional classifications. Our results support a well-resolved monophyletic group that contains the geographically widespread O. bombyliflora, O. speculum, O. tenthredinifera, and the O. fusca-lutea lineage. Relationships in the other group are poorly resolved. Based on our observations that taxa with identical sequences at presumably rapidly evolving loci clearly differ in floral morphology, we hypothesize that the diversity in the genus Ophrys is the result of a recent radiation in this orchid lineage.     

The phylogenetic position of an <Emphasis Type="Italic">Armillaria</Emphasis> species from Amami-Oshima,a subtropical island of Japan,based on elongation factor and ITS sequences

An undetermined Armillaria species was collected on Amami-Oshima, a subtropical island of Japan. The phylogenetic position of the Armillaria sp. was determined using sequences of the elongation factor-1α (EF-1α) gene and the internal transcribed spacer (ITS) region (ITS1-5.8S-ITS2) of ribosomal DNA (rDNA). The phylogenetic analyses based on EF-1α and ITS sequences showed that this species differs from known Japanese taxa of Armillaria. The sequences of this species and A. novae-zelandiae from Southeast Asia were contained in a strongly supported clade, which was adjacent to a well-supported sister clade containing A. novae-zelandiae from Australia and New Zealand.     

衣藻属的系统发育分析——基于形态形状和nrDNA ITS序列

通过实验分析莱茵衣藻 ( Chlamydomonas reinhardtii) 1个种和互连网获得衣藻属 1 5个种及丝藻属 1个种 ( Ulothrix zonata) ,共 1 7个种的 nr DNA ITS序列 ,并以 U.zonata为外类群 ,采用计算机分析软件包对其进行分析及构建分子系统发育树图。同时以 1 2个传统分类性状 ,对此 1 6种衣藻构建数据矩阵 ;以 U.zonata动孢子的相应性状为外类群原始性状 ,用Wagner法在计算机上对其进行分枝分析 ;然后比较并分析分子系统树和表征性状分支分析树的异同。初步尝试以 ITS分子序列系统发育分析作为传统性状分析的补充来研究衣藻种间的亲缘关系。     

Primula jiugongshanensis sp. nov. (Primulaceae) from China,based on morphological and molecular evidence

Primula jiugongshanensis J. W. Shao, a new species from Mt Jiugongshan in Hubei Province, China, is described and illustrated. This species is a biennial glabrous herb with pinnately compound leaves, and affiliated to P. sect. Ranunculoides C. M. Hu. It is distinguished from other species of the section by outer simple leaves ovate, flowers distylous, corolla lobe apex conspicuously emarginate, and lack of bulblets. Molecular phylogenetic analysis of nrDNA internal transcribed spacer sequences (ITS) was conducted and the results also supported the recognition of P. jiugongshanensis as a new species. The species is known from only three separated unprotected populations and is categorized as ‘Endangered’ (EN).