Clarification of the relationship beteen Apiaceae and Araliaceae based on matK and rbcL sequence data

Apiaceae and Araliaceae (Apiales) represent a particularly troublesome example of the difficulty in understanding evolutionary relationships between tropical-temperate family pairs. Previous studies based on rbcL sequence data provided insights at higher levels, but were unable to resolve fully the family-pair relationship. In this study, sequence data from a more rapidly evolving gene, matK, was employed to provide greater resolution. In Apiales, matK sequences evolve an average of about two times faster than rbcL sequences. Results of phylogenetic analysis of matK sequences were first compared to those obtained previously from rbcL data; the two data sets were then combined and analyzed together. Molecular analyses confirm the polyphyly of apiaceous subfamily Hydrocotyloideae and suggest that some members of this subfamily are more closely related to Araliaceae than to other Apiaceae. The remainder of Apiaceae forms a monophyletic group with well-defined subclades corresponding to subfamilies Apioideae and Saniculoideae. Both the matK and the combined rbcL-matK analyses suggest that most Araliaceae form a monophyletic group, including all araliads sampled except Delarbrea and Mackinlaya. The unusual combination of morphological characters found in these two genera and the distribution of matK and rbcL indels suggest that these taxa may be the remnants of an ancient group of pro-araliads that gave rise to both Apiaceae and Araliaceae. Molecular data indicate that the evolutionary history of the two families is more complex than simple derivation of Apiaceae from within Araliaceae. Rather, the present study suggests that there are two well-defined "families," both of which may have been derived from a lineage (or lineages) or pro-araliads that may still have extant taxa.     

The phylogenetic potential of entire 26S rDNA sequences in plants

18S ribosomal RNA genes are the most widely used nuclear sequences for phylogeny reconstruction at higher taxonomic levels in plants. However, due to a conservative rate of evolution, 18S rDNA alone sometimes provides too few phylogenetically informative characters to resolve relationships adequately. Previous studies using partial sequences have suggested the potential of 26S or large-subunit (LSU) rDNA for phylogeny retrieval at taxonomic levels comparable to those investigated with 18S rDNA. Here we explore the patterns of molecular evolution of entire 26S rDNA sequences and their impact on phylogeny retrieval. We present a protocol for PCR amplification and sequencing of entire (approximately 3.4 kb) 26S rDNA sequences as single amplicons, as well as primers that can be used for amplification and sequencing. These primers proved useful in angiosperms and Gnetales and likely have broader applicability. With these protocols and primers, entire 26S rDNA sequences were generated for a diverse array of 15 seed plants, including basal eudicots, monocots, and higher eudicots, plus two representatives of Gnetales. Comparisons of sequence dissimilarity indicate that expansion segments (or divergence domains) evolve 6.4 to 10.2 times as fast as conserved core regions of 26S rDNA sequences in plants. Additional comparisons indicate that 26S rDNA evolves 1.6 to 2.2 times as fast as and provides 3.3 times as many phylogenetically informative characters as 18S rDNA; compared to the chloroplast gene rbcL, 26S rDNA evolves at 0.44 to 1.0 times its rate and provides 2.0 times as many phylogenetically informative characters. Expansion segment sequences analyzed here evolve 1.2 to 3.0 times faster than rbcL, providing 1.5 times the number of informative characters. Plant expansion segments have a pattern of evolution distinct from that found in animals, exhibiting less cryptic sequence simplicity, a lower frequency of insertion and deletion, and greater phylogenetic potential.      

Phylogenetic assessment of the Branchiobdellidae (Annelida, Clitellata) using 18S rDNA, mitochondrial cytochrome c oxidase subunit I and morphological characters

Using 18S rDNA, mitochondrial cytochrome c oxidase subunit I and morphological characters, the Branchiobdellidae (Annelida, Clitellata) were shown to form a monophyletic group distinct from the leeches using two distant 'oligochaetes' as outgroups. The study used 20 branchiobdellid species from 14 genera in four subfamilies with these representing each of the taxon's three distributional regions in the Holarctic realm. No monophyletic groups were found using the gene sequence data that related to either geographical regions or currently recognized subfamilies. However, two monophyletic groups were strongly supported; the two European species of Branchiobdella and the combination of Sathodrilus attenuatus and Xironogiton victoriensis . The latter pair is taxonomically diverse, but sympatric on Signal crayfish, Pacifastacus leniusculus , in California, USA.     

Molecular phylogenetics of Phyllanthaceae: evidence from plastid MATK and nuclear PHYC sequences

Plastid matK and a fragment of the low-copy nuclear gene PHYC were sequenced for 30 genera of Phyllanthaceae to evaluate tribal and generic delimitation. Resolution and bootstrap percentages obtained with matK are higher than that of PHYC, but both regions show nearly identical phylogenetic patterns. Phylogenetic relationships inferred from the independent and combined data are congruent and differ from previous, morphology-based classifications but are highly concordant with those of the plastid gene rbcL previously published. Phyllanthaceae is monophyletic and gives rise to two well-resolved clades (T and F) that could be recognized as subfamilies. DNA sequence data for Keayodendron and Zimmermanniopsis are presented for the first time. Keayodendron is misplaced in tribe Phyllantheae and belongs to the Bridelia alliance. Zimmermanniopsis is sister to Zimmermannia. Phyllanthus and Cleistanthus are paraphyletic. Savia and Phyllanthus subgenus Kirganelia are not monophyletic.     

The phylogenetic history of Selaginellaceae based on DNA sequences from the plastid and nucleus: extreme substitution rates and rate heterogeneity

Molecular phylogenetic research on Selaginellaceae has focused on the plastid gene rbcL, which in this family has unusually high substitution rates. Here we develop a molecular data set from the nuclear 26S ribosomal DNA gene with the aim of evaluating and extending the results of previous phylogenetic research. The 26S rDNA and the rbcL regions were sequenced for a sample of 23 species, which represent the main elements of species diversity in the family. The data were analysed independently and in combination using both maximum parsimony and Bayesian inference. Although several between genome differences were found, the general pattern of relationships uncovered by all analyses was very similar. Results corroborate the previous study supporting new groupings not previously recognised on morphological grounds. Substitution rates in the 26S rDNA were also found to be high (26% informative) for the region analysed, but lower than for rbcL (37% informative). These data indicate that high substitution rates might be widespread in all three genomes (i.e., plastid, mitochondrion, and nucleus).     

The circumscription and phylogenetic relationships of Callitropsis and the newly described genus Xanthocyparis (Cupressaceae)

A new species of conifer was recently discovered in northern Vietnam. In a preliminary phylogenetic analysis of morphological data a possible sister species, Chamaecyparis nootkatensis (D. Don) Spach, was identified; however, because of the presumed phylogenetic remoteness of these two species to the remainder of the Cupressaceae, a new genus-Xanthocyparis-was described to accommodate both species. Here an analysis of ITS (nrDNA), matK, and rbcL sequence data in combination with 58 informative morphological characters was aimed at testing the monophyly of the remainder of Chamaecyparis and evaluating the placement and monophyly of Xanthocyparis. Chamaecyparis, minus C. nootkatensis, was resolved as a monophyletic group, remote from Cupressus and Xanthocyparis. Cupressus, Juniperus, and Xanthocyparis formed a very highly supported monophyletic group. However, Cupressus was not monophyletic. Instead the Old World species sampled were resolved sister to a clade containing a monophyletic Juniperus, a monophyletic Xanthocyparis, and a clade of New World Cupressus species. If both species of Xanthocyparis are to be treated as members of the same genus, then due to the principal of priority they will have to be recognized in the genus Callitropsis. Research is continuing to resolve the status of New World and Old World Cupressus.     

Phylogenetic relationships within Serpulidae (Sabellida, Annelida) inferred from molecular and morphological data

We assessed phylogenetic relationships within Serpulidae (including Spirorbinae) using parsimony and Bayesian analyses of 18S rDNA, the D1 and D9−D10 regions of 28S rDNA, and 38 morphological characters. In total, 857 parsimony informative characters were used for 31 terminals, 29 serpulids and sabellid and sabellariid outgroups. Following ILD assessment the two sequence partitions and morphology were analysed separately and in combination. The morphological parsimony analysis was congruent with the results of the 2003 preliminary analysis by Kupriyanova in suggesting that a monophyletic Serpulinae and Spirorbinae form a clade, while the remaining serpulids form a basal grade comprising what are normally regarded as Filograninae. Bremer support values were, however, quite low throughout. In contrast, the combined analyses of molecular and morphological data sets provided highly resolved and well-supported trees, though with some conflict when compared to the morphology-only analysis. Spirorbinae was recovered as a sister group to a monophyletic group comprising both 'filogranin' taxa ( Salmacina , Filograna , Protis , and Protula ) and 'serpulin' taxa such as Chitinopoma , Metavermilia , and Vermiliopsis . Thus the traditionally formulated subfamilies Serpulinae and Filograninae are not monophyletic. This indicates that a major revision of serpulid taxonomy is needed at the more inclusive taxonomic levels. We refrain from doing so based on the present analyses since we feel that further taxon sampling and molecular sequencing are required. The evolution of features such as the operculum and larval development are discussed.     

Phylogenetic relationships of Cornaceae and close relatives inferred from matK and rbcL sequences

Phylogenetic relationships were inferred using nucleotide sequences of the chloroplast gene matK for members of Cornales, a well-supported monophyletic group comprising Cornaceae and close relatives. The shortest trees resulting from this analysis were highly concordant with those based on previous phylogenetic analysis of rbcL sequences. Analysis of a combined matK and rbcL sequence data set (a total of 2652 bp [base pairs]) provided greater resolution of relationships and higher internal support for clades compared to the individual data sets. Four major clades (most inclusive monophyletic groups) of Cornales are indicated by both sets of genes: (1) Cornus-Alangium, (2) nyssoids (Nyssa-Davidia-Camptotheca)- mastixioids (Mastixia, Diplopanax), (3) Curtisia, and (4) Hydrangeaceae-Loasaceae. The combined evidence indicates that clades 2 and 3 are sisters, with clade 4 sister to the remainder of Cornales. These relationships are also supported by other lines of evidence, including synapomorphies in fruit and pollen morphology and gynoecial vasculature. Comparisons of matK and rbcL sequences based on one of the most parsimonious rbcL-matK trees indicate that matK has a much higher A-T content (66.9% in matK vs. 55.8% in rbcL) and a lower transition:transversion ratio (1.23 in matK vs. 2.21 in rbcL). The total number of nucleotide substitutions per site for matK is 2.1 times that of rbcL in Cornales. These findings are similar to recent comparisons of matK and rbcL in other dicots. Variable sites of matK are almost evenly distributed among the three codon positions (1.0:1.0:1.3), whereas variable sites of rbcL are mostly at the third position (1.8:1.0 :7.5). Among- lineages rates of nucleotide substitutions in rbcL are basically homogeneous throughout Cornales, but are more heterogeneous in matK.     

Molecular phylogeny of Kalyptorhynchia (Rhabdocoela,Platyhelminthes) inferred from ribosomal sequence data

With 556 species described to date, Kalyptorhynchia includes about one‐third of all species of rhabdocoel flatworms. In this study, we present the first molecular phylogenetic analysis of this taxon. The final analysis comprises 110 species. These represent 11 of the 17 known families. The largest family (241 known species), Polycystididae, is represented by nine of 10 subfamilies and 33 of the 58 genera. Sequence data from 18S rDNA and 28S rDNA were analysed using maximum likelihood and Bayesian approaches. Of the two taxa traditionally recognised within Kalyptorhynchia, Eukalyptorhynchia and Schizorhynchia, only Schizorhynchia is monophyletic. All eukalyptorhynch families, except Cicerinidae, are monophyletic. On the other hand, two of the three schizorhynch families included are not monophyletic. Within Polycystididae, the traditional taxonomy was not reflected in our phylogenetic analyses and most subfamilies are polyphyletic. These results suggest that current classification, mostly based on characters of the genital system, suffers from homoplasy. Where possible, a revised classification, taking into account these new findings, is given.     

基于28S rDNA D2基因片段与形态特征的优茧蜂亚科系统发育研究

本研究选取优茧蜂亚科Euphorinae(膜翅目Hymenoptera:茧蜂科Braconidae)的8族19属23种作为内群,茧蜂其它6个亚科的8属8种作外群,首次结合同源核糖体28S rDNA D2基因序列片段和41个形态学特征对该亚科进行了系统发育学研究。利用"圆口类"的内茧蜂亚科Rogadinae、茧蜂亚科Braconinae、矛茧蜂亚科Doryctinae的3个亚科为根,以PAUP*4.0和MrBayes3.0B4软件分别应用最大简约法(MP)和贝叶斯法对优茧蜂亚科的分子数据和分子数据与非分子数据的结合体进行了分析;并以PAUP*4.0对优茧蜂亚科的28S rDNA D2基因序列的片段的碱基组成与碱基替代情况进行了分析。结果表明:优茧蜂亚科的28S rDNA D2基因序列片段的GC%含量在40.00%~49.25%之间变动,而对于碱基替代情况来讲,优茧蜂亚科各个成员间序列变异位点上颠换(transversion)大于转换(transition);不同的分析和算法所产生的系统发育树都表明目前根据形态定义出的优茧蜂亚科Euphorinae不是一个单系群,而是一个与蚁茧蜂亚科Neoneurinae和高腹茧蜂亚科Cenocoelinae混杂在一起的并系群;在优茧蜂亚科内部,悬茧蜂族Meterorini和食甲茧蜂族Microctonini(排除猎户茧蜂属Orionis)为单系群,而宽鞘茧蜂族Centistini、大颚茧蜂族Cosmophorini、优茧蜂族Euphorini、瓢虫茧蜂族Dinocampini为并系群;悬茧蜂族Meterorini在优茧蜂亚科Euphorinae内位于基部位置的观点得到部分的支持,同时食甲茧蜂族Microctonini被判定为相对进化的类群。此外对于优茧蜂亚科内各属之间的相互亲缘关系,不同算法所得到的系统发育属的结果不完全一致,这表明优茧蜂亚科内(属及族)的系统发育关系还有待于进一步研究。     

Molecular phylogenetic analysis of Passifloraceae sensu lato (Malpighiales) based on plastid and nuclear DNA sequences

A phylogenetic analysis of Passifloraceae sensu lato was performed using rbcL, atpB, matK, and 18S rDNA sequences from 25 genera and 42 species. Parsimony analyses of combined data sets resulted in a single most parsimonious tree, which was very similar to the 50% majority consensus tree from the Bayesian analysis. All nodes except three were supported by more than 50% bootstrap. The monophyly of Passifloraceae s.l. as well as the former families, Malesherbiaceae, Passifloraceae sensu stricto, and Turneraceae were strongly supported. Passifloraceae s.s. and the Turneraceae are sisters, and form a strongly supported clade. Within Passifloraceae s.s., the tribes Passifloreae and Paropsieae are both monophyletic. The intergeneric relationships within Passifloraceae s.s. and Turneraceae are roughly correlated with previous classification systems. The morphological character of an androgynophore/gynophore is better used for characterizing genera grouping within Passifloraceae s.s. Other morphological characters such as the corona and aril are discussed.     

Prospects for discriminating Zingiberaceae species in India using DNA barcodes

We evaluated nine plastid(matK, rbcL, rpoC1, rpoB,rpl36-rps8, ndhJ, trnL-F, trnH-psbA, accD) and two nuclear(ITS and ITS2) barcode loci in family Zingiberaceae by analyzing 60 accessions of 20 species belonging to seven genera from India.Bidirectional sequences were recovered for every plastid locus by direct sequencing of polymerase chain reaction(PCR)amplicons in all the accessions tested. However, only 35(58%)and 40 accessions(66%) yielded ITS and ITS2 sequences,respectively, by direct sequencing. In different bioinformatics analyses, matK and rbcL consistently resolved 15 species(75%)into monophyletic groups and five species into two paraphyletic groups. The 173 ITS sequences, including 138 cloned sequences from 23 accessions, discriminated only 12 species(60%), and the remaining species were entered into three paraphyletic groups. Phylogenetic and genealogic analyses of plastid and ITS sequences imply the possible occurrence of natural hybridizations in the evolutionary past in giving rise to species paraphyly and intragenomic ITS heterogeneity in the species tested. The results support using matK and rbcL loci for barcoding Zingiberaceae members and highlight the poor utility of ITS and the highly regarded ITS2 in barcoding this family, and also caution against proposing ITS loci for barcoding taxa based on limited sampling.     

5 The origin of the dinoflagellate plastid

The Dasycladales is an ancient order of tropical benthic marine green algae, unique in their radially arranged unicellular thalli and well-preserved fossil record due to extensive calcification of the thallus. The inference of an accurate phylogeny for the Dasycladales is important in order to better understand stratigraphy, character evolution, and classification. Previous analyses ( rbc L and 18S rDNA) suggested that the Family Acetabulariaceae is monophyletic, but that the Family Dasycladaceae is a basal paraphyletic assemblage. However, the two data sets disagreed regarding genus- and species-level relationships within the Dasycladales. For example, the placement of the genera, Halicoryne , Bornetella and Cymopolia were incongruent. Given the conflicting results of these previous analyses, the current project examined a third highly conserved nuclear-encoded gene, 26S rDNA. Aligned 26S rDNA sequences were analyzed with parsimony and model-based methods and compared to previous results based on18S and rbc L sequences. Family-level relationships based on 26S rDNA were congruent with previous studies: the Acetabulariaceae is monophyletic while the Dasycladaceae is paraphyletic. In addition, acetabulariacean genera are not monophyletic, suggesting that the presence of a corona inferior or calcification of gametes may not be appropriate to define genera. Within the Dasycladaceae, the basal position of Cymopolia is supported by 26S rDNA, a result congruent with rbcL and stratigraphy but not with 18S data. These results will be discussed in the context of morphological character evolution, fossil stratigraphy and family, tribal and generic relationships among these living algal fossils. Supported in part by NSF grant DEB-0128977 to FWZ.     

Phylogeny and biogeography of Juglans (Juglandaceae) based on matK and ITS sequence data

We investigated phylogenetic and biogeographic relationships within Juglans (walnuts), a Tertiary disjunct genus, using 15 species of Juglans and related (Juglandaceae) outgroups. The relationships were analyzed using nucleotide sequences of the chloroplast gene matK and its flanking spacers and of the internal transcribed spacers (ITS) and 5.8S gene of the nuclear ribosomal DNA. The DNA sequences provided 246 informative characters for parsimony analysis. ITS data supported as monophyletic groups the four generic sections, Cardiocaryon, Dioscaryon, Rhysocaryon, and Trachycaryon. Within Rhysocaryon, the temperate black walnuts and the tropical black walnuts were supported as monophyletic groups. When the two data sets were combined, J. cinerea was nested within Cardiocaryon. Combined analysis with published nuclear DNA restriction site data placed J. cinerea in a monophyletic group with Cardiocaryon. These analyses consistently supported Juglans as a monophyletic group and as the sister group to the genus Pterocarya. The results of this work are consistent with the known geological history of Juglans. The fossil record suggests that the butternuts had evolved by the early Oligocene in North America. The presence of butternuts in Eurasia could be the result of migration from North America to Eurasia during the warming trend of the mid Oligocene.     

Phylogeny and evolution of the Betulaceae as inferred from DNA sequences,morphology, and paleobotany

Phylogeny of the Betulaceae is assessed on the basis of rbcL, ITS, and morphological data. Based upon 26 rbcL sequences representing most “higher” hamamelid families, the Betulaceae are monophyletic, with Casuarinaceae as its sister group, regardless of whether the outgroup is Cunoniaceae, Cercidiphyllaceae, Hamamelidaceae, or Nothofagus. Within the Betulaceae, two sister clades are evident, corresponding to the subfamilies Betuloideae and Coryloideae. However, with only 13 phylogenetically informative sites, the rbcL sequences provide limited intra-subfamilial resolution. Internal transcribed spacer (ITS) sequences provided 96 phylogenetically informative sites from 491 aligned sites resulting in a single most parsimonious tree of 374 steps (consistency index = 0.791) with two major lineages corresponding to the two traditional subfamilies: Betuloideae (Alnus, Betula) and Coryloideae (Corylus, Ostryopsis, Carpinus, Ostrya). This arrangement is mostly consistent with those from rbcL and morphology and is greatly reinforced by analyses with the three data sets combined. In the Coryloideae, the Ostryopsis–Carpinus–Ostrya clade is well supported, with Corylus as its sister group. The sister-group relationship between Ostryopsis and the Carpinus–Ostrya clade is well supported by ITS, rbcL, and morphological data. Phylogenetic relationships among the extant genera deduced by these analyses are compatible with inferences from ecological evolution and the extensive fossil record.