A phylogeny of legumes (Leguminosae) based on analysis of the plastid matK gene resolves many well-supported subclades within the family

Phylogenetic analysis of 330 plastid matK gene sequences, representing 235 genera from 37 of 39 tribes, and four outgroup taxa from eurosids I supports many well-resolved subclades within the Leguminosae. These results are generally consistent with those derived from other plastid sequence data (rbcL and trnL), but show greater resolution and clade support overall. In particular, the monophyly of subfamily Papilionoideae and at least seven major subclades are well-supported by bootstrap and Bayesian credibility values. These subclades are informally recognized as the Cladrastis clade, genistoid sensu lato, dalbergioid sensu lato, mirbelioid, millettioid, and robinioid clades, and the inverted-repeat-lacking clade (IRLC). The genistoid clade is expanded to include genera such as Poecilanthe, Cyclolobium, Bowdichia, and Diplotropis and thus contains the vast majority of papilionoids known to produce quinolizidine alkaloids. The dalbergioid clade is expanded to include the tribe Amorpheae. The mirbelioids include the tribes Bossiaeeae and Mirbelieae, with Hypocalypteae as its sister group. The millettioids comprise two major subclades that roughly correspond to the tribes Millettieae and Phaseoleae and represent the only major papilionoid clade marked by a macromorphological apomorphy, pseudoracemose inflorescences. The robinioids are expanded to include Sesbania and members of the tribe Loteae. The IRLC, the most species-rich subclade, is sister to the robinioids. Analysis of the matK data consistently resolves but modestly supports a clade comprising papilionoid taxa that accumulate canavanine in the seeds. This suggests a single origin for the biosynthesis of this most commonly produced of the nonprotein amino acids in legumes.     

A phylogeny of the chloroplast gene rbcL in the Leguminosae: taxonomic correlations and insights into the evolution of nodulation

Phylogenetic analysis of the chloroplast-encoded rbcL gene in Leguminosae are consistent with previous hypotheses in suggesting that the family as a whole is monophyletic, but that only two of its three subfamilies are natural. The earliest dichotomies in the family appear to have involved tribes Cercideae or Cassieae (subtribe Dialiinae), followed by Detarieae/ Macrolobieae, all of which are members of subfamily Caesalpinioideae. The remainder of the family is divided into two clades: (1) Mimosoideae and the caesalpinioid tribes Caeasalpinieae and Cassieae (subtribes Ceratoniinae and Cassiinae); (2) Papilionoideae. Basal groups within Papilionoideae are, as expected, elements of the grade tribes Sophoreae and Swartzieae. Major clades within Papilionoideae include: (1) a Genistoid Alliance comprising Genisteae, Crotalarieae, Podalyrieae, Thermopsideae, Euchresteae, and also some Sophoreae; (2) a clade marked by the absence of one copy of the chloroplast inverted repeat, with which are associated Robinieae. Loteae, and some Sophoreae; (3) Phaseoleae, Desmodieae. Psoraleeae, and most Millettieae, a group also marked by presence of pseudoracemose inflorescences; and (4) a well-supported clade comprising Aeschynomeneae, Adesmieae, and some Dalbergieae. Nodulation is most parsimoniously optimized on the rbcL strict consensus tree as three parallel gains, occurring in Papilionoideae, the caesalpioioid ancestors of Mimosoideae, and in the genus Chamaecrista (Caesalpinieae: Cassieae).     

A phylogeny of cycads (Cycadales) inferred from chloroplast matK gene, trnK intron, and nuclear rDNA ITS region

Phylogenetic relationships among the three families and 12 living genera of cycads were reconstructed by distance and parsimony criteria using three markers: the chloroplast matK gene, the chloroplast trnK intron and the nuclear ITS/5.8S rDNA sequence. All datasets indicate that Cycadaceae (including only the genus Cycas) is remotely related to other cycads, in which Dioon was resolved as the basal-most clade, followed by Bowenia and a clade containing the remaining nine genera. Encephalartos and Lepidozamia are closer to each other than to Macrozamia. The African genus Stangeria is embedded within the New World subfamily Zamiodeae. Therefore, Bowenia is an unlikely sister to Stangeria, contrary to the view that they form the Stangeriaceae. The generic status of Dyerocycas and Chigua is unsupportable as they are paraphyletic with Cycas and the Zamia, respectively. Nonsense mutations in the matK gene and indels in the other two datasets lend evidence to reinforce the above conclusions. According to the phylogenies, the past geography of the genera of cycads and the evolution of character states are hypothesized and discussed. Within the suborder Zamiieae, Stangeria, and the tribe Zamieae evolved significantly faster than other genera. The matK gene and ITS/5.8S region contain more useful information than the trnK intron in addressing phylogeny. Redelimitations of Zamiaceae, Stangeriaceae, subfamily Encephalartoideae and subtribe Macrozamiineae are necessary.     

Phylogeny of amorpheae (fabaceae: papilionoideae)

The legume tribe Amorpheae comprises eight genera and 240 species with variable floral form. In this study, we inferred a phylogeny for Amorpheae using DNA sequence data from the plastid trnK intron, including matK, and the nuclear ribosomal ITS1, 5.8S, and ITS2. Our data resulted in a well-resolved phylogeny in which the tribe is divided into the daleoids (Dalea, Marina, and Psorothamnus), characterized by generally papilionaceous corollas, and the amorphoids (Amorpha, Apoplanesia, Errazurizia, Eysenhardtia, and Parryella), characterized by non-papilionaceous flowers. We found evidence for the paraphyly of Psorothamnus and for the monophyly of Dalea once D. filiciformis is transferred to monophyletic Marina. Errazurizia rotundata is more closely related to Amorpha than to the other errazurizias, and Eysenhardtia is supported to be monophyletic. The monotypic Parryella and Apoplanesia are placed within the amorphoids. Among Papilionoideae, trnK/matK sequence data provide strong evidence for the monophyly of Amorpheae and place Amorpheae as sister to the recently discovered dalbergioid clade.     

The dalbergioid legumes (Fabaceae): delimitation of a pantropical monophyletic clade

A monophyletic pantropical group of papilionoid legumes, here referred to as the "dalbergioid" legumes, is circumscribed to include all genera previously referred to the tribes Aeschynomeneae and Adesmieae, the subtribe Bryinae of the Desmodieae, and tribe Dalbergieae except Andira, Hymenolobium, Vatairea, and Vataireopsis. This previously undetected group was discovered with phylogenetic analysis of DNA sequences from the chloroplast trnK (including matK) and trnL introns, and the nuclear ribosomal 5.8S and flanking internal transcribed spacers 1 and 2. All dalbergioids belong to one of three well-supported subclades, the Adesmia, Dalbergia, and Pterocarpus clades. The dalbergioid clade and its three main subclades are cryptic in the sense that they are genetically distinct but poorly, if at all, distinguished by nonmolecular data. Traditionally important taxonomic characters, such as arborescent habit, free stamens, and lomented pods, do not provide support for the major clades identified by the molecular analysis. Short shoots, glandular-based trichomes, bilabiate calyces, and aeschynomenoid root nodules, in contrast, are better indicators of relationship at this hierarchical level. The discovery of the dalbergioid clade prompted a re-analysis of root nodule structure and the subsequent finding that the aeschynomenoid root nodule is synapomorphic for the dalbergioids.     

Phylogenetic analysis of Pinguicula (Lentibulariaceae): chloroplast DNA sequences and morphology support several geographically distinct radiations

The genus Pinguicula is one of the three genera of the carnivorous Lentibulariaceae, comprising approximately 80 species. Phylogeny inference using nucleotide sequences of the chloroplast gene matK and the trnK group II intron, as well as a set of 32 morphological characters revealed five well-supported, major lineages within the genus. These lineages largely reflect radiations in clearly defined geographic regions, whereas most previously recognized sections of the genus are shown to be para- or polyphyletic. A species-rich Mexican-Central American-Caribbean clade has the Eurasian P. alpina and an East Asian clade as successive sisters. All three are characterized by a production of flower buds on winter-resting plants, a specific corolla hair structure and a very large corolla lower central lobe. Another diverse clade is composed of species with primarily European distribution including the widespread type species P. vulgaris. For this clade, vegetative reproduction during dormancy is synapomorphic. Species native to SE North America and the South American Andes and a group of Mediterranean and NE Atlantic coast species together appear in a fifth well-supported clade, that is characterized by a tropical-type growth habit. It is the only clade that has reached temperate zones of the southern hemisphere.     

Occurrence of matK in a trnK group II intron in charophyte green algae and phylogeny of the Characeae

A group II intron containing the matK gene, which encodes a splicing-associated maturase, was found in the trnK (lysine tRNA) exon in the chloroplast genome of the six extant genera of green algae in the family Characeae, which among green algae are the sister group to embryophytes (land plants). The characean trnK intron (～2.5 kilobases [kb]) and matK ORF (～1.5 kb) are comparable in size to the intron and ORF of land plants, in which they are similarly found inserted in the trnK exon. Domain X, a sequence of conserved amino acid residues within matK, occurs in the Characeae. Phylogenetic analysis using maximum likelihood (GTR + I + gamma likelihood model) and parsimony (branch and bound search) yielded one tree with high bootstrap support for all branches. The matK tree was congruent with the rbcL tree for the same taxa. The number and proportion of informative sites was higher in matK (501, 31% of matK sequence) compared to rbcL (122, 10%). Characeae branch lengths were on average more than five times longer for matK compared to rbcL and provided better resolution within the Characeae. These findings along with recent genomic analyses demonstrate that the intron and matK invaded the chloroplast genome of green algae prior to the evolution of land plants.     

Evolution of Piperales--matK gene and trnK intron sequence data reveal lineage specific resolution contrast

Piperales represent the largest basal angiosperm order with a nearly worldwide distribution. The order includes three species rich genera, Piper (ca. 2000 species), Peperomia (ca. 1500-1700 species), and Aristolochia s. l. (ca. 500 species). Sequences of the matK gene and the non-coding trnK group II intron are analysed for a dense set of 105 taxa representing all families (except Hydnoraceae) and all generic segregates (except Euglypha within Aristolochiaceae) of Piperales. A large number of highly informative indels are found in the Piperales trnK/matK dataset. Within a narrow region approximately 500 nt downstream in the matK coding region (CDS), a length variable simple sequence repeat (SSR) expansion segment occurs, in which insertions and deletions have led to short frame-shifts. These are corrected shortly afterwards, resulting in a maximum of six amino acids being affected. Furthermore, additional non-functional matK copies were found in Zippelia begoniifolia, which can easily be discriminated from the functional open reading frame (ORF). The trnK/matK sequence data fully resolve relationships within Peperomia, whereas they are not effective within Piper. The resolution contrast is correlated with the rate heterogeneity between those lineages. Parsimony, Bayesian and likelihood analyses result in virtually the same topology, and converge on the monophyly of Piperaceae and Saururaceae. Lactoris gains high support as sister to Aristolochiaceae subf. Aristolochioideae, but the different tree inference methods yield conflicting results with respect to the relationships of subfam. Asaroideae. In Piperaceae, a clade formed by the monotypic genus Zippelia and the small genus Manekia (=Sarcorhachis) is sister to the two large genera Piper and Peperomia.     

Phylogenetic signal in matK vs. trnK: a case study in early diverging eudicots (angiosperms)

The matK gene has been among the most useful loci for resolving plant phylogenetic relationships at different evolutionary time-scales, but much less is known about the phylogenetic utility of the flanking trnK intron, especially for deep level phylogenetics. We compared the relative performance of matK and trnK intron regions for resolving the relationships of the early diverging eudicots (angiosperms). The two regions display similar nucleotide compositions and distributions of rate variation among sites. The trnK intron sequences also provide similar levels of phylogenetic information per-site as matK. Combining the trnK intron sequences with matK increases overall bootstrap support for the early diverging eudicots compared to analyses of matK alone. MP, ML and Bayesian analyses provide strong support for eudicots, the sister group relationship of Ranunculales to remaining eudicots, and a Buxales+Trochodendraceae+core eudicots clade. matK and the trnK intron support conflicting positions for Buxales and Trochodendrales in relation to the core eudicots.     

The Linderniaceae and Gratiolaceae are further lineages distinct from the Scrophulariaceae (Lamiales)

The Lamiales are one of the largest orders of angiosperms, with about 22,000 species. The Scrophulariaceae, as one of their most important families, has recently been shown to be polyphyletic. As a consequence, this family was re-classified and several groups of former scrophulariaceous genera now belong to different families, such as the Calceolariaceae, Plantaginaceae, or Phrymaceae. In the present study, relationships of the genera Craterostigma, Lindernia and its allies, hitherto classified within the Scrophulariaceae, were analyzed. Sequences of the chloroplast trnK intron and the matK gene (approximately 2.5 kb) were generated for representatives of all major lineages of the Lamiales and the former Scrophulariaceae. Bayesian and parsimony analyses revealed two isolated lineages, one of which consists of Lindernia and its allies, the other of Gratiola and allies. Gratiola was previously assumed to be related to Lindernia and was therefore included here. It is proposed to treat the two clades as separate families, Linderniaceae and Gratiolaceae. For the Linderniaceae, several morphological synapomorphies exist in addition to molecular data, such as conspicuous club-shaped stamen appendages.     

Infrafamilial Phylogeny of the Aquatic Angiosperm Podostemaceae Inferred from the Nucleotide Sequences of the matK Gene

Abstract: The infrafamilial relationships of Podostemaceae were deduced from nucleotide sequences of the chloroplast matK gene. The matK phylogenetic analyses show that Podostemaceae are composed of two major clades that correspond to the subfamily Tristichoideae sensu stricto and Weddellina and the subfamily Podostemoideae. Weddellina, which has long been recognized as a member of the Tristichoideae, is sister to the Podostemoideae, supporting the classification that recognized a third subfamily Weddellinoideae. Malaccotristicha malayana and Terniopsis sessilis form a basal clade in Tristichoideae sensu stricto. Tristichoideae show a high morphological diversity and, surprisingly, a close relationship exists between Dalzellia zeylanica and Indotristicha ramosissima, which remarkably differ in their body plans. A few genera defined by particular characters, such as Synstylis and Torrenticola, merge into clades of other larger genera. The Podostemoideae taxa studied are composed of two American clades, an Asian-Australian clade and a Madagascan clade, and may suggest that the subfamily perhaps originated in America and migrated to the Old World.     

Generic phylogeny and historical biogeography of Alismataceae, inferred from multiple DNA sequences

Alismataceae is an aquatic or semi-aquatic herb family with a subcosmopolitan distribution. The family is one of the oldest lineages within monocots and plays an important role in the systematics, biogeography and evolutionary processes of flowering plants. However, the generic relationships of the family are still a subject of debate, and its historical biogeography is less studied. In the present study, we carried out a comprehensive phylogenetic analysis based on multiple DNA sequences (nuclear: ITS; chloroplast: psbA, rbcL, matK, rpoB, rpoC1, trnK 5' intron and trnK 3' intron; mitochondria: cob and atp1). The result supports merging Limnocharitaceae into Alismataceae as one family. Two well-supported clades were obtained based on the combined ITS, psbA, rbcL and matK dataset. Clade B consists of Luronium, Damasonium, Baldellia and Alisma; and clade A consists of the remaining genera of Alismataceae as well as Limnocharitaceae. Biogeographic analysis and bayesian molecular dating suggested that Alismataceae originated in West Palearctic or Afrotropical area during the Late Cretaceous, and subsequently split into two clades. Clade A and clade B diversified in Afrotropical area and West Palearctic area, respectively. The intercontinental distribution of this family mainly resulted from dispersals involving migration across land bridges and long-distance dispersal.     

Molecular systematic studies on the tribe Polygonateae (s. l.) in China based on RFLPs data of PCR-amplified chloroplast DNA fragments

Phylogenetic relationships in the tribe Polygonateae s. l. of Liliaceae were investigated with an RFLP analysis of two PCR-amplified chloroplast genome DNA fragments. One fragment is the transfer RNA gene for lysine( trnK gene) including the maturase-encoding gene matK, and the other is the rpl16 gene which codes for a chloroplast ribosomal protein in large subunit. The trnK gene is ca. 2600 bp in length in all the taxa, but the rpl16 gene ranges from ca. 1140 bp to ca. 1320 bp in length among different genera and in Polygonatum. The results suggested that the tribe Polygonateae s. str. consisting of Polygonatum, Disporopsis, Smilacina and Maianthemum is closely related to Convallaria in Convallarieae, and supported the earlier results that Streptopus and Disporum should be removed out from the tribe Polygonateae s. l.. The results also showed that Polygonatum formed a clad with Disporopsis, while Smilacina lumped with Maianthemum in the tribe Polygonateae s. str.. In addition, the present study supports the view that S. ginfoshanicumshould be transferred from Smilacina to Polygonatum.     

Broad screening of the legume family for variability in seed insecticidal activities and for the occurrence of the A1b-like knottin peptide entomotoxins

Pea albumin 1b (PA1b) is a small sulphur-rich peptide from pea seeds, also named leginsulin because of the binding characteristics of its soybean orthologue. Its insecticidal properties were discovered more recently. By using a combination of molecular, biochemical and specific insect bioassays on seed extracts, we characterised genes from numerous Papilionoideae, but not from Caesalpinioideae or Mimosoideae, although the last group harboured species with partially positive cues (homologous biological activities). The A1b defence peptide family, therefore, appears to have evolved relatively late in the legume lineage, maybe from the sophoroid group (e.g. Styphnolobium japonicum). However, unambiguous sequence information is restricted to a group of tribes within the subfamily Papilionoideae (Psoraleae, Millettieae, Desmodieae, Hedysareae, Phaseoleae, Vicieae, and the now clearly polyphyletic "Trifolieae" and "Galegeae"). Recent diversification by gene duplications has occurred in many species, or longer ago in some lineages (Medicago truncatula), as well as probable gene or expression losses at different taxonomic levels (Loteae, Vigna subterranea).     

Phylogeny reconstruction in the Caesalpinieae grade (Leguminosae) based on duplicated copies of the sucrose synthase gene and plastid markers

The Caesalpinieae grade (Leguminosae) forms a morphologically and ecologically diverse group of mostly tropical tree species with a complex evolutionary history. This grade comprises several distinct lineages, but the exact delimitation of the group relative to subfamily Mimosoideae and other members of subfamily Caesalpinioideae, as well as phylogenetic relationships among the lineages are uncertain. With the aim of better resolving phylogenetic relationships within the Caesalpinieae grade, we investigated the utility of several nuclear markers developed from genomic studies in the Papilionoideae. We cloned and sequenced the low copy nuclear gene sucrose synthase (SUSY) and combined the data with plastid trnL and matK sequences. SUSY has two paralogs in the Caesalpinieae grade and in the Mimosoideae, but occurs as a single copy in all other legumes tested. Bayesian and maximum likelihood phylogenetic analyses suggest the two nuclear markers are congruent with plastid DNA data. The Caesalpinieae grade is divided into four well-supported clades (Cassia, Caesalpinia, Tachigali and Peltophorum clades), a poorly supported clade of Dimorphandra Group genera, and two paraphyletic groups, one with other Dimorphandra Group genera and the other comprising genera previously recognized as the Umtiza clade. A selection analysis of the paralogs, using selection models from PAML, suggests that SUSY genes are subjected to a purifying selection. One of the SUSY paralogs, under slightly stronger positive selection, may be undergoing subfunctionalization. The low copy SUSY gene is useful for phylogeny reconstruction in the Caesalpinieae despite the presence of duplicate copies. This study confirms that the Caesalpinieae grade is an artificial group, and highlights the need for further analyses of lineages at the base of the Mimosoideae.     

RRNA and dnaK relationships of Bradyrhizobium sp. nodule bacteria from four papilionoid legume trees in Costa Rica

Enzyme electrophoresis and sequencing of rRNA and dnaK genes revealed high genetic diversity among root nodule bacteria from the Costa Rican trees Andira inermis, Dalbergia retusa, Platymiscium pinnatum (Papilionoideae tribe Dalbergieae) and Lonchocarpus atropurpureus (Papilionoideae tribe Millettieae). A total of 21 distinct multilocus genotypes [ETs (electrophoretic types)] was found among the 36 isolates analyzed, and no ETs were shared in common by isolates from different legume hosts. However, three of the ETs from D. retusa were identical to Bradyrhizobium sp. isolates detected in prior studies of several other legume genera in both Costa Rica and Panama. Nearly full-length 16S rRNA sequences and partial 23S rRNA sequences confirmed that two isolates from D. retusa were highly similar or identical to Bradyrhizobium strains isolated from the legumes Erythrina and Clitoria (Papilionoideae tribe Phaseoleae) in Panama. rRNA sequences for five isolates from L. atropurpureus, P. pinnatum and A. inermis were not closely related to any currently known strains from Central America or elsewhere, but had affinities to the reference strains Bradyrhizobium japonicum USDA 110 (three isolates) or to B. elkanii USDA 76 (two isolates). A phylogenetic tree for 21 Bradyrhizobium strains based on 603 bp of the dnaK gene showed several significant conflicts with the rRNA tree, suggesting that genealogical relationships may have been altered by lateral gene transfer events.     

A Molecular Phylogeny of Costaceae (Zingiberales)

The phylogenetic relationships of Costaceae, a tropical monocotyledonous family sister to the gingers (Zingiberaceae), were investigated with a combination of two chloroplast loci (the trnL-F locus, including the trnL intron, the 3'trnL exon, and the trnL-F intergenic spacer, and the trnK locus, including the trnK intron and the matK coding region) and one nuclear locus (ITS1-5.8s-ITS2). The resulting parsimony analysis of selected taxa that demonstrate the range of floral morphological variation in the family shows that the Cadalvena-type [corrected] floral morphology is ancestral to the group and that both Tapeinochilos species and a Monocostus + Dimerocostus clade represent recent divergences. The genus Costus is broadly paraphyletic but Costus subgenus Eucostus K. Schum. represents a large monophyletic radiation that is poorly resolved. Within this clade, secondary analyses suggest that pollination syndrome, traditionally used for taxonomic and classification purposes within the genus Costus, is a relatively plastic trait of limited phylogenetic utility. This represents the first detailed investigation into intrageneric and interspecific evolutionary relationships within the family Costaceae and presents some novel evolutionary trends with respect to floral morphology and biogeography.     

Evolution of carnivory in Lentibulariaceae and the Lamiales

As a basis for analysing the evolution of the carnivorous syndrome in Lentibulariaceae (Lamiales), phylogenetic reconstructions were conducted based on coding and non-coding chloroplast DNA (matK gene and flanking trnK intron sequences, totalling about 2.4 kb). A dense taxon sampling including all other major lineages of Lamiales was needed since the closest relatives of Lentibulariaceae and the position of "proto-carnivores" were unknown. Tree inference using maximum parsimony, maximum likelihood, and Bayesian approaches resulted in fully congruent topologies within Lentibulariaceae, whereas relationships among the different lineages of Lamiales were only congruent between likelihood and Bayesian optimizations. Lentibulariaceae and their three genera (Pinguicula, Genlisea, and Utricularia) are monophyletic, with Pinguicula being sister to a Genlisea-Utricularia clade. Likelihood and Bayesian trees converge on Bignoniaceae as sister to Lentibulariaceae, albeit lacking good support. The "proto-carnivores" (Byblidaceae, Martyniaceae) are found in different positions among other Lamiales but not as sister to the carnivorous Lentibulariaceae, which is also supported by Khishino-Hasegawa tests. This implies that carnivory and its preliminary stages ("proto-carnivores") independently evolved more than once among Lamiales. Ancestral states of structural characters connected to the carnivorous syndrome are reconstructed using the molecular tree, and a hypothesis on the evolutionary pathway of the carnivorous syndrome in Lentibulariaceae is presented. Extreme DNA mutational rates found in Utricularia and Genlisea are shown to correspond to their unusual nutritional specialization, thereby hinting at a marked degree of carnivory in these two genera.