Molecular data delineate four genera of "Thryothorus" wrens

Wrens of the genus Thryothorus comprise over a third of the species diversity in the family Troglodytidae. In addition to this species diversity, these wrens vary in a number of behavioral characteristics, in particular in the presence and structure of vocal duets, which makes them an interesting target for comparative evolutionary ecological and behavioral study. However, no phylogenetic hypothesis for this group-which would provide a sound basis for comparative analysis-is currently available. While previous molecular phylogenetic work established conclusively that the type of this genus, Thryothorus ludovicianus (Latham), was not part of a monophyletic group with other Thryothorus, the exact limits of the genus could not be established due to limited taxon sampling. Here, we present molecular data from all but four currently recognized species of Thryothorus. These data confirm that Thryothorus is paraphyletic, and that the type T. ludovicianus does not form a monophyletic group with any other member of the genus. Based on analyses of our data, we resurrect two previously recognized wren genera, Pheugopedius and Thryophilus, and erect a new genus-Cantorchilus-to house the remaining ex-Thryothorus species. Our hypothesis of relationships will provide a firm basis for future behavioral and morphological analyses of these species.     

Phylogenetic relationships within the leaf-mining flies (Diptera: Agromyzidae) inferred from sequence data from multiple genes

The leaf-mining flies (Diptera: Agromyzidae) are a diverse group whose larvae feed internally in leaves, stems, flowers, seeds, and roots of a wide variety of plant hosts. The systematics of agromyzids has remained poorly known due to their small size and morphological homogeneity. We investigated the phylogenetic relationships among genera within the Agromyzidae using parsimony and Bayesian analyses of 2965 bp of DNA sequence data from the mitochondrial COI gene, the nuclear ribosomal 28S gene, and the single copy nuclear CAD gene. We included 86 species in 21 genera, including all but a few small genera, and spanning the diversity within the family. The results from parsimony and Bayesian analyses were largely similar, with major groupings of genera in common. Specifically, both analyses recovered a monophyletic Phytomyzinae and a monophyletic Agromyzinae. Within the subfamilies, genera found to be monophyletic given our sampling include Agromyza, Amauromyza, Calycomyza, Cerodontha, Liriomyza, Melanagromyza, Metopomyza, Nemorimyza, Phytobia, and Pseudonapomyza. Several genera were found to be polyphyletic or paraphyletic including Aulagromyza, Chromatomyia, Phytoliriomyza, Phytomyza, and Ophiomyia. We evaluate our findings and discuss host-use evolution in light of current agromyzid taxonomy and two recent hypotheses of relationships based on morphological data.     

Phylogenetic relationships, character evolution, and taxonomic implications within the slipper lobsters (Crustacea: Decapoda: Scyllaridae)

The slipper lobsters belong to the family Scyllaridae which contains a total of 20 genera and 89 species distributed across four subfamilies (Arctidinae, Ibacinae, Scyllarinae, and Theninae). We have collected nucleotide sequence data from regions of five different genes (16S, 18S, COI, 28S, H3) to estimate phylogenetic relationships among 54 species from the Scyllaridae with a focus on the species rich subfamily Scyllarinae. We have included in our analyses at least one representative from all 20 genera in the Scyllaridae and 35 of the 52 species within the Scyllarinae. Our resulting phylogenetic estimate shows the subfamilies are monophyletic, except for Ibacinae, which has paraphyletic relationships among genera. Many of the genera within the Scyllarinae form non-monophyletic groups, while the genera from all other subfamilies form well supported clades. We discuss the implications of this history on the evolution of morphological characters and ecological transitions (nearshore vs. offshore) within the slipper lobsters. Finally, we identify, through ancestral state character reconstructions, key morphological features diagnostic of the major clades of diversity within the Scyllaridae and relate this character evolution to current taxonomy and classification.     

A species-level phylogenetic study of the Verbena complex (Verbenaceae) indicates two independent intergeneric chloroplast transfers

Two major impediments to infer plant phylogenies at inter- or intra- species level include the lack of appropriate molecular markers and the gene tree/species tree discordance. Both of these problems require more extensive investigations. One of the foci of this study is examining the phylogenetic utility of a combined chloroplast DNA dataset (>5.0kb) of seven non-coding regions, in comparison with that of a large fragment (ca. 3.0kb) of a low-copy nuclear gene (waxy), in a recent, rapidly diversifying group, the Verbena complex. The complex includes three very closely related genera, Verbena (base chromosome number x=7), Glandularia (x=5), and Junellia (x=10), comprising some 150 species distributed predominantly in South and North America. Our results confirm the inadequacy of non-coding cpDNA in resolving relationships among closely related species due to lack of variation, and the great potential of low-copy nuclear gene as source of variation. However, this study suggests that when both cpDNA and nuclear DNA are employed in low-level phylogenetic studies, cpDNA might be very useful to infer organelle evolutionary history (e.g., chloroplast transfer) and more comprehensively understand the evolutionary history of organisms. The phylogenetic framework of the Verbena complex resulted from this study suggests that Junellia is paraphyletic and most ancestral among the three genera; both Glandularia and Verbena are monophyletic and have been derived from within Junellia. Implications of this phylogenetic framework to understand chromosome number evolution and biogeography are discussed. Most interestingly, the comparison of the cpDNA and nuclear DNA phylogenies indicates two independent intergeneric chloroplast transfers, both from Verbena to Glandularia. One is from a diploid North American Verbena species to a polyploid North American Glandularia species. The other is more ancient, from the South American Verbena group to the common ancestor of a major Glandularia lineage, which has radiated subsequently in both South and North America. The commonly assumed introgressive hybridization may not explain the chloroplast transfers reported here. The underlying mechanism remains uncertain.     

Cytotaxonomy of Commelinaceae: chromosome numbers of some African and Asiatic species

Chromosome counts are reported for 32 taxa (31 species and 1 subspecies) belonging to 10 genera of Commelinaceae from seven African and Asiatic countries. Counts for 13 species and 1 subspecies are recorded for the first time. Published chromosome numbers for Anhicopsis and Polyspatha are confirmed. It is suggested that Pdisota, Pollia and Stanfieldidla each has a single basic number (x = 20, 16 and 11, respectively). The known cytological diversity in Floscopa is extended. The third continental African species of Coleolrype is found to have the same chromosome number (2n = 36) as the other two. The preponderance of the basic number x = 15 in Commelina is supported. The uncommon basic number x = 13 is reported in four taxa of Cyanotis together with karyotypic differences. The basic number x = 6 is found in a second species of Murdannia . Karyotypic data in addition to chromosome numbers are presented for 24 of the 32 taxa investigated. Karyotypes are found to be useful in assessing relationships in the family, and evolutionary trends in the karyotype are noted.     

Brassicales: an update on chromosomal evolution and ancient polyploidy

Brassicales comprise 17 families, c. 400 genera and more than 4600 species. Despite the mustard family (crucifers, Brassicaceae) continuing to be the subject of intensive research, the remaining 16 families are largely under studied. Here I summarize the available data on chromosome number and genome size variation across Brassicales in the context of a robust phylogenetic framework. This analysis has revealed extensive knowledge gaps in karyological data for non-crucifer and species-rich families in particular (i.e., Capparaceae, Cleomaceae, Resedaceae and Tropaeolaceae). A parsimonious interpretation of the combined chromosomal and phylogenetic data set suggests that the ancestral pre-Brassicales genome had 9 or 14 chromosome pairs, later multiplied by the At-β (beta) whole-genome duplication (WGD) to n?=?18 or 28. This WGD was followed by post-polyploid diploidization marked by diversification to 12 or 13 families and independent decreases in chromosome numbers. Family-specific WGDs are proposed to precede the diversification of Capparaceae, Resedaceae and Tropaeolaceae.     

Phylogeny and biogeography of the core babblers (Aves: Timaliidae)

The avian family Timaliidae is a species rich and morphologically diverse component of African and Asian tropical forests. The morphological diversity within the family has attracted interest from ecologists and evolutionary biologists, but systematists have long suspected that this diversity might also mislead taxonomy, and recent molecular phylogenetic work has supported this hypothesis. We produced and analyzed a data set of 6 genes and almost 300 individuals to assess the evolutionary history of the family. Although phylogenetic analysis required extensive adjustment of program settings, we ultimately produced a well-resolved phylogeny for the family. The resulting phylogeny provided strong support for major subclades within the family but extensive paraphyly of genera. Only 3 genera represented by more than 3 species were monophyletic. Biogeographic reconstruction indicated a mainland Asian origin for the family and most major clades. Colonization of Africa, Sundaland, and the Philippines occurred relatively late in the family's history and was mostly unidirectional. Several putative babbler genera, such as Robsonius, Malia, Leonardina, and Micromacronus are only distantly related to the Timaliidae.     

A molecular phylogenetic analysis of the family Rhacophoridae with an emphasis on the Asian and African genera

Using characters from mitochondrial DNA to construct maximum parsimony and maximum likelihood trees, we performed a phylogenetic analysis on representative species of 14 genera: 12 that belong to the treefrog family Rhacophoridae and two, Amolops and Rana, that are not rhacophorids. Our results support a phylogenetic hypothesis that depicts a monophyletic family Rhacophoridae. In this family, the Malagasy genera Aglyptodactylus, Boophis, Mantella, and Mantidactylus form a well-supported sister clade to all other rhacophorid genera, and Mantella is the sister taxon to Mantidactylus. Within the Asian/African genera, the genus Buergeria forms a well-supported clade of four species. The genera, except for Chirixalus, are generally monophyletic. An exception to this is that Polypedates dennysii clusters with species of Rhacophorus, suggesting that the taxonomy of the rhacophorids should be revised to reflect this relationship. Chirixalus is not monophyletic. Unexpectedly, there is strong support for Chirixalus doriae from Southeast Asia forming a clade with species of the African genus Chiromantis, suggesting that Chiromantis dispersed to Africa from Asia. Also, there is strong support for the sister taxon relationship of Chirixalus eiffingeri and Chirixalus idiootocus apart from other congeners.     

基于染色体计数和ITS序列初步探讨横断山区柴胡属植物(伞形科)的系统发育

横断山区是中国柴胡属Bupleurum植物的分布中心。本文对横断山区6个种2变种进行了染色体记数报道,其中4个种2变种是首次报道。对横断山区的10个种4个变种、中国北方（河北和黑龙江）的3个种的nrDNA ITS进行测序,同时从GenBank里面下载同属的来自非洲和地中海西部的16个nrDNA ITS序列数据,结合染色体数目变化结果,初步探讨了横断山区柴胡属植物的系统发育。结果表明横断山区可能是现代柴胡属植物的频度中心和多样分布中心之一。它们的祖先种可能是非洲北部的木本柴胡属植物B.fruticosum,或者是地中海西部的柴胡属植物,推测是通过中东和高加索扩散而形成的,其中与非洲南部特有种B.mundtii的亲缘关系也较近;染色体基数演化趋势是：8是较原始基数,6和7是次生基数,其染色体异基数变异和多倍化可能是物种形成、进化以及向外扩散的主要方式;在ITS系统发育树中,中国柴胡属植物染色体基数为8的种类聚为一支,染色体基数为6和7的种类聚为了一支,不支持舒璞等（1998）关于中国柴胡属的属下分类系统。结合已有的形态学、细胞学、孢粉学证据和ITS系统发育树,建议窄竹叶柴胡B.marginatum var.stenophyllum独立成种。     

Generic delimitation and phylogenetic relationships within the subtribe Chironiinae (Chironieae: Gentianaceae), with special reference to Centaurium: evidence from nrDNA and cpDNA sequences

To better understand the evolutionary history of the genus Centaurium and its relationship to other genera of the subtribe Chironiinae (Gentianaceae: Chironieae), molecular analyses were performed using 80 nuclear ribosomal ITS and 76 chloroplast trnLF (both the trnL UAA intron and the trnL-F spacer) sequences. In addition, morphological, palynological, and phytochemical characters were included to a combined data matrix to detect possible non-molecular synapomorphies. Phylogenetic reconstructions support the monophyly of the Chironiinae and an age estimate of ca. 22 million years for the subtribe. Conversely, both molecular data sets reveal a polyphyletic Centaurium, with four well-supported main clades hereafter treated as separate genera. The primarily Mediterranean Centaurium s.s. is closely related to southern African endemics Chironia and Orphium, and to the Chilean species Centaurium cachanlahuen. The resurrected Mexican and Central American genus Gyrandra is closely related to Sabatia (from eastern North America). Lastly, the monospecific genus Exaculum (Mediterranean) forms a monophyletic group together with the two new genera: Schenkia (Mediterranean and Australian species) and Zeltnera (all other indigenous American centauries). Several biogeographical patterns can be inferred for this group, supporting a Mediterranean origin followed by dispersals to (1) North America, Central America, and South America, (2) southern Africa (including the Cape region), and (3) Australia and Pacific Islands.     

A molecular phylogeny of the fern family Pteridaceae: assessing overall relationships and the affinities of previously unsampled genera

The monophyletic Pteridaceae accounts for roughly 10% of extant fern diversity and occupies an unusually broad range of ecological niches, including terrestrial, epiphytic, xeric-adapted rupestral, and even aquatic species. In this study, we present the results of the first broad-scale and multi-gene phylogenetic analyses of these ferns, and determine the affinities of several previously unsampled genera. Our analyses of two newly assembled data sets (including 169 newly obtained sequences) resolve five major clades within the Pteridaceae: cryptogrammoids, ceratopteridoids, pteridoids, adiantoids, and cheilanthoids. Although the composition of these clades is in general agreement with earlier phylogenetic studies, it is very much at odds with the most recent subfamilial classification. Of the previously unsampled genera, two (Neurocallis and Ochropteris) are nested within the genus Pteris; two others (Monogramma and Rheopteris) are early diverging vittarioid ferns, with Monogramma resolved as polyphyletic; the last previously unsampled genus (Adiantopsis) occupies a rather derived position among cheilanthoids. Interestingly, some clades resolved within the Pteridaceae can be characterized by their ecological preferences, suggesting that the initial diversification in this family was tied to ecological innovation and specialization. These processes may well be the basis for the diversity and success of the Pteridaceae today.     

Molecular phylogenetics of the family Cyprinidae (Actinopterygii: Cypriniformes) as evidenced by sequence variation in the first intron of S7 ribosomal protein-coding gene: further evidence from a nuclear gene of the systematic chaos in the family

The family Cyprinidae is the largest freshwater fish group in the world, including over 200 genera and 2100 species. The phylogenetic relationships of major clades within this family are simply poorly understood, largely because of the overwhelming diversity of the group; however, several investigators have advanced different hypotheses of relationships that pre- and post-date the use of shared-derived characters as advocated through phylogenetic systematics. As expected, most previous investigations used morphological characters. Recently, mitochondrial DNA (mtDNA) sequences and combined morphological and mtDNA investigations have been used to explore and advance our understanding of species relationships and test monophyletic groupings. Limitations of these studies include limited taxon sampling and a strict reliance upon maternally inherited mtDNA variation. The present study is the first endeavor to recover the phylogenetic relationships of the 12 previously recognized monophyletic subfamilies within the Cyprinidae using newly sequenced nuclear DNA (nDNA) for over 50 species representing members of the different previously hypothesized subfamily and family groupings within the Cyprinidae and from other cypriniform families as outgroup taxa. Hypothesized phylogenetic relationships are constructed using maximum parsimony and Basyesian analyses of 1042 sites, of which 971 sites were variable and 790 were phylogenetically informative. Using other appropriate cypriniform taxa of the families Catostomidae (Myxocyprinus asiaticus), Gyrinocheilidae (Gyrinocheilus aymonieri), and Balitoridae (Nemacheilus sp. and Beaufortia kweichowensis) as outgroups, the Cyprinidae is resolved as a monophyletic group. Within the family the genera Raiamas, Barilius, Danio, and Rasbora, representing many of the tropical cyprinids, represent basal members of the family. All other species can be classified into variably supported and resolved monophyletic lineages, depending upon analysis, that are consistent with or correspond to Barbini and Leuciscini. The Barbini includes taxa traditionally aligned with the subfamily Cyprininae sensu previous morphological revisionary studies by Howes (Barbinae, Labeoninae, Cyprininae and Schizothoracinae). The Leuciscini includes six other subfamilies that are mainly divided into three separate lineages. The relationships among genera and subfamilies are discussed as well as the possible origins of major lineages.     

Phylogenetic inference of the genus Bupleurum (Apiaceae) in Hengduan Mountains based on chromosome counts and nuclear ribosomal DNA ITS sequences

pleurum of Apiaceae in China. This paper reports chromosome numbers of six species and two varieties of Bupleurum, and for four species and two varieties their chromosome numbers are reported for the first time. The phylogeny of Bupleurum was investigated based on the ITS region of the nuclear ribosomal DNA (nrDNA) of 14 taxa from the Hengduan Mountains, 3 taxa from the North China (Hebei and Heilongjiang), and 16 taxa from Africa and the Mediterranean region. Varia-tions in chromosome numbers and the ITS sequences were used to infer phylogenetic relationships between Bupleurum species in Hengduan Mountains. The results showed that the Hengduan Mountains might represent one of the frequency and diversity centers for Bupleurum. The ancestors of Bupleurum species in the Hengduan Mountains may be related to the woody B. fruticosum in North Africa, or the species in the western Mediterranean region. It is postulated that the ancestral population migrated into Hengduan Mountains through the Middle East and the Caucasus. Furthermore, the neo-endemic B. mundtii in South Africa appeared to be a close relative of the species in the Hengduan Mountains. In the trend of basic chromosome number evolution, x = 8 should be regarded as the ancestral basic number, while x = 6, 7 as the derived ones. The Bupleurum species in the Hengduan Moun-tains have been undergoing changes in the basic chromosome numbers or the ploidy level. The ITS phylogenetic tree showed that the Chinese species were divided into two clades: one with the basic chromosome number x = 8, and the other with x = 6, 7. The results rejected the previous infrageneric classification of Bupleurum in China. We further suggested to raise B. marginatum var. stenophyllum to species rank based on the combined evidence from morphology, karyology, pollen morphology, and the ITS phylogenetic tree.     

Phylogenetic analyses and chromosome evolution in Convallarieae (Ruscaceae sensu lato), with some taxonomic treatments

The tribe Convallarieae, comprising 10 genera and 95 species, has recently been transferred from its own family to Ruscaceae sensu lato. In this study, sequence data from trnK and rbcL were analyzed for 19 species in 8 genera, and chromosome morphology was analyzed for 17 species in 7 genera. The parsimony analysis of trnK and rbcL sequences showed that Convallarieae are monophyletic. Although early branches did not receive strong bootstrap support, Convallaria diverged at the first branch, followed by Speirantha. The rest of the tribe was split into three, well-supported clades: one with Reineckea, the second with Campylandra and Rohdea, and the third with Tupistra, Tricalistra, and Aspidistra. Two monotypic genera, Rohdea and Tricalistra, were embedded in a clade of Campylandra and of Tupistra, respectively. Three karyotypes were distinguished in the tribe on the basis of the basic number and morphology of metaphase chromosomes: Convallaria type (with x=19 and unimodal chromosome length), Tupistra type (with x=19 and trimodal chromosome length), and Aspidistra-elatior type (with x=18 and trimodal chromosome length). The character-state distribution in the molecular tree showed that the Convallaria type is plesiomorphic, from which was derived the Tupistra type and subsequently the Aspidistra-elatior type. Taxonomic treatments to transfer Campylandra to Rohdea and Tricalistra to Tupistra are also given.     

Phylogenetic relationships in the family Resedaceae L.

A study was made on the phylogenetic relationships of species of the family Resedaceae, based on morphological features, chromosome meiotic behaviour, karyotype features, size and fertility of pollen grains, nucleotypic parameters, seed protein profiles and esterase isozyme patterns.For the comparison of the seed protein profiles among species a method was developed based on the presence or absence of the bands by means of a computer program. The dendrogram obtained by such a method is in line, to a great extent, with the clusters (sections) obtained within the family based on morphological features.Data on meiotic behaviour and on morphology, such as the type of fruit, carpel numbers and others, suggest that x=5 is the primitive basic chromosome number of this family. x₁=6 and x₂=7 are considered as secondary basic numbers derived from x=5 through aneuploid changes.The results support a proposed phylogenetic tree of the genera and sections of the genus Reseda represented in the European Flora.The principal phenomena that have operated in the evolution of the Resedaceae seem to be aneuploid changes, polyploidy and structural rearrangements. A trend towards DNA increase in the evolution of this group is also apparent.     

Phylogeny of the tribe Fumarieae (Papaveraceae s.l.) based on chloroplast and nuclear DNA sequences: Evolutionary and biogeographic implications

? Premise of the Study: Little research has been done at the molecular level on the tribe Fumarieae (Papaveraceae). Papaveraceae is a model plant group for studying evolutionary patterns despite the lack of a reference phylogeny for this tribe. We investigated the phylogenetic relationships within the tribe to complete the molecular data for this family in order to help understand its character evolution and biogeographic pattern. ? Methods: We used maximum-parsimony and Bayesian approaches to analyze five DNA regions for 25 species representing 10 of the 11 Fumarieae genera and five outgroups. Evolutionary pathways of four characters (habit, life span, type of fruit, and number of seeds per fruit) were inferred on the phylogeny using parsimony. The ancestral distribution areas were reconstructed using dispersal-vicariance analysis. ? Key Results: Fumarieae is monophyletic and includes three groups that agree with the morphology-based subtribes: Discocapninae, Fumariinae, and Sarcocapninae. Within subtribes, the relationships among genera were different from those obtained with morphological data. Annual life span, nonchasmophytic habit, and a several-seeded capsule were the basal character states for the tribe. The ancestor occupied a continuous area between West Eurasia and Africa. Vicariances explain the divergence between lineages Discocapninae (South Africa) and Fumariinae-Sarcocapninae (Mediterranean), and the disjunction of Fumariinae (Mediterranean-Central Asia). ? Conclusions: Molecular phylogeny confirms the subtribal classification of Fumarieae based on morphology. However it provides different results regarding the relationships among genera within each subtribe, which affects the inference of the evolutionary pathway followed by the four selected characters. The disjunct distribution of the tribe is explained by different vicariance scenarios.     

A molecular phylogenetic framework for the subfamily Ocenebrinae (Gastropoda,Muricidae)

The Ocenebrinae is a subfamily of marine predatory gastropods known as oyster and mussel drills. Their current phylogenetic framework is traditionally based on shell and radular characters, but a consensus on relationships among genera is still lacking. We investigated the molecular phylogeny of Ocenebrinae using 50 species and DNA data from one nuclear (28S) and two mitochondrial (COI and 16S) genes, the largest data set so far assembled for this subfamily. We found support for the monophyly of the Ocenebrinae, and species were divided into four major lineages. Within groups, genera had similar geographic distributions, suggesting that except in a few cases, species diversification within clades occurred without range expansions. We discuss the phylogenetic distribution of a labral tooth and a sealed siphonal canal, two characteristic ocenebrine features. We also show that Ocinebrina species in the north‐eastern Pacific are not monophyletic with north‐eastern Atlantic and Mediterranean species, and that the Ocinebrina edwardsii species complex belongs to Ocenebra.     

Evolution of Lycopodiaceae from results of sequencing spacer sequences of chloroplast rRNA genes

Nucleotide sequences of a chloroplast rDNA region including 8 bp from the 3' end of 23S rDNA-ITS2-4.5S rDNA-ITS3-5S rDNA-ITS4 (approximately 800 bp) were determined in 25 species of Lycopodiaceae and two species of the genus Isoetes. The rate of molecular evolution of spacers significantly varied in different Lycopsida taxa. A phylogenetic analysis by the neighbor-joining (NJ) method revealed that the family Lycopodiaceae is monophyletic. The topology of phylogenetic trees suggests the isolation of four or probably five genera in family Lycopodiaceae. For these genera, synapomorphic indels were detected. The obtained data were compared with the results of phylogenetic analysis of Lycopsida with regard to other sequences. The relationships of taxa within the family Lycopodiaceae is discussed.     

Molecular phylogenetics of the clover genus (Trifolium--Leguminosae)

Trifolium, the clover genus, is one of the largest genera of the legume family. We conducted parsimony and Bayesian phylogenetic analyses based on nuclear ribosomal DNA internal transcribed spacer and chloroplast trnL intron sequences obtained from 218 of the ca. 255 species of Trifolium, representatives from 11 genera of the vicioid clade, and an outgroup Lotus. We confirm the monophyly of Trifolium, and propose a new infrageneric classification of the genus based on the phylogenetic results. Incongruence between the nrDNA and cpDNA results suggests five to six cases of apparent hybrid speciation, and identifies the putative progenitors of the allopolyploids T. dubium, a widespread weed, and T. repens, the most commonly cultivated clover species. Character state reconstructions confirm 2n=16 as the ancestral chromosome number in Trifolium, and infer a minimum of 19 instances of aneuploidy and 22 of polyploidy in the genus. The ancestral life history is hypothesized to be annual in subgenus Chronosemium and equivocal in subgenus Trifolium. Transitions between the annual and perennial habit are common. Our results are consistent with a Mediterranean origin of the genus, probably in the Early Miocene. A single origin of all North and South American species is hypothesized, while the species of sub-Saharan Africa may originate from three separate dispersal events.