Origin and evolution of North American polyploid Silene (Caryophyllaceae)

Nuclear DNA sequences from introns of the low-copy nuclear gene family encoding the second largest subunit of RNA polymerases and the ribosomal internal transcribed spacer (ITS) regions, combined with the psbE-petL spacer and the rps16 intron from the chloroplast genome were used to infer origins and phylogenetic relationships of North American polyploid Silene species and their closest relatives. Although the vast majority of North American Silene species are polyploid, which contrasts to the diploid condition dominating in other parts of the world, the phylogenetic analyses rejected a single origin of the North American polyploids. One lineage consists of tetraploid Silene menziesii and its diploid allies. A second lineage, Physolychnis s.l., consists of Arctic, European, Asian, and South American taxa in addition to the majority of the North American polyploids. The hexaploid S. hookeri is derived from an allopolyploidization between these two lineages. The tetraploid S. nivea does not belong to any of these lineages, but is closely related to the European diploid S. baccifera. The poor resolution within Physolychnis s.l. may be attributed to rapid radiation, recombination among homoeologues, homoplasy, or any combination of these factors. No extant diploid donors could be identified in Physolychnis s.l.     

Polyploid origins in a circumpolar complex in Draba (Brassicaceae) inferred from cloned nuclear DNA sequences and fingerprints

Polyploid evolution has been of major importance in the arctic flora, but rarely addressed on the full circumpolar scale. Herein we study the allopolyploid Draba lactea and its close allies, which form a taxonomically intricate arctic-alpine complex including diploids, tetraploids, and hexaploids. Based on samples from the entire circumpolar area, we inferred the origins of polyploids in this complex using cloned DNA sequences from two nuclear regions (one intron from a gene encoding a second largest subunit in the RNA polymerase family, RPD2, and the ribosomal internal transcribed spacer region, ITS) and DNA fingerprints (random amplified polymorphic DNAs, RAPDs). Although D. lactea and all other polyploids examined in Draba are genetic alloploids showing fixed heterozygosity, the data obtained in the present study suggest that each of the polyploids analyzed here may have originated from a single diploid lineage: hexaploid D. lactea via tetraploid D. lactea from the D. palanderiana lineage (not from the D. fladnizensis and D. nivalis lineages as previously hypothesized), the tetraploid D. turczaninovii from the D. fladnizensis lineage, the tetraploid D. porsildii from the D. lonchocarpa lineage, and a tetraploid here named Draba spB from the D. nivalis lineage. Draba lactea has probably originated several times in the Beringian area, and it is not necessary to invoke complex origins based on a combination of different species lineages as previously suggested.     

Molecular evidence for hybrid origins of the invasive polyploids Carthamus creticus and C. turkestanicus (Cardueae, Asteraceae)

The hexaploids Carthamus creticus and C. turkestanicus are noxious weeds with wide but non-overlapping Mediterranean distributions, and C. creticus, together with another polyploid, C. lanatus, have also invaded similar climatic regions in North and South America, South Africa and Australia. Here we infer their origins using sequences of the plastid intergenic spacer trnH-psbA and the intron trnK and three introns of nuclear low-copy genes of the RNA Polymerase family (RPD2 and the duplicated RPC2), as well as RAPD markers (random amplified polymorphic DNA). Phylogenetic analyses of the nuclear introns and additivity analysis of the RAPD markers support the hypotheses that the two hexaploids are allopolyploids sharing a tetraploid progenitor lineage represented by the broadly Mediterranean C. lanatus, combined with different diploid progenitor lineages consistent with the different geographic distributions of the hexaploids. Whereas C. leucocaulos from the south-eastern Greek Islands represents the diploid progenitor lineage of the western C. creticus, the Irano-Turanian C. glaucus represents the diploid progenitor lineage of the eastern C. turkestanicus. The plastid data suggest that the diploid lineages served as the maternal progenitors of the hexaploids.     

Evolution of parental ITS regions of nuclear rDNA in allopolyploid Aegilops (Poaceae) species

The genus Aegilops comprises approximately 25 diploid, tetraploid and hexaploid species, in which the genome types of all allopolyploids involve either U or D genome, or both of them. The internal transcribed spacer (ITS) region of 18S-26S nuclear ribosomal DNA (rDNA) from 11 allopolyploid species and 7 related diploid species in the genus were directly sequenced by pooled PCR products. Phylogenetic analyses for tracing evolutionary patterns of parental rDNA in allopolyploid species were performed using the neighbor-joining method. The D genome involved tree included three clades (CC-DDCC, DDMM-DDMMSS-DDMMUU, and MM-MhMh-DDNN), but did not include Ae. squarrosa (DD). It indicated that the rDNA of ancestral D genome had been somewhat differentiated in allopolyploids. The U genome involved tree showed that the allopolyploids and their common ancestor, Ae. umbellulata, formed a clade, suggesting that rDNA in UUMM and UUSS genomes has been homogenizing toward that of ancestral U genome. The phylogenetic pattern of U genome based on ITS sequences also supported the "pivotal-differential" hypothesis.     

Polyploid speciation inHedera (Araliaceae): Phylogenetic and biogeographic insights based on chromosome counts and ITS sequences

Variation in chromosome number and internal transcribed sequences (ITS) of nrDNA is used to infer phylogenetic relationships of a wide range ofHedera species. Polyploidy was found to be frequent inHedera, with diploid, tetraploid, hexaploid and octoploid populations being detected. Nucleotide additivity occurs in the ITS sequences of one tetraploid (H. hibernica) and two hexaploid species (H. maderensis, H. pastuchovii), suggesting that all three species originated by allopolyploidisation. ITS sequence polymorphism and nucleotide characters may indicate the presence of an ancient genome persistent only in some allopolyploid species. Phylogenetic analyses of ITS sequence data reveal two lineages ofHedera: one containing all sequences belonging to extant diploids plus the tetraploidH. algeriensis, and a second that includes this ancient ITS type and others exclusive to several polyploid species. The origin of the polyploids is evaluated on the basis of morphology, chromosome counts, ITS sequence polymorphism, and phylogenetic analyses. Reconstruction of reticulate evolution inHedera agrees with two allopolyploid areas on both sides of the Mediterranean basin. Morphological, molecular and cytological evidence also suggests an active dispersal ofHedera populations that may account for three independent introductions in Macaronesia.     

Ancient allopolyploid speciation in Geinae (Rosaceae): evidence from nuclear granule-bound starch synthase (GBSSI) gene sequences

A nuclear low-copy gene phylogeny provides strong evidence for the hybrid origin of seven polyploid species in Geinae (Rosaceae). In a gene tree, alleles at homologous loci in an allopolyploid species are expected to be sisters to orthologues in the ancestral taxa rather than to each other. Alleles at a duplicated locus in an autopolyploid, however, are expected to be more closely related to each other than they are to any orthologous copies in closely related species. We cloned and sequenced about 1.9 kilobases from the 5' end of the GBSSI-1 gene from two diploid, one tetraploid, and six hexaploid species. Each of the three loci in the hexaploid species forms a separate group, two of which are more closely related to copies in other species than they are to each other. This finding indicates that the hexaploid lineage evolved through two consecutive allopolyploidization events. Based on the GBSSI-1 gene tree, we hypothesized that there was an initial hybridization between a diploid species from the ancestral lineage of Coluria and Waldsteinia and an unknown diploid species to form the tetraploid Geum heterocarpum lineage. Backcrossing of G. heterocarpum with a representative of the unknown diploid lineage then resulted in a hexaploid lineage that has radiated considerably since its origin, comprising at least 40 extant species with various morphologies. A penalized likelihood analysis indicated that Geinae may be about 17 million years old, implying that the hypothesized allopolyploid speciation events are relatively ancient. Six of the 22 cloned Geinae GBSSI-1 copies in this study, which all are duplicate copies in polyploid taxa, may have become pseudogenes. We compared the GBSSI-1 phylogeny with one from chloroplast data and explored implications for the evolution of some fruit characters.     

Polyploids with different origins and ancestors form a single sexual polyploid species

Polyploidization is one of the few mechanisms that can produce instantaneous speciation. Multiple origins of tetraploid lineages from the same two diploid progenitors are common, but here we report the first known instance of a single tetraploid species that originated repeatedly from at least three diploid ancestors. Parallel evolution of advertisement calls in tetraploid lineages of gray tree frogs has allowed these lineages to interbreed, resulting in a single sexually interacting polyploid species despite the separate origins of polyploids from different diploids. Speciation by polyploidization in these frogs has been the source of considerable debate, but the various published hypotheses have assumed that polyploids arose through either autopolyploidy or allopolyploidy of extant diploid species. We utilized molecular markers and advertisement calls to infer the origins of tetraploid gray tree frogs. Previous hypotheses did not sufficiently account for the observed data. Instead, we found that tetraploids originated multiple times from extant diploid gray tree frogs and two other, apparently extinct, lineages of tree frogs. Tetraploid lineages then merged through interbreeding to result in a single species. Thus, polyploid species may have complex origins, especially in systems in which isolating mechanisms (such as advertisement calls) are affected directly through hybridization and polyploidy.     

Duplication of floral regulatory genes in the Lamiales

Duplication of some floral regulatory genes has occurred repeatedly in angiosperms, whereas others are thought to be single-copy in most lineages. We selected three genes that interact in a pathway regulating floral development conserved among higher tricolpates (LFY/FLO, UFO/FIM, and AP3/DEF) and screened for copy number among families of Lamiales that are closely related to the model species Antirrhinum majus. We show that two of three genes have duplicated at least twice in the Lamiales. Phylogenetic analyses of paralogs suggest that an ancient whole genome duplication shared among many families of Lamiales occurred after the ancestor of these families diverged from the lineage leading to Veronicaceae (including the single-copy species A. majus). Duplication is consistent with previous patterns among angiosperm lineages for AP3/DEF, but this is the first report of functional duplicate copies of LFY/FLO outside of tetraploid species. We propose Lamiales taxa will be good models for understanding mechanisms of duplicate gene preservation and how floral regulatory genes may contribute to morphological diversity.     

Molecular phylogeny of the Southeast Asian freshwater fish family Botiidae (Teleostei: Cobitoidea) and the origin of polyploidy in their evolution

The freshwater fish family Botiidae is represented by seven genera on the Indian subcontinent and in East and Southeast Asia and includes diploid as well as evolutionary tetraploid species. We present a phylogeny of Botiidae including 33 species representing all described genera using the mitochondrial cytochrome b and 12s rRNA genes to reconstruct the phylogenetic relationships among the genera and to estimate the number of polyploidisation events during their evolution. Our results show two major lineages, the subfamilies Leptobotiinae with the genera Leptobotia and Parabotia and Botiinae with the genera Botia, Chromobotia, Sinibotia, Syncrossus, and Yasuhikotakia. Our results suggest that two species that were traditionally placed into the genus Yasuhikotakia form a monophyletic lineage with the species of Sinibotia. A review of the data on the ploidy level of the included species shows all diploid species to belong to Leptobotiinae and all tetraploid species to Botiinae. A single polyploidisation event can therefore be hypothesised to have occurred in the ancestral lineage leading to the Botiinae.     

Molecular phylogeny and reticulate origins of the polyploid Bromus species from section Genea (Poaceae)

The origin of polyploid Bromus species of section Genea was investigated using molecular data. This group of annual species native from the Old-World is composed of three diploids, two tetraploids, one hexaploid, and one octoploid. Molecular cloning, sequencing, and phylogenetic analyses were performed on several accessions per species. We used the low copy nuclear gene Waxy, repeated rDNA spacers ITS1 and ITS2 and chloroplast spacers trnT-trnL and trnL-trnF. Our analyses revealed four different lineages involved in the parentage of the polyploids and confirmed their reticulate origin. Three of these lineages are closely related to the diploid species B. sterilis, B. tectorum, and B. fasciculatus. The fourth lineage could not be related to any diploid according to the available data. Our data gave insights on the origin of all the polyploids of section Genea, and chloroplast data allowed us to identify the maternal lineages. The Waxy gene was the most informative regarding origin of the polyploids. The Waxy copies duplicated by polyploidy appear selectively maintained in the polyploid species. No sequence heterogeneity was encountered in the ITS region, where concerted evolution seems to have occurred toward either maternal or paternal repeats. These results provide new information about the origin and molecular evolution of these polyploids and will allow a more accurate taxonomic treatment of the concerned species, based on their evolutionary history.     

Systematic Evaluation of Isoëtes asiatica Makino (Isoëtaceae) based on AFLP, nrITS, and Chloroplast DNA Sequences

On the basis of amplified fragment length polymorphism (AFLP) and nucleotide sequence data from nuclear ribosomal internal transcribed spacer (nrITS) and three chloroplast DNA regions (rbcL, cpITS, and trnS-psbC spacer), we investigated the species delimitation and the evolutionary lineage of Isoëtes asiatica from Hokkaido, Japan. The neighbor-joining (NJ) dendrogram based on AFLP markers revealed the well-defined clusters (bootstrap value?=?100%) of I. asiatica. Results from the principal component analysis are largely congruent with those obtained in the NJ dendrogram. The maximum parsimony analysis, based on data from nrITS and three chloroplast DNA sequences, supported a monophyly of three species, I. asiatica, Isoëtes echinospora, and Isoëtes maritima from Hokkaido, Kamchatka, and Alaska regions. The distinct species status of I. asiatica was also well supported in the combined chloroplast DNA phylogeny. Therefore, I. asiatica appear to represent example of gradual speciation due to spatial isolation of ancestral populations followed by genetic divergence. Our results also suggest that I. asiatica is probably not the ancestral diploid of the polyploids occurring in East Asia.     

Molecular phylogenetics of Fouquieriaceae: evidence from nuclear rDNA ITS studies

Molecular sequence data from the 18S-26S rDNA internal transcribed spacer (ITS) region support the monophyly of Fouquieria sensu lato (Fouquieriaceae) and the three subgenera (subg. Fouquieria, subg. Bronnia, subg. Idria) previously recognized within it. Resolution within subg. Fouquieria differs somewhat between parsimony and maximum likelihood (ML) trees. Section Fouquieria and sect. Ocotilla within subg. Fouquieria are not well supported as monophyletic groups. Uncertainty regarding placement of the root within Fouquieriaceae makes discussion of character evolution within the family difficult. Three root positions are consistent with rate-constant evolution of ITS sequences: (1) along the branch to subg. Idria, (2) along the branch to subg. Bronnia, and (3) along the branch to subg. Fouquieria. The first root position listed is equivalent to an outgroup rooting. The third root position listed is equivalent to a midpoint rooting. Of the three root positions above, only the third is along a branch that may be sufficiently long to act as a long-branch attractor. The first two root positions would result in character reconstruction suggesting that succulent growth forms and white floral pigmentation are ancestral within the family, with shifts to woody growth forms and to red floral pigmentation. The third root position results in equivocal reconstruction of the ancestral growth form, equivocal reconstruction of ancestral floral pigmentation in parsimony trees, and a suggestion of white floral pigmentation as ancestral in ML trees. Two previous hypotheses of polyploid origins are compatible with the molecular data presented here: (1) origin of the tetraploid F. diguetii from F. macdougalii, and (2) allopolyploid origin of the hexaploid F. burragei from the tetraploid F. diguetii and a diploid species similar to F. splendens. Direct descent of the hexaploid F. columnaris from the subg. Bronnia lineage is not supported by our data. An amphiploid origin of F. columnaris involving a member of the subg. Bronnia lineage and an extinct taxon outside subg. Bronnia, however, cannot be ruled out.     

Evolution and polyploid origins in North American Arctic Puccinellia (Poaceae) based on nuclear ribosomal spacer and chloroplast DNA sequences

The proportion of polyploid plant species increases at higher latitudes, and it has been suggested that original postglacial Arctic immigrants of some large groups, including grasses, were polyploid. We analyzed noncoding nuclear and chloroplast DNA of all North American diploid Puccinellia (Poaceae) and a subset of arctic polyploids to hypothesize evolutionary relationships among diploids and to evaluate the parentage of polyploids. Diploids formed three lineages: one uniting arctic species P. arctica and P. banksiensis; a second comprising arctic species P. tenella, P. alaskana, P. vahliana, and P. wrightii; and a third uniting the two temperate species P. lemmonii and P. parishii. The arctic species P. angustata (hexaploid) and P. andersonii (primarily octoploid) apparently derive from the P. arctica-P. banksiensis lineage based on ITS and chloroplast sequences, and share an ancestor with arctic triploid/tetraploid P. phryganodes based on nrDNA sequences. Sequence comparisons also suggest tetraploid P. bruggemannii evolved from two arctic lineages: P. vahliana-P. wrightii and P. arctica-P. banksiensis. These patterns and the predominance of arctic rather than temperate diploid species support the idea that diploid Puccinellia recolonized the Arctic from northern glacial refugia like Beringia, and also formed stabilized polyploid hybrids during these refugial events or subsequently during postglacial colonization.     

Phylogeography of two European Reticulitermes (Isoptera) species: the Iberian refugium

Chemical, i.e. cuticular hydrocarbons, and molecular data were used to probe the phylogeography of Reticulitermes termites collected from various parts of France, Spain and Portugal. Phylogenetic relationships were inferred from sequences of the internal transcribed spacer (ITS2) of nuclear ribosomal RNA genes as well as from two partial mitochondrial DNA segments, the cytochrome oxidase II gene and a sequence combining the tRNA-Leu gene and fragments of the NADH dehydrogenase I and ribosomal 16S genes. Two species, namely, R. grassei and R. banyulensis, were identified based on an analysis of cuticular hydrocarbons and the identification was confirmed by ITS2 haplotyping. However, phylogeny based on the analysis of mitochondrial DNA was not completely in agreement with the conclusions drawn from the chemical and nuclear data. An analysis of 56 R. grassei colonies revealed intraspecific differentiation into two major lineages with distinct geographical ranges. Whereas analysis of cuticular hydrocarbons showed that R. banyulensis was chemically distinct from R. grassei, analysis of mitochondrial DNA showed its close kinship with the R. grassei lineage occurring in southern Spain. This kinship could be explained by their evolution from a common polymorphic ancestor species in this ice age refugium.     

The evolution of genome size and rDNA in diploid species of Chenopodium s.l. (Amaranthaceae)

The evolution of genome size and ribosomal DNA (rDNA) locus organization was analysed in 23 diploid species of Chenopodium s.l., all of which share the same base chromosome number of x = 9. Phylogenetic relationships among these species were inferred from plastid and nuclear ribosomal internal transcribed spacer (nrITS) DNA sequences. The molecular phylogenetic analyses assigned all analysed species of Chenopodium s.l. to six evolutionary lineages, corresponding to the recent new generic taxonomic treatment of Chenopodium s.l. The distribution of rDNA loci for four species is presented here for the first time using fluorescence in situ hybridization (FISH) with 5S and 35S rDNA probes. Most of the 23 analysed diploid Chenopodium spp. possessed a single subterminally located 35S rDNA locus, except for three species which possessed two 35S rDNA loci. One or two 5S rDNA loci were typically localized subterminally on chromosomes, rarely interstitially. Analyses of rDNA locus numbers in a phylogenetic context resulted in the reconstruction of one locus each of 35S rDNA and 5S rDNA, both in subterminal positions, as the ancestral state. Genome sizes determined using flow cytometry were relatively small (2C value < 2.8 pg), ranging from 0.734 pg in C. schraderianum to 2.721 pg in C. californicum (nearly four‐fold difference), and were often conserved within major phylogenetic lineages, suggesting an adaptive value. The reconstructed ancestral genome size was small for all evolutionary lineages, and changes have probably coincided with the divergence of major lineages. © 2015 The Linnean Society of London, Botanical Journal of the Linnean Society, 2015, 179 , 218–235.     

Nuclear ribosomal DNA sequence variation and evolution of spotted marsh-orchids (Dactylorhiza maculata group)

Sequences of both internal and external transcribed spacers of nuclear ribosomal DNA were sequenced for four species belonging to the Dactylorhiza maculata group or "spotted marsh-Orchids". These four species are D. fuchsii, D. saccifera, D. foliosa, and D. maculata. Extensive nuclear ribosomal DNA polymorphism was uncovered within the diploid D. fuchsii and the putative autotetraploid D. maculata. Within the phylogenetic trees reconstructed using parsimony and Bayesian analyses, four main lineages (A, B, C, and D) were well supported. While D. saccifera, D. maculata, and D. foliosa were confined to clades B, C, and D, respectively, D. fuchsii accessions were spread over three clades (A, B, and C). Lineage C, which included accessions of the diploid D. fuchsii and the tetraploid D. maculata, was closely related to the lineage of D. foliosa (lineage D), an endemic diploid species from Madeira. Moreover, intra-individual polymorphism was found within accessions of D. maculata, D. fuchsii, and D. saccifera. It is shown that in some instances two lineages, contributed to the observed intra-individual polymorphism (C and A in D. maculata, A and B in D. fuchsii and D. saccifera). Evolutionary scenarios leading to this extensive nuclear ribosomal DNA polymorphism are discussed in the light of results from maternally inherited chloroplast DNA markers and an autopolyploid origin of D. maculata from a D. foliosa-like ancestor is postulated.     

Sequence Divergence of Microsatellites and Phylogeny Analysis in Tetraploid Cotton Species and Their Putative Diploid Ancestors

To determine the level of microsatellite sequence differences and to use the information to construct a phylogenetic relationship for cultivated tetraploid cotton （Gossypium spp.） species and their putative diploid ancestors, 10 genome-derived microsatellite primer pairs were used to amplify eight species, including two tetraploid and six diploid species, in Gossypium. A total of 92 unique amplicons were resolved using polyacrylamide gel electrophoresis. Each amplicon was cloned, sequenced, and analyzed using standard phylogenetic software. Allelic diversities were caused mostly by changes in the number of simple sequence repeat （SSR） motif repeats and only a small proportion resulted from interruption of the SSR motif within the locus for the same genome. The frequency of base substitutions was 0.5%-1.0% in different genomes, with only few indels found. Based on the combined 10 SSR flanking sequence data, the homology of A-genome diploid species averaged 98.9%, even though most of the amplicons were of the same size, and the sequence homology between G. gossypioides （Ulbr.） Standl. and three other D-genome species （G. raimondii Ulbr., G. davidsonii Kell., and G. thurberi Tod.） was 98.5%, 98.6%, and 98.5%, respectively. Phylogenetic trees of the two allotetraploid species and their putative diploid progenitors showed that homoelogous sequences from the A- and D-subgenome were still present in the polyploid subgenomes and they evolved independently. Meanwhile, homoelogous sequence interaction that duplicated loci in the polyploid subgenomes became phylogenetic sisters was also found in the evolutionary history of tetraploid cotton species. The results of the present study suggest that evaluation of SSR variation at the sequence level can be effective in exploring the evolutionary relationships among Gossypuim species.     

Inferring the history of the polyploid Silene aegaea (Caryophyllaceae) using plastid and homoeologous nuclear DNA sequences

The origin of the rare allotetraploid Silene aegaea was inferred from plastid rps16 intron sequences, homoeologous copies of nuclear ribosomal internal transcribed spacer (ITS) sequences, and an intron from the nuclear gene coding for the second largest subunit of RNA polymerase II (RPB2). The nuclear DNA regions support the S. sedoides and S. pentelica lineages as most closely related to the two S. aegaea paralogues. A few recombinant ITS sequences were found, but as PCR recombination could be demonstrated, no true recombination could be demonstrated. No recombination was found in the RPB2 sequences. Plastid rps16 intron sequences strongly support S. pentelica as the maternal lineage. The strength of the approach of using homoeologous sequences of several loci is demonstrated, and its usefulness for the study of phylogenies of groups including polyploids is emphasized.     

The influence of tectonics,sea‐level changes and dispersal on migration and diversification of Isonandreae (Sapotaceae)

Internal transcribed spacer (ITS) ribosomal DNA sequence data were generated for 80 of the c. 200 species of Isonandreae and were added to data from African and Neotropical representatives in subfamily Sapotoideae and outgroups in Sapotaceae. Bayesian dating and ancestral area reconstruction indicated that Isonandreae are derived from within an African grade. Multiple Australasian species or lineages are derived from Sundanian lineages in South‐East Asia with stem ages originating from the late Oligocene. Sri Lankan and Indian lineages are also derived from Sundanian lineages. Our results are consistent with migration from Africa into Sundania followed by numerous over‐water dispersal events across Wallace's Line into Australasia and migration from Sundania to the Indian subcontinent. Pleistocene speciation indicates that sea‐level changes during that epoch could have been responsible for some species diversification in Sundania. © 2013 The Linnean Society of London, Botanical Journal of the Linnean Society, 2014, 174 , 130–140.