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A taxonomic survey of the relict and poorly known circum-Mediterranean Anagyris (Fabaceae) was conducted using a total evidence principal approach based on morphological and molecular data. Statistical analysis of both qualitative and quantitative morphological and chemically-derived traits allowed us to separate the Mediterranean A. foetida and the Canarian A. latifolia as independent species. Newly analyzed traits relating to the odour of leaves, number of inflorescences per branch, colour of fresh calyx, and degree of corolla opening, were diagnostic for identifying these two previously almost indistinguishable taxa. Significant differences in quantitative morphometric traits indicate that vegetative and floral characters are overall larger in A. latifolia than in A. foetida, supporting the hypothesis of increased size and woodiness of Macaroneisan endemic plants compared to the reduced sizes of thier continental counterparts adapted to the more xeric Mediterranean climate. Combined plastid trnLF and nuclear ribosomal ITS data recovered a monophyletic origin of Anagyris within the central-western Eurasian Thermopsideae clade and its split into two lineages. The dated divergences of the Anagyris lineages, calculated by Bayesian relaxed-clock methods with the combined sequence data, showed that Anagyris diverged in the late Miocene (8.2 ± 4.5 Ma), and that the origin of A. foetida (3.6 ± 3.2 Ma) pre-dated that of A. latifolia (1.9 ± 2.1 Ma).

Background and AimsTo date chloroplast genomes are available only for members of the non-protein amino acid-accumulating clade (NPAAA) Papilionoid lineages in the legume family (i.e. Millettioids, Robinoids and the ‘inverted repeat-lacking clade’, IRLC). It is thus very important to sequence plastomes from other lineages in order to better understand the unusual evolution observed in this model flowering plant family. To this end, the plastome of a lupine species, Lupinus luteus, was sequenced to represent the Genistoid lineage, a noteworthy but poorly studied legume group.MethodsThe plastome of L. luteus was reconstructed using Roche-454 and Illumina next-generation sequencing. Its structure, repetitive sequences, gene content and sequence divergence were compared with those of other Fabaceae plastomes. PCR screening and sequencing were performed in other allied legumes in order to determine the origin of a large inversion identified in L. luteus.Key ResultsThe first sequenced Genistoid plastome (L. luteus: 155 894 bp) resulted in the discovery of a 36-kb inversion, embedded within the already known 50-kb inversion in the large single-copy (LSC) region of the Papilionoideae. This inversion occurs at the base or soon after the Genistoid emergence, and most probably resulted from a flip–flop recombination between identical 29-bp inverted repeats within two trnS genes. Comparative analyses of the chloroplast gene content of L. luteus vs. Fabaceae and extra-Fabales plastomes revealed the loss of the plastid rpl22 gene, and its functional relocation to the nucleus was verified using lupine transcriptomic data. An investigation into the evolutionary rate of coding and non-coding sequences among legume plastomes resulted in the identification of remarkably variable regions.ConclusionsThis study resulted in the discovery of a novel, major 36-kb inversion, specific to the Genistoids. Chloroplast mutational hotspots were also identified, which contain novel and potentially informative regions for molecular evolutionary studies at various taxonomic levels in the legumes. Taken together, the results provide new insights into the evolutionary landscape of the legume plastome.

Cistus pouzolzii is a species distributed in the western Mediterranean. Its populations are located in France and North Africa, being absent in the Iberian Peninsula.. It was described by Delile, but its taxonomic and nomenclatural status has been discussed by several authors, mainly based on its particular combination of morphological characters. We have recently found several populations of a Cistus species whose morphological characters match those of C. pouzolzii. To confirm the taxonomic identity of these populations, a comparative study was performed using herbarium specimens of C. pouzolzii from France and North Africa. In addition, two chloroplast regions were sequenced to study the phylogenetic relationships between the newly discovered Iberian populations and the rest of the species of the genus Cistus, including the sequences belonging to C. pouzolzii from France and Morocco previously used. Both analyses (morphological and phylogenetic studies) confirmed that the specimens sampled in the Iberian populations were C. pouzolzii. Furthermore, both the nomenclature of this species and its conservation status are briefly discussed.

One of the longstanding questions in phylogenetic systematics is how to address incongruence among phylogenies obtained from multiple markers and how to determine the causes. This study presents a detailed analysis of incongruent patterns between plastid and ITS/ETS phylogenies of Tribe Senecioneae (Asteraceae). This approach revealed widespread and strongly supported incongruence, which complicates conclusions about evolutionary relationships at all taxonomic levels. The patterns of incongruence that were resolved suggest that incomplete lineage sorting (ILS) and/or ancient hybridization are the most likely explanations. These phenomena are, however, extremely difficult to distinguish because they may result in similar phylogenetic patterns. We present a novel approach to evaluate whether ILS can be excluded as an explanation for incongruent patterns. This coalescence-based method uses molecular dating estimates of the duration of the putative ILS events to determine if invoking ILS as an explanation for incongruence would require unrealistically high effective population sizes. For four of the incongruent patterns identified within the Senecioneae, this approach indicates that ILS cannot be invoked to explain the observed incongruence. Alternatively, these patterns are more realistically explained by ancient hybridization events.

Phylogenetic studies have demonstrated the largest morning glory genus, Ipomoea, is not monophyletic, and nine other segregate genera are derived from within Ipomoea. Therefore, systematic research is focused on the monophyletic tribe Ipomoeeae (c. 650-900 species). We used whole plastid genomes to infer relationships among exemplar species distributed across Ipomoeeae. Whole plastomes were sequenced, assembled and annotated for twenty-eight morning glory species, representing major Ipomoeeae lineages. Phylogenies were estimated for twenty-eight species sequenced here and one published plastome using alignments of: (a) eighty-two chloroplast genes and (b) whole plastomes. In addition, divergence times were estimated to date key divergence events within the Ipomoeeae. Two nodes were calibrated with fossil pollen for molecular dating analyses. Phylogenies estimated from the two plastome datasets had identical topologies. Phylogenetic results are generally consistent with prior phylogenetic analyses of morning glories. Two major clades, previously named Astripomoeinae and Argyreiinae, are well-supported. There also is support for the monophyly of Ipomoea subgenus Quamoclit. Higher-level relationships with weak support in previous analyses were recovered in this analysis with strong support. Results from the molecular dating analysis suggest a middle Eocene divergence time for the Ipomoeeae. Furthermore, the Argyreiinae clade was found to have diversified before the Astripomoeinae; however, error bars overlap between divergence time estimates of these two clades. Phylogenetic results presented here provide greater confidence in relationships among major lineages of the Ipomoeeae, and divergence time estimation results provide a temporal context for the diversification of this fascinating group of angiosperms.

Background Next-generation sequencing has provided a wealth of plastid genome sequence data from an increasingly diverse set of green plants ( Viridiplantae ). Although these data have helped resolve the phylogeny of numerous clades (e.g., green algae, angiosperms, and gymnosperms), their utility for inferring relationships across all green plants is uncertain. Viridiplantae originated 700-1500 million years ago and may comprise as many as 500,000 species. This clade represents a major source of photosynthetic carbon and contains an immense diversity of life forms, including some of the smallest and largest eukaryotes. Here we explore the limits and challenges of inferring a comprehensive green plant phylogeny from available complete or nearly complete plastid genome sequence data. Results We assembled protein-coding sequence data for 78 genes from 360 diverse green plant taxa with complete or nearly complete plastid genome sequences available from GenBank. Phylogenetic analyses of the plastid data recovered well-supported backbone relationships and strong support for relationships that were not observed in previous analyses of major subclades within Viridiplantae . However, there also is evidence of systematic error in some analyses. In several instances we obtained strongly supported but conflicting topologies from analyses of nucleotides versus amino acid characters, and the considerable variation in GC content among lineages and within single genomes affected the phylogenetic placement of several taxa. Conclusions Analyses of the plastid sequence data recovered a strongly supported framework of relationships for green plants. This framework includes: i) the placement of Zygnematophyceace as sister to land plants ( Embryophyta ), ii) a clade of extant gymnosperms ( Acrogymnospermae ) with cycads +  Ginkgo sister to remaining extant gymnosperms and with gnetophytes ( Gnetophyta ) sister to non- Pinaceae conifers (Gnecup trees), and iii) within the monilophyte clade ( Monilophyta ), Equisetales  +  Psilotales are sister to Marattiales  + leptosporangiate ferns. Our analyses also highlight the challenges of using plastid genome sequences in deep-level phylogenomic analyses, and we provide suggestions for future analyses that will likely incorporate plastid genome sequence data for thousands of species. We particularly emphasize the importance of exploring the effects of different partitioning and character coding strategies.

Petunia secreta is a rare and endemic species, that was found in two different landscapes, approximately 21 Km apart from each other. In this study we showed that P. secreta presented high genetic diversity that was equivalent to or even higher than that of widespread Petunia species. Two evolutionary lineages were found and they are correlated to the different landscapes where P. secreta grows: open areas in conglomerate sandstone towers at an elevation of approximately 300-400 m or along the road growing in an open vegetation flat area. Therefore the major risk to P. secreta maintenance is its rarity, suggesting the necessity of a preservation program. The analysis of genetic structure and variability of isolated species is of critical importance in evaluating whether stochastic or human-caused factors are affecting rare species. Low genetic diversity compromises the ability of populations to evolve and reduces their chances of survival under environmental changes. Petunia secreta, a rare and endemic species, is an annual and heliophilous herb that is bee-pollinated and easily recognizable by its purple and salverform corolla. It was described as a new species of the Petunia genus in 2005. Few individuals of P. secreta have been observed in nature and little is known about this species. All the natural populations of P. secreta that were found were studied using 15 microsatellite loci, two intergenic plastid sequences and morphological traits. Statistical analysis was performed to describe the genetic diversity of this rare species and the results compared with those of more widespread and frequent Petunia species from the same geographic area to understand whether factors associated with population size could affect rare species of this genus. The results showed that despite its rarity, P. secreta presented high genetic diversity that was equivalent to or even higher than that of widespread Petunia species. It was shown that this species is divided into two evolutionary lineages, and the genetic differentiation indices between them and other congeneric species presented different patterns. The major risk to P. secreta maintenance is its rarity, suggesting the necessity of a preservation programme and more biological and evolutionary studies that handle the two evolutionary lineages independently.

The contrasting evolutionary histories of endemic versus related cosmopolitan species provide avenues to understand the spatial drivers and limitations of biodiversity. Here, we investigated the evolutionary history of three New Zealand endemic Deschampsia species, and how they are related to cosmopolitan D. cespitosa. We used RADseq to test species delimitations, infer a dated species tree, and investigate gene flow patterns between the New Zealand endemics and the D. cespitosa populations of New Zealand, Australia and Korea. Whole plastid DNA analysis was performed on a larger worldwide sampling. Morphometrics of selected characters were applied to New Zealand sampling. Our RADseq review of over 55 Mbp showed the endemics as genetically well-defined from each other. Their last common ancestor with D. cespitosa lived during the last ten MY. The New Zealand D. cespitosa appears in a clade with Australian and Korean samples. Whole plastid DNA analysis revealed the endemics as members of a southern hemisphere clade, excluding the extant D. cespitosa of New Zealand. Both data provided strong evidence for hybridization between D. cespitosa and D. chapmanii. Our findings provide evidence for at least two migration events of the genus Deschampsia to New Zealand and hybridization between D. cespitosa and endemic taxa.

Premise of the study: The subfamily Panicoideae (Poaceae) encompasses nearly one-third of the diversity of grass species, including important crops such as maize and sugarcane. Previous analyses recovered strong support for a Panicoideae+Centothecoideae lineage within the diverse Panicoideae+Arundinoideae+Chloridoideae+Micrairoideae+Aristidoideae+Danthonioideae (PACMAD) clade, although support for internal relationships was inconsistent. The objectives of this research were to (1) further test the monophyly of each subfamily and previously recovered clades within the Panicoideae+Centothecoideae lineage, (2) establish phylogenetic relationships among these groups, and (3) propose a new tribal classification for this lineage based explicitly on the phylogeny. METHODS: Maximum parsimony and Bayesian inference analyses of 37 taxa were based on previously published sequences (ndhF and rpl16 intron) and on new plastid and nuclear (rbcL and granule-bound starch synthase I) sequence data as well as structural data. Key results. The Panicoideae+Centothecoideae lineage and a majority of the clades identified in previous analyses continue to be robustly supported, but resolution along the backbone of the topology remains elusive. Support for the monophyly of both subfamilies was lacking although support values for some clades increased. The tribes Centotheceae and Arundinelleae were confirmed as polyphyletic. CONCLUSIONS: Subfamily Centothecoideae is formally submerged into the Panicoideae, and a new tribal classification for the expanded Panicoideae is proposed based explicitly on the phylogeny. This classification includes 12 tribes of which Chasmanthieae and Zeugiteae are segretated from the Centotheceae; Tristachyideae is segregated from Arundinelleae, and a new tribe, Cyperochloeae, is validated to accommodate two isolated genera. A key to the tribes is provided.

Aim The post-glacial range dynamics of many European plant species have been widely investigated, but information rapidly diminishes as one moves further back in time. Here we infer the historical range shifts of Laurus, a paradigmatic tree of the Tethyan flora that has covered southern Eurasia since the Oligo-Miocene, by means of phylogenetic and phylogeographical analyses. Location Mediterranean Basin, Black Sea and Macaronesian archipelagos (Azores, Madeira, Canary Islands). Methods We analysed plastid DNA (cpDNA) sequence (trnK–matK, trnD–trnT) variation in 57 populations of Laurus and three Lauraceae genera. Phylogenetic methods (maximum parsimony and Bayesian inference) and statistical parsimony networks were used to reconstruct relationships among haplotypes. These results were contrasted with the fossil record and bioclimatic niche-based model predictions of past distributions to infer the migration routes and location of refugia. Results The phylogenetic tree revealed monophyly for Laurus. Overall sequence variability was low within Laurus, but six different haplotypes were distinguished and a single network retrieved, portraying three lineages primarily related to geography. A strongly divergent eastern lineage occupied Turkey and the Near East, a second clade was located in the Aegean region and, lastly, a western clade grouped all Macaronesian and central and western Mediterranean populations. A close relationship was observed between the Macaronesian populations of L. azorica and the western populations of L. nobilis. Main conclusions The phylogeographical structure of Laurus preserves the imprints of an ancient contraction and break-up of the range that resulted in the evolution of separate cpDNA lineages in its western- and easternmost extremes. Intense range dynamics in the western Mediterranean and multiple glacial refugia contributed to the generation and long-term conservation of this phylogeographical pattern, as shown by the fit between the haplotype ranges and past suitable areas inferred from bioclimatic models. Finally, our results challenge the taxonomic separation of Laurus into two distinct species.