Non-coding Plastid Regions Research Articles

Dates and rates in grape's plastomes: evolution in slow motion.

The family Vitaceae includes the domesticated grapevine (Vitis vinifera), one of the most economically important crops in the world. Despite the importance of Vitaceae, there is still considerable controversy surrounding their phylogenetic relationships and evolutionary timescales. Moreover, variation in rates of molecular evolution among Vitaceae remains mostly unexplored. The present research aims to fill these knowledge gaps through the analysis of plastome sequences. Thirteen newly sequenced grape plastomes are presented and their phylogenetic relationships examined. Divergence times and absolute substitution rates are inferred under different molecular clocks by the analysis of 95 non-coding plastid regions and 43 representative accessions of the major lineages of Vitaceae. Furthermore, the phylogenetic informativeness of non-coding plastid regions is investigated. We find strong evidence in favor of the random local clock model and rate heterogeneity within Vitaceae. Substitution rates decelerate in Ampelocissus, Ampelopsis, Nekemias, Parthenocissus, Rhoicissus, and Vitis, with genus Vitis showing the lowest values up to a minimum of ~ 4.65 × 10-11s/s/y. We suggest that liana-like species of Vitaceae evolve slower than erect growth habit plants and we invoke the "rate of mitosis hypothesis" to explain the observed pattern of the substitution rates. We identify a reduced set of 20 non-coding regions able to accurately reconstruct the phylogeny of Vitaceae and we provide a detailed description of all 152 non-coding regions identified in the plastomes of subg. Vitis. These polymorphic regions will find their applications in phylogenetics, phylogeography, and population genetics as well in grapes identification through DNA barcoding techniques.

Out of Amazonia and Back Again: Historical Biogeography of the Species-Rich Neotropical Genus Philodendron (Araceae)

The origin of Neotropical species diversity is strongly associated with the geological history of South America. Since the Miocene, a number of species radiations across different Neotropical lineages coincided with the rise of the Andes and the formation of the Isthmus of Panama. The species-rich genus Philodendron Schott (Araceae) is widely distributed across Neotropical rainforests, originating in the Late Oligocene and diversifying more intensely from the Miocene onward. It is likely that its diversification process and distribution patterns are associated with recent geological changes in the Americas. To test this hypothesis, we sampled the species diversity of Philodendron across its entire geographic range and used a combination of three non-coding plastid regions (petD, rpl16, and trnK/matK) to obtain a comprehensive time-calibrated phylogeny. We then inferred geographic range evolution and explored the impact of the Andean orogeny on speciation, extinction, and dispersal. The genus Philodendron originated ~29 million years ago (mya) and experienced the earliest diversification events ~25 mya in the Pan-Amazonian rainforests. From the Middle Miocene onward, multiple geographic range expansion events occurred from Amazonia to southeast Brazil and to the area which would become the Chocó and the northern Andes. From the Pliocene onward, Philodendron reached Central America and the Caribbean islands, and Andean lineages recolonized and diversified in Amazonia. In Philodendron, higher diversification rates are found in the adjacent lowland rainforests of the northern Andes compared with other regions in the Neotropics, demonstrating a potential indirect impact of the Andean uplift on species radiations in lowland regions.

Inferring the potential of plastid DNA-based identification of derived ferns: a case study on the Asplenium trichomanes aggregate in Europe

The utility of three plastid DNA regions to identify fern species was explored with focus on the European representatives of the Asplenium trichomanes aggregate. The sampling included representatives of the three diploid and the four tetraploid taxa recognized in the European flora plus Macaronesia. Besides European samples, the compiled data set comprised specimens of a putative Hawaiian endemic and one species occurring in Southeast Asia. By combining the sequences of three non-coding plastid regions, 13 haplotypes were recovered of which four were found in more than one taxon. Evidences for four distinct diploid lineages were found that correspond to Asplenium anceps, A. inexpectans, A. trichomanes s.s., and A. tripteropus. The four tetraploids occurring in Europe shared haplotypes with A. inexpectans. Thus, DNA barcoding can successfully identify the diploids, but fail to separate the tetraploids from their diploid ancestors. As a consequence, barcoding analyses of ferns need to take into account the differences of ploidy level measured by evidence independent from the DNA barcode. Evidence for uneven accumulation of intra-species DNA variation was recovered by comparing all species. Furthermore, the study provided evidence that the current taxonomy of these ferns requires to be revised. The two European diploids form well-separated clades and need to be recognized as A. inexpectans and A. trichomanes s.s. To keep name consistency for all European tetraploids, a new name Asplenium jessenii is introduced to replace A. trichomanes subsp. hastatum.

Cryptic genetic diversity in Solanum elaeagnifolium (Solanaceae) from South America

Solanum elaeagnifolium is a global invasive weed that is polyploid in the South American range. Our aim was to establish the origin and distribution of these polyploid lineages, together with their relationship with the invasion of new habitats. Ploidy level was determined in 26 natural populations using chromosomal counts, and two non-coding plastid regions trnL-trnF and rpl32F-trnL were sequenced. Relationships among haplotypes were examined by a median-joining network, and groups of populations were identified by employing a spatial analysis of molecular variance. Phylogenetic relationships among haplotypes were reconstructed using Bayesian inference, and divergence times were estimated using BEAST. We found 21 haplotypes structured into three lineages: one with diploids in North America and invaded areas, and two in South America that developed independent polyploidisation events. The separation of lineages took place during the Pliocene with more recent divergences in the Early Pleistocene. The existence of three lineages can be attributed to the uplift of the Andes and the emergence of the Isthmus of Panama. Diversification within each lineage may be related to the recent cyclical glacial variation throughout the Pleistocene creating haplotype diversity and recurrent polyploidy. Thus, the South American populations of S. elaeagnifolium did not originate from a recent colonisation but are established ancient lineages.

Melting pot of biodiversity: first insights into the evolutionary patterns of the Colchic bramble flora (RubussubgenusRubus, Rosaceae)

The Caucasus is a biodiversity hotspot of global significance, containing a number of highly diverse and species-rich plant taxa. The region is also thought to be an important evolutionary hotspot for Rubus subgenus Rubus (brambles). However, Caucasian brambles have only been poorly studied to date and our knowledge of their evolutionary mechanisms, systematics and taxonomic variability remains rudimentary. Therefore, the objectives of this study were to shed light on the evolution, diversity and reproduction modes of Rubus in one of the two Caucasian glacial refugia, Colchis. Flow cytometry measurements were used to estimate DNA ploidy, a flow cytometric seed screen was conducted to determine reproduction mode and Sanger sequencing of two non-coding plastid regions was used to reveal phylogenetic patterns. The most common ploidy level was tetraploid, followed by diploid and (rarely) triploid. Intra-individual variation in reproduction mode was low, as the morphoseries Glandulosi and Radula exhibited strict sexuality and other taxa were mostly apomictic. A few exceptions were observed that deserve further attention, e.g. sexuality induced hypothetically by haploid pollen or by environmental conditions, a high proportion of triploid embryos or polyspermy. Plastid haplotype variability revealed specific, ancient evolutionary patterns with limited involvement of extant diploid taxa and recent isolation from European brambles. We provide the first insight into the variability and evolution of Colchic brambles, which serves as a starting point for further systematic and evolutionary studies.

Intra and interspecific sequence variation in closely related species of Cereus (CACTACEAE)

In comparative phylogenetic and population genetic studies, one of the most crucial steps is to select appropriate DNA markers, a decision based primarily on the estimated variation in markers in cross-taxonomic surveys. To assess whether genetic variation at the intraspecific level in one species predicts the variation in another closely related species we used two congeneric species of Cereus (Cactaceae: Cereeae). We screened and characterized eight noncoding plastid regions (trnS-trnG, atpI-atpH, trnT-trnL, psbD-trnT, petL-psbE, 3'rps16-5'trnk, trnG intron, and trnL intron), and one nuclear gene (PhyC) in Cereus fernambucensis and C. hildmannianus. A total of 40 individuals from 15 populations were characterized according to nucleotide diversity, number of haplotypes, and number of potentially informative characters. The results revealed that nucleotide substitutions and indels are the main source of variation, with the largest divergence between species found in trnS-trnG. The trnL intron and petL-psbE showed intraspecific variability in both species. The psbD-trnT, atpI-atpH, trnS-trnG, and trnT-trnL, which are the most variable regions in one species, showed no variation in the other. Finally, the nuclear gene PhyC showed more resolution between Cereus species than within species. We thus found considerable heterogeneity among widely used plastid markers, even between closely related species, and suggest the use of PhyC as a marker for phylogenetic inference in these species. These results reinforce the need of screening as a preliminary step to conduct phylogeographic or phylogenetic studies in face of unpredictable sequence variation of molecular markers in plants.

Revisiting <I>Luffa</I> (Cucurbitaceae) 25 Years After C. Heiser: Species Boundaries and Application of Names Tested with Plastid and Nuclear DNA Sequences

Abstract The circumscription of the five to eight species of Luffa, as well as their correct names, have long been problematic. Experts on the genus, most recently C. Heiser and C. Jeffrey, have disagreed on the number of species in the New World and the application of the name L. operculata, which in turn affected the names L. quinquefida and L. sepium. Heiser used classic biosystematic methods, including experimental crossing, to infer species boundaries, but neither researcher had today's option of using DNA sequences for this purpose. We sequenced 51 accessions of Luffa, representing the geographic range of the genus and as much as possible topotypical or type material. Phylogenies from four non-coding plastid regions and the nuclear ribosomal DNA spacer region show that eight clades of specimens have geographic-morphological coherence. Heiser's view that Luffa has three species in the New World is supported, and there are four species in tropical and subtropical Asia. Australia has an endemic species...

Predicting Plastid Marker Variation: Can Complete Plastid Genomes from Closely Related Species Help?

Rapidly evolving non-coding plastid regions (NCPs) are currently widely used in evolutionary biology especially in plant systematic studies where NCPs have become one of the most commonly used tools in clarifying species relationships. Currently, the generally small amount of sequence variation provided by NCPs compared to nuclear regions makes plastid phylogeny reconstruction challenging at the species-level, especially so in species rich clades such as Solanum with c. 1,200 species. Previous studies have established that the set of most highly variable NCPs vary between major plant families, and here we explore whether this variation extends beyond family level to genera and major clades within genera. Using full plastome data, we identify the most highly variable plastid markers in the Potato clade of Solanum. We then compare sequence variation between the Potato and the closely related Morelloid clade. Results show that whilst a narrow set of NCPs show consistently high variation, levels of sequence variation in most NCPs differ greatly between the two closely related clades. The high variation detected between closely related groups implies that repeated screening studies will be needed for individual projects despite the potential availability of results from closely related taxa, and indicates a narrower applicability of family-specific screening studies than previously thought.

Complex evolution in Arundinarieae (Poaceae: Bambusoideae): Incongruence between plastid and nuclear GBSSI gene phylogenies

The monophyly of tribe Arundinarieae (the temperate woody bamboos) has been unequivocally recovered in previous molecular phylogenetic studies. In a recent phylogenetic study, 10 major lineages in Arundinarieae were resolved based on eight non-coding plastid regions, which conflicted significantly with morphological classifications both at the subtribal and generic levels. Nevertheless, relationships among and within the 10 lineages remain unclear. In order to further unravel the evolutionary history of Arundinarieae, we used the nuclear GBSSI gene sequences along with those of eight plastid regions for phylogenetic reconstruction, with an emphasis on Chinese species. The results of the plastid analyses agreed with previous studies, whereas 13 primary clades revealed in the GBSSI phylogeny were better resolved at the generic level than the plastid phylogeny. Our analyses also revealed many inconsistencies between the plastid DNA and the nuclear GBSSI trees. These results implied that the nuclear genome and the plastid genome had different evolutionary trajectories. The patterns of incongruence suggested that lack of informative characters, incomplete lineage sorting, and/or hybridization (introgression) could be the causes. Seven putative hybrid species were hypothesized, four of which are discussed in detail on the basis of topological incongruence, chromosome numbers, morphology, and distribution patterns, and those taxa probably resulted from homoploid hybrid speciation. Overall, our study indicates that the tribe Arundinarieae has undergone a complex evolution.

Forensic botany: species identification of botanical trace evidence using a multigene barcoding approach

Forensic botany can provide significant supporting evidence during criminal investigations. However, it is still an underutilized field of investigation with its most common application limited to identifying specific as well as suspected illegal plants. The ubiquitous presence of plant species can be useful in forensics, but the absence of an accurate identification system remains the major obstacle to the present inability to routinely and correctly identify trace botanical evidence. Many plant materials cannot be identified and differentiated to the species level by traditional morphological characteristics when botanical specimens are degraded and lack physical features. By taking advantage of a universal barcode system, DNA sequencing, and other biomolecular techniques used routinely in forensic investigations, two chloroplast DNA regions were evaluated for their use as "barcoding" markers for plant identification in the field of forensics. We therefore investigated the forensic use of two non-coding plastid regions, psbA-trnH and trnL-trnF, to create a multimarker system for species identification that could be useful throughout the plant kingdom. The sequences from 63 plants belonging to our local flora were submitted and registered on the GenBank database. Sequence comparison to set up the level of identification (species, genus, or family) through Blast algorithms allowed us to assess the suitability of this method. The results confirmed the effectiveness of our botanic universal multimarker assay in forensic investigations.

Phylogenetic Relationships of Zeugites (Poaceae: Centothecoideae) Inferred from Plastid and Nuclear DNA Sequences and Morphology

The genus Zeugites P. Browne encompasses ten species of neotropical grasses. To infer the phylogenetic relationships of the species, parsimony analyses of morphological data and DNA sequences from a nuclear region (ITS) and two non-coding plastid regions (the trnL intron and the trnL-trnF intergenic spacer) were undertaken for each data set separately and in combination. The combined analysis is presented as our preferred phylogenetic hypothesis because of its better resolution and higher support for the clades. Zeugites is monophyletic only if Calderonella sylvatica is included in it. Our analyses recover two major clades within Zeugites: Clade I, formed by Zeugites pittieri, Zeugites latifolia, Zeugites sylvatica (Calderonella sylvatica), Zeugites sagittata, and Zeugites capillaris; and Clade II, including Zeugites hackelii, Zeugites smilacifolia, Zeugites munroana, Zeugites pringlei, Zeugites mexicana, and Zeugites americana. The monospecific genus Pohlidium is strongly supported as sister of Zeugites (including Calderonella) but it is maintained as a different genus due to noticeable differences in spikelet and inflorescence morphology.

Phylogenetic Relationships of Zeugites (Poaceae: Centothecoideae) Inferred from Plastid and Nuclear DNA Sequences and Morphology

The genus Zeugites P. Browne encompasses ten species of neotropical grasses. To infer the phylogenetic relationships of the species, parsimony analyses of morphological data and DNA sequences from a nuclear region (ITS) and two non-coding plastid regions (the trnL intron and the trnL-trnF intergenic spacer) were undertaken for each data set separately and in combination. The combined analysis is presented as our preferred phylogenetic hypothesis because of its better resolution and higher support for the clades. Zeugites is monophyletic only if Calderonella sylvatica is included in it. Our analyses recover two major clades within Zeugites: Clade I, formed by Zeugites pittieri, Zeugites latifolia, Zeugites sylvatica (Calderonella sylvatica), Zeugites sagittata, and Zeugites capillaris; and Clade II, including Zeugites hackelii, Zeugites smilacifolia, Zeugites munroana, Zeugites pringlei, Zeugites mexicana, and Zeugites americana. The monospecific genus Pohlidium is strongly supported as sister of Zeugites (including Calderonella) but it is maintained as a different genus due to noticeable differences in spikelet and inflorescence morphology.

Cryptic Speciation in the Caesalpinia hintonii Complex (Leguminosae: Caesalpinioideae) in a Seasonally Dry Mexican Forest

[corrected] The Caesalpinia hintonii group comprises six species of endemic shrubs or trees, C. epifanioi, C. hintonii, C. laxa, C. macvaughii, C. melanadenia and C. oyamae, found in scattered patches of seasonally dry forest in the Río Balsas depression and the neighbouring Tehuacán-Cuicatlán valley, which are part of the Mexican morphotectonic province of Sierra Madre del Sur. An evaluation is made of phylogeographic patterns and genetic diversity with a phylogenetic analysis of the C. hintonii complex in order to study the dynamics of speciation in this endemic group of legumes. A phylogeographic study based on four highly variable non-coding plastid regions (trnL intron, trnL-F intergenic spacer, trnH-psbA intergenic spacer, and accD-psaI intergenic spacer) was carried out for the Caesalpinia hintonii complex. Five of the six taxa of the C. hintonii complex were included. The plastid analyses involving multiple accessions of each taxon from throughout their ranges resolved C. epifanioi and C. hintonii as well-supported clusters, but C. oyamae has two unexpectedly divergent lineages. Two well-supported geographic clades: eastern (C. epifanioi, C. melanadenia and C. oyamae) and western (C. hintonii and C. macvaughii) were established. The analyses performed provide evidence of recent morphostatic radiation in C. oyamae resulting from isolation and local adaptation. This pattern of genetic differentiation without morphological divergence may be a model that fits many groups of tropical woody taxa inhabiting similarly dry forests subjected to shifting selection.

Phylogenetic relationships of the recently discovered species — Clivia mirabilis

A phylogenetic study of Clivia, including the newly described species, Clivia mirabilis, recently discovered in the Northern Cape, was carried out to elucidate the relationship of this geographically isolated taxon to the remaining four species. DNA sequence data were collected from four non-coding plastid regions: the rpoBtrnC intergenic spacer, the rps16 intron, the trnL intron and the trnL-F intergenic spacer (the latter two collectively known as the trnL-F region). In the combined tree C. mirabilis is placed as sister to a clade comprising the other four taxa.