trnL Intron Research Articles

The phylogenetic position of Aglaodorum Schott (Araceae – Aroideae – Aglaonemateae)

Analysis of chloroplast DNA sequences (trnL intron and trnLtrnF intergenic spacer) of the helophytic colony-forming Aglaodorum griffithii (Schott) Schott and other representative species of Araceae supported its placement within Aglaonema Schott. It is therefore proposed that Aglaodorum Schott should be recognised as a generic synonym of Aglaonema.

Genetic Diversity and Phylogenetic Analyses of Laurus nobilis L. (Lauraceae) Populations Revealed Chloroplast (cpDNA) trnL Intron and trnL-F Region

ABSTRACT In this study, genetic diversity and phylogenetic analyzis based on cpDNA trnL intron and trnL-F sequences of L. nobilis L. populations were made. Persea schiedeana and Endiandra wolfei taxa were used as outgroups. Plant samples were collected from various different provinces and districts of the Aegean region and brought to the laboratory. Genomic DNAs were isolated from fresh and green leaves of the plants. trnC and trnD primers were used for the trnL intron region, and trnE and trnF primers were used for the trnL-F region. The obtained DNA sequences were then corrected using BioEdit and FinchTV programs. MEGA 6.0 program was used both phylogenetic analyzis and the detection of genetic distances. For the intron region of the trnL, the mean nucleotide composition was determined as 36.5% T, 20.2% C, 26.4% A and 16.9% G. These values were found as 30.7% T for 15.9% C, 31.9% A and 21.6% G for the trnL-F region. The lowest distance for the trnL intron region was found 0.000 while the highest distance was 0.004. The value was found 0.000 for the trnL-F region. The phylogenetic tree, which was obtained using both trnL intron and trnL-F sequences, consisted of two large branches. As a result, according to the data obtained from trnL intron and trnL-F sequences, the trnL intron sequence found a solvent in the phylogenetic relationship between L. nobilis populations, while the phylogenetic relationship between trnL-F sequence and L. nobilis populations was not found.

Incongruence of morphological determinations and DNA barcode sequences: a case study in Cinnamomum (Lauraceae)

In the course of molecular systematic studies of Lauraceae we received a sample of a plant cultivated under the name Cinnamomum porrectum in the Botanical Garden Munchen-Nymphenburg. Preliminary determinations, both morphologically based on the Flora of China (Li & al. 2008) and by chloroplast sequences (psbA-trnH spacer, trnK intron including matK gene, trnL intron, trnL-trnF spacer and trnQ-rps16 spacer) obtained by Sanger sequencing suggested that it was C. camphora, still the plant looked different from other individuals of C. camphora cultivated in the botanical gardens of Berlin, Hamburg, Mainz, Munich and Oldenburg. Attempts to sequence the more informative nuclear internal transcribed spacer repeatedly led to mixed signals. We therefore used Illumina sequencing on a set of pre-amplified molecular markers (ITS, trnK 3′ and 5′ intron, trnL intron, and the intergenic spacers psbA-trnH, trnL-trnF, as well as parts of the trnQ-rps16 spacer), and downloaded available sequences of C. camphora and C. parthenoxylon from GenBank for comparison. Considerable differences were found among these sequences, but the haplotype groups do not coincide with the current species determinations. Particularly the internal transcribed spacer sequences are rather diverse, suggesting possible misidentifications, contaminations, and/or a common gene pool that is larger than anticipated. Concerning the plant in question, our results suggest that it may be a hybrid, with C. camphora as the maternal and another species, possibly C. parthenoxylon, as the paternal parent.Citation: Rohwer J. G., Trofimov D., Mayland-Quellhorst E. & Albach D. 2019: Incongruence of morphological determinations and DNA barcode sequences: a case study in Cinnamomum (Lauraceae). – Willdenowia 49: 383–400. doi: https://doi.org/10.3372/wi.49.49309Version of record first published online on 4 December 2019 ahead of inclusion in December 2019 issue.

Aeschynomene chicocesariana, a Striking New Unifoliolate Legume Species from the Brazilian Chapada Diamantina and its Phylogenetic Placement in the Dalbergioid Clade

Abstract—Unifoliolate leaves are relatively rare in the species-rich dalbergioid clade of papilionoid legumes (Leguminosae, Papilionoideae). For the pantropical, diverse genus Aeschynomene (ca. 180 species), the only known record of unifoliolate species described so far is the savanna-inhabiting A. simplicifolia from Chapada dos Veadeiros in Goiás, Brazil. The current work provides a description, illustrations, and a distribution map of another unifoliolate species, Aeschynomene chicocesariana, which was newly discovered in the rupestrian grasslands of Chapada Diamantina in Bahia, Brazil. Bayesian phylogenetic analyses of DNA sequence data (nuclear ribosomal ITS/5.8S and the plastid genes matK and trnL intron) have unequivocally placed A. chicocesariana with the rest of Aeschynomene species that are traditionally placed within Aeschynomene sect. Ochopodium. This result agrees with the shared morphology of A. chicocesariana and members of that section.

First Report of Sieversian Wormwood (Artemisia sieversiana) as a New Host of Dodder (Cuscuta australis) in China

Sieversian wormwood (Artemisia sieversiana Ehrhart ex Willd; Compositae), annual or biennial herb, is a widely used medicinal plant in both traditional Chinese medicine and Tibetan medicine (Jiangsu New Medical College 1977). Sieversian wormwood has a broad spectrum of pharmacological activities such as antiphlogosis, detumescence, and hemostatic. This wormwood is widely distributed in many provinces of China, including the three northeastern provinces of Liaoning, Jilin, and Heilongjiang, and has been planted and grows in Shandong and Jiangsu. On 22 July 2018, dodder was first found on Sieversian wormwood on the north embankment of the Xinkai River (N 41°48′36.51″, E 123°33′21.32″, 53 m elevation) in Shenyang (temperate continental monsoon climate), Liaoning. About 15 Sieversian wormwood plants in approximately 10 m² were parasitized by the dodder. The dodder was mostly located in the middle-third to upper canopy, where the Sieversian wormwood stalks and petioles were firmly entwined by the dodder. The branched dodder stems spread among neighboring Sieversian wormwood plants and were also found parasitizing surrounding Chenopodium album L., Kochia scoparia (L.) Schrad., Setaria viridis (L.) Beauv., and Artemisia annua L. The Sieversian wormwood plants parasitized by the dodder showed chlorotic and wilting leaves, droopy branches, and stunted growth. All wilt symptoms were most evident on warm, sunny days. However, the affected host had no clear lesions around the plant tissues where the dodder haustoria were clearly observed penetrating. The orange, leafless dodder was 0.6044 ± 0.093 mm in diameter (mean ± SD, n = 15), reproduced synchronically with its new host. The dodder inflorescences were compact cymose and subsessile. Corolla were creamy white and cupulate, abutted by oblong, not overlapping calyces, corolla lobes were obtuse, upright to spreading. Capsules were depressed-globose, not circumscissile, and only its lower half was typically enclosed by the withered corolla. There were four seeds per capsule. Seeds were brownish, ovoid, scabrous, and with a short line hilum. The dodder was tentatively identified as Cuscuta australis R. Brown, Prodr. according to its inflorescence, petal, calyx, pedicle, capsule, ovary, and seed characteristics (other specific features see Wu 1979; Yuncker 1932). The 436-bp internal transcribed spacer 2 region (ITS2) of our dodder rDNA (deposited in GenBank, accession no. MK106064) had the highest similarity with C. australis references (GenBank nos. KF454371 [99.03%], MG548813 [99.31%], and MH370812 [99.54%]). The 209-bp chloroplast trnL intron of our dodder (GenBank no. MK879383) had 100% similarity with C. australis reference (GenBank no. MK879385), as well as 99.62% with another reference (AY558828). The two gene sequences together confirmed its identity as C. australis, albeit the dodder ITS2 was also with high similarity (99%) with C. campestris. The dodder, C. australis, has been reported on many plants of the families Compositae, Leguminosae, and Polygonaceae, mainly on widely cultivated crops and local-specialty medicinal herbs in agricultural fields, such as chickpea (Cicer arietinum L.) and niuxi (Achyranthes bidentate Blume) (Chen et al. 2014, 2016). To our knowledge, this is the first time that C. australis has been observed on Sieversian wormwood in China. The dodder, as a pest plant, may inevitably limit the availability of the wild Sieversian wormwood resource.

Phylogeny and biogeography of Polygala (Polygalaceae)

AbstractWe estimated phylogenetic relationships of Polygala inferred from nrITS, matK‐trnK, trnL intron, trnL‐trnF intergenic spacer, and rbcL sequence data, with emphasis on the New World clade. Our results did not support the New World and Old World clades as sister groups in Polygala. Relationships among Polygala and allied genera are being discussed, with the recovered clades compared to those of the current taxonomic delimitations. A biogeographic analysis using an ultrametric phylogenetic tree and calibrated with the fossil Paleosecuridaca as well as a reconstruction of the ancestral area are also provided.

Молекулярно-генетический анализ Malus sieversii – сравнение Джунгарской популяции in situ и ex situ

Malus sieversii (Ledeb.) M. Roem. is recognized as the ancestor of the cultivated apple tree. It has a valuable gene pool, and in the last decades, it becomes endangered in Kazakhstan. The genetic diversity of 13 clone-varieties of Sievers apple tree (Malus sieversii) of Dzungarian population from the introduction collection of the Main Botanical Garden in Almaty (Kazakhstan) and 31 samples from three natural populations were assessed using eight polymorphic ISSR markers. The phylogenetic relationship of the clone-varieties with the natural population was estimated using ribosomal (ITS) and chloroplast DNA (rps16 intron, trnL intron, trn L-trn F). The data revealed a high genetic diversity of various clones and samples of Sievers apple tree of the Dzungarian populations. As a result of phylogenetic and cluster analysis the studied clones and samples of M. sieversii from natural populations formed a cluster, samples of supposedly hybrid origin formed 2 clusters, and no clustering was detected depending on geographical distance. The data demonstrate that there is an active information exchange between the populations.

Three lonely Argentines: Toward a new generic delimitation in Polygalaceae

Three enigmatic lineages of the Polygalaceae endemic to the semi‐arid region of Argentina were sampled within the framework of a molecular phylogenetic study of its genera. The regions nrITS, matK, the trnL intron, the trnL‐trnF intergenic spacer, and rbcL were analyzed, and the history of some morphological characters was reconstructed in the family. Results of our phylogenetic analyses and morphological study support the recognition of three monotypic genera historically treated in the genera Bredemeyera and Polygala. A new genus, Rhamphopetalum, and a new combination, R. microphyllum, are proposed together with resurrection of Hualania and Monrosia. The emergence of two species (Bredemeyera colletioides, B. microphylla) as new genera resolves the polyphyletic status of Bredemeyera. Relationships and morphological characters among the genera of Polygalaceae are discussed. Provided here are photographic plates of Hualania colletioides, Monrosia pterolopha, and Rhamphopetalum microphyllum habits, with details of floral structures, a distribution map, typifications, and taxonomic keys for the tribes of Polygalaceae and genera within the tribe Polygaleae.

Encyclia, Epidendrum, or Prosthechea? Clarifying the Phylogenetic Position of a Rare Amazonian Orchid (Laeliinae-Epidendroideae-Orchidaceae)

Abstract—The phylogenetic position of a rare orchid species, which was described by Mansfeld from the Brazilian Amazon, is discussed. Although originally described under Epidendrum despite having a column that is not fully adnate to the lip claw, a feature not common in this genus, Epidendrum apuahuense has been transferred to Encyclia and Prosthechea by other authors. About 90 yr after its description, we found a new population of this species in Brazil, which allowed us to study its taxonomy. A phylogenetic reconstruction was performed using nuclear (ITS rDNA) and plastid (matK exon, rpl32-trnL spacer, trnL intron, and trnL-trnF spacer) datasets, aiming to clarify the generic placement of this taxon. Our results strongly support keeping E. apuahuense under Epidendrum. We also neotypify this name, since the original specimens were destroyed and no illustration is available. The morphological affinities of E. apuahuense and allied taxa are discussed.

A molecular phylogeny of the genus Drimia (Asparagaceae: Scilloideae: Urgineeae) in India inferred from non-coding chloroplast and nuclear ribosomal DNA sequences

The evolutionary history of the medicinally important bulbous geophyte Drimia (subfamily: Scilloideae) has long been considered as a matter of debate in the monocot systematics. In India the genus is represented by a species complex, however, the taxonomic delimitation among them is ill-defined till date. In the present study, a comprehensive phylogenetic relationship among Indian species of this genus has been inferred for the first time based on chloroplast DNA trnL intron, rps16-trnK intergenic spacer, atpB-rbcL intergenic spacer and ribosomal DNA ITS1-5.8S-ITS2 sequences, leaf morphology, anatomy, stomatal characteristics and pollen exine ornamentations. The present findings revealed the monophyletic origin of the Indian members of Drimia and grouped them into two possible lineages (clade- I and II). The phylogenetic tree based on cpDNA concatenated sequences further resolved the clade-I into two distinct subclades (I and II) and clarified the intraspecies relationship among the studied members. The present study suggested a strong relationship between the molecular phylogeny and the morphological characteristics of the species studied. A possible trend of evolution of two important traits: ‘type of palisade cells’ in leaf and ‘pollen exine patterns’ among the members of Drimia in India was also suggested.

Phylogenetic patterns and molecular evolution among ‘True citrus fruit trees’ group (Rutaceae family and Aurantioideae subfamily)

Cytoplasmic chloroplast DNAs was explored to establish phylogenetic relationships among ‘True citrus’ group and elucidate its molecular evolution. Conserved group I intron (PQRS) sequence motifs were detected in Citrus genus and showed differences of nucleotide variation between elements and some species. The P8 stem-loop region was the only variable region in trnL intron of Citrus species. The deletion of 6 bp has been detected between sites (40–45) in the partial alignment of P8 stem-loop resulted in a change in conformation of the secondary structures in the ancestral species and other derived species and this could be explained by the evolution and speciation of this genus. The consistency index (CI=1) and the retention index (RI=1) values suggest that the changes are mostly apomorphic despite the absence of homoplasy (HI=0). The phylogenetic tree confirms and shows a branching of C. limon, C. sinensis, C. paradisi, C. tangelo and C. aurantium in a monophyletic clade. This result so far to C. reticulata ruled out the maternal contribution of mandarin gene pool in the elaboration of these secondary Citrus species. The Maximum Likelihood tree indicated that the genus Citrus is quite distant from the related genera Poncirus, Microcitrus, Eremocitrus and Fortunella except only Fortunella margarita species. Here new insight is given about mechanism involving in the evolution of the trnL intron in ‘True Citrus’. From the study it appears that an explicit rejection of the null hypothesis and a scenario of a demography history and recent expansion of ‘True Citrus’ genotypes seem to be retained that may be an important factor influencing the power of genetic diversity in ‘True Citrus’.

A Useful Technical Application of the Identification of Nucleotide Sequence Polymorphisms and Gene Resources for Cinnamomum osmophloeum Kaneh. (Lauraceae)

The plant genus Cinnamomum contains economically important evergreen aromatic trees and shrubs belonging to the laurel family, Lauraceae. Our study tree species Cinnamomum osmophloeum Kaneh. (CO) has high economic value in Taiwan. The present study attempts to identify the gene resources of Cinnamomum osmophloeum Kaneh. by analyzing the nucleotide sequences of the partial noncoding internal transcribed spacer 2 (pITS2) of the ribosomal DNA and the trnL-trnF chloroplast genome. Seventy-three geographical strains of Cinnamomum osmophloeum, preserved in the Lien Hua-Chin Research Center of the Forestry Research Institute and the Hua-Lin Forestry Center of Chinese Culture University, were collected and analyzed by PCR amplification and DNA sequencing to study the genetic diversity and nucleotide sequence polymorphisms of the tested specimens. Our results allowed us to accurately identify the lineage of Cinnamomum osmophloeum and to conclude that the strains belonging to the Lien Hua-Chin Research Center had much higher genetic diversity than those preserved in the Hua-Lin Forestry Center. Multiple sequence alignments demonstrated that the variability of the nucleotide sequence polymorphisms for the pITS2 region was higher than those of the trnL intron and trnL-trnF intergenic spacer (IGS) regions among the 73 tested specimens of Cinnamomum osmophloeum. Cluster analyses, using the neighbor-joining and maximum parsimony methods, for the 73 tested geographical strains of Cinnamomum osmophloeum and species of Cinnamomum registered in the GenBank and EMBL databases were performed to demonstrate the genus and species distribution of the samples. Here, we describe the use of pITS2 polymorphisms as a genetic classifier and report the establishment of a DNA sequence database for CO gene resource identification. The sequence database described in this study can be used to identify CO specimens at the inter- or intraspecies level using pITS2 DNA sequences, which illustrates its value in gene resource identification. Our study results can be used further for correctly identifying the true Cinnamomum osmophloeum Kaneh.

Expanding and testing fluorescent amplified fragment length polymorphisms for identifying roots of boreal forest plant species.

Premise of the StudyIdentifying roots to species is challenging, but is a common problem in ecology. Fluorescent amplified fragment length polymorphisms (FAFLPs) can distinguish species within a mixed sample, are high throughput, and are inexpensive. To broaden the use of this tool across ecosystems, unique size profiles must be established for species, and its limits identified.MethodsFragments of three noncoding cpDNA regions were used to create size profiles for 193 species common to the western Canadian boreal forest. We compared detection success among congeners using FAFLPs and Sanger sequencing of the trnL intron. We also simulated and experimentally created communities to test the influence of species richness, cpDNA regions used, and extraction/amplification biases on detection success.ResultsOf the 193 species, 54% had unique size profiles. This value decreased when fewer cpDNA regions were used. In simulated communities, ambiguous species identifications were positively related to the species richness of the community. In mock communities, some species evaded detection owing to poor extraction or amplification. Sequencing did not increase detection success compared to FAFLPs for a subset of 24 species across nine genera.DiscussionWe recommend FAFLPs are best suited to confirm rather than discover species occurring belowground.

Short Communication: Species identity and taxonomical position of selected species of Annonaceae based on trnL molecular marker

Abstract. Lestari DA, Azrianingsih R. 2019. Species identity and taxonomical position of selected species of Annonaceae based on trnL molecular marker. Biodiversitas 20: 1012-1019. Identification based on morphological characters could be difficult when some characters are not visible. The absence of some morphological characters could effect for identification of species identity and taxonomical position of the species. Confirmation by the DNA data is needed to support species identification. The sequence was used in this research is trnL intron as non-coding sequence DNA, based on chloroplast DNA. The research aimed to estimate of species identity and determine of taxonomical position of species of Annonaceae based on trnL sequences. Methods were used through steps of DNA extraction, DNA amplification, DNA sequencing, and data analysis to selected species of Annonaceae from Purwodadi Botanic Garden (PBG), the trnL intron sequences of 10 Annonaceae species from GenBank database and two species of Magnoliaceae as out-group. Results showed that trnL sequence as non-coding gene explains the different groupings with the previous groupings in Annonaceae to this observed species. trnL sequence can estimate of species identity as much as 30%, caused by changes of nucleotide bases from mutation and missing data. Polymorphism of DNA sequences showed that 61.18% sites as conserved region, 24.05% sites as polymorphic variation and 14.76% sites as alignment gaps. Oxymitra sp. is in group (monophyletic) with Mitrephora javanica because they are genetically in close relationship (Uvariae tribes and Annonoideae sub-family), Popowia sp. is in group with Orophea enterocarpa, because they are genetically in close relationship (Miliuseae tribes and Malmeoideae sub-family).

Molecular Phylogeography and Ecological Niche Modeling of Sibbaldia procumbens s.l. (Rosaceae).

The phylogeographical analysis and ecological niche modeling (ENM) of the widely distributed Northern Hemisphere Sibbaldia procumbens s.l. can help evaluate how tectonic motion and climate change helped shape the current distribution patterns of this species. Three chloroplast regions (the atpI-atpH and trnL-trnF intergenic spacers and the trnL intron) were obtained from 332 (156 from present study and 176 from the previous study) individuals of S. procumbens s.l. An unrooted haplotype network was constructed using the software NETWORK, while BEAST was used to estimate the divergence times among haplotypes. ENM was performed by MAXENT to explore the historical dynamic distribution of S. procumbens s.l. The haplotype distribution demonstrates significant phylogeographical structure (NST > GST; P < 0.01). The best partitioning of genetic diversity by SAMOVA produced three groups, while the time to the most recent common ancestor of all haplotypes was estimated to originate during the Miocene, with most of the haplotype diversity having occurred during the Quaternary. The MAXENT analysis showed S. procumbens s.l. had a wider distribution range during the last glacial maximum and a narrower distribution range during the last interglacial, with predictions into the future showing the distribution range of S. procumbens s.l. shrinking.

Evolutionary history of the Pasque-flowers (Pulsatilla, Ranunculaceae): Molecular phylogenetics, systematics and rDNA evolution

Pulsatilla (Anemoneae, Ranunculaceae) is sister to Anemone s.s. and contains ca 40 perennial species of considerable horticultural and medical importance. We sequenced 31 of those species, plus nine subspecies, two cultivars and six outgroups, for two nuclear regions (high-copy nrITS and low-copy MLH1) and three plastid regions (rbcL, accD–psaI, trnL intron) in order to generate the first comprehensive species-level phylogeny of the genus. Phylogenetic trees were constructed using both concatenation-based (maximum likelihood and Bayesian inference) and coalescence methods. The better supported among the internal nodes were subjected to molecular clock dating and ancestral area reconstruction, and karyotypic characters identified by us using Fluorescence In Situ Hybridization were mapped across the tree. The preferred species tree from the coalescence analysis formed the basis of a new infrageneric classification based on monophyly plus degree of divergence. The earliest divergent of the three subgenera, Kostyczewianae, is represented by only a single species that is morphologically intermediate between Anemone s.s. and ‘core’ Pulsatilla. Subgenus Pulsatilla is considerably richer in species than its sister subgenus Preonanthus and contains three monophyletic sections. Species possessing nodding flowers and pectinately dissected leaves are phylogenetically derived compared with groups possessing erect flowers and palmately lobed leaves. Pulsatilla separated from Anemone s.s. at ca 25 Ma. Our results indicate a central Asian mountain origin of the genus and an initial diversification correlated with late Tertiary global cooling plus regional mountain uplift, aridification and consequent expansion of grasslands. The more rapid and extensive diversification within subgenus Pulsatilla began at ca 3 Ma and continued throughout the Quaternary, driven not only by major perturbations in global climate but also by well-documented polyploidy.

Belowground plant parts are crucial for comprehensively estimating total plant richness in herbaceous and woody habitats.

Most studies consider aboveground plant species richness as a representative biodiversity measure. This approach inevitably assumes that the partitioning of total plant species richness into above- and belowground components is constant or at least consistent within and across vegetation types. However, with studies considering belowground plant richness still scarce and completely absent along vegetation gradients, this assumption lacks experimental support. Novel DNA sequencing techniques allow economical, high-throughput species identification of belowground environmental samples, enabling the measurement of the contributions of both above- and belowground plant components to total plant richness. We investigated above- and belowground plant species richness in four vegetation types (birch forest, heath, low alpine tundra, high alpine tundra) at the scale of herbaceous plant neighborhoods (dm) using 454 sequencing of the chloroplast trnL (UAA) intron to determine the plant species richness of environmental root samples and combined it with aboveground data from vegetation surveys to obtain total plant species richness. We correlated the measured plant species richness components with each other and with their respective plant biomass components within and across vegetation types. Total plant species richness exceeded aboveground richness twice on average and by as much as three times in low alpine tundra, indicating that a significant fraction of belowground plant richness cannot be recorded aboveground. More importantly, no consistent relationship among richness components (above- and belowground) was found within or across vegetation types, indicating that aboveground richness alone cannot predict total plant richness in contrasting vegetation types. Finally, no consistent relationship between plant richness and the corresponding biomass component was found. Our results clearly show that aboveground plant richness alone is a poor estimator of total plant species richness within and across different vegetation types. Consequently, it is crucial to account for belowground plant richness in future plant ecological studies in order to validate currently accepted plant richness patterns, as well as to measure potential changes in plant community composition in a changing environment.

Rapid detection of several endangered agarwood-producing Aquilaria species and their potential adulterants using plant DNA barcodes coupled with high-resolution melting (Bar-HRM) analysis

Abstract Aquilaria is an endangered agarwood-producing genus that is currently protected by international laws. The agarwood trade is strictly monitored to prevent illegal harvesting, which has caused high demand for this natural product. Other plant sources of similar appearance or fragrance as agarwood are used as adulterant species in counterfeit products. To promote species identification via the DNA barcoding technique, the existing DNA barcoding database in our laboratory was enriched with seven plant barcoding sequences from a commercially important Aquilaria species (Aquilaria beccariana) and seven adulterant species (Cocos nucifera, Dalbergia latifolia, Pinus contorta var. latifolia, Santalum album, Strychnos ignatii, Thuja sp. and Terminalia catappa). DNA barcoding with high-resolution melting analysis (Bar-HRM) showed that the mini-barcode internal transcribed spacer 1 (ITS1) was an effective gene locus that allows for a rapid and species-specific detection of Aquilaria and their adulterants, while four other mini-barcodes (rbcL, trnL intron, ITS2 and 5.8s) functioned as a support and a crosscheck for the barcoding results. The accuracy of the Bar-HRM technique in species origin identification was further assessed with seven agarwood blind specimens. The Bar-HRM technique is a potential tool for validating agarwood-species origin and detecting products with adulterant species.

Phylogenetic Relationships and Morphological Evolution in the Carnivorous Genus Philcoxia (Plantaginaceae, Gratioleae)

Philcoxia (Plantaginaceae, Gratioleae) is a genus with seven species of small herbs endemic to Brazil. It is characterized by underground stems and petioles, peltate leaves, and zigzag racemose inflorescences, in addition to a remarkable carnivorous syndrome. The phylogenetic placement of Philcoxia within tribe Gratioleae has been uncertain since its discovery due to its unique morphology and limited data from phylogenetic studies. We tested the phylogenetic placement of Philcoxia within Gratioleae, increasing the number of species sampled within both the genus and the rest of the tribe, and sequencing four regions from cpDNA (rpl16, rps16, and trnL introns and the trnL-trnF intergenic spacer) and one from nrDNA (ITS1 spacer); we used both parsimony and Bayesian inference to reconstruct the phylogeny. Moreover, we assessed both the monophyly of the genus and interspecific relationships. Finally, we performed ancestral character state reconstructions of 10 morphological characters to infer synapomorphies for the genus and specific clades within it. Philcoxia was recovered as monophyletic, with maximum support values, as sister to Stemodia stellata. Five morphological character states were reconstructed as potential synapomorphies for the Philcoxia clade (rosetted herb habit, underground stems and petioles, floral resupination, number of stamens reduced to two, and number of fertile thecae per anther reduced to one), whereas fourwere reconstructed as potential synapomorphies for specific clades within the genus (peltate leaves, irregular rosettes, vegetative propagation by rhizomes, and presence of tubers). Further anatomical studies, in association with ecological data, would provide insight into the structure of the underground systems and its adaptive significance, and would further elucidate the evolution of carnivory in Philcoxia. Our study highlights the need to reassess the concepts and circumscriptions of some currently accepted genera of the Gratioleae such as Bacopa, Conobea, and Stemodia, and for the tribe Angelonieae.

Evolutionary history of the tribe Astereae in the Flora Iranica area: Systematic implications

The present study reconstructs the phylogenetic relationships of the tribe with emphasis on Psychrogeton using both nuclear (ITS, ETS) and chloroplast (trnL intron and trnL-trnF intergenic spacer) markers. Divergence times for main lineages were estimated using BEAST analysis. Based on results of molecular analyses, tribe Astereae is circumscribed here as containing 16 genera and 38 species and comprising strongly supported five major clades: Aster, Chamaegeron, Erigeron, Galatella and Psychrogeton. Among the Eurasian asteroid taxa, Aster bachtiaricus formed the basal most diverging lineage far distantly from the Aster s.str. clade. Lachnophllum and Chamaegeron are sister taxa in nuclear tree, although weakly united in plastid topology. Galatella with the inclusion of Crinitina (= Crinitaria) constitutes a well-supported clade, which along with Tripolium forms the Galatella group. Eurasian Erigeron species were derived within the Erigeron clade. Erigeron uniflorus subsp. daenensis and subsp. elborsensis are distinct from the type subspecies and are resurrected here as species in their own right. Our analyses of the datasets revealed that all examined species of Psychrogeton, except P. obovatus, emerged in a single clade comprising four distinct subclades. Molecular dating analyses indicate that tribe Astereae originated in the Late Eocene at 38.6 Ma. The most genera of Astereae diverged during the Middle Miocene whereas the diversification of lineages began mostly through the Pliocene and Pleistocene. On the basis of the molecular data as well as the morphological characteristics, Aster bachtiaricus was elevated to the generic rank and this new monospecific genus was named Iranoaster. Psychrogeton obovatus was treated as a member of the recently established genus Neobrachyactis. The conflicting position of some taxa including Lachnophyllum gossypinum, Dichrocephala, Myriactis and Asterothamnus in nuclear and plastid trees might be the result of ancient hybridization/introgression events.