Molecular authentication of commercial "Qian-hu" through the integration of nrDNA internal transcribed spacer 2 and nucleotide signature.

BACKGROUND: Peucedani Radix, also known as "Qian-hu" is a traditional Chinese medicine derived from Peucedanum praeruptorum Dunn. It is widely utilized for treating wind-heat colds and coughs accompanied by excessive phlegm. However, due to morphological similarities, limited resources, and heightened market demand, numerous substitutes and adulterants of Peucedani Radix have emerged within the herbal medicine market. Moreover, Peucedani Radix is typically dried and sliced for sale, rendering traditional identification methods challenging. MATERIALS AND METHODS: We initially examined and compared 104 commercial "Qian-hu" samples from various Chinese medicinal markets and 44 species representing genuine, adulterants or substitutes, utilizing the mini barcode ITS2 region to elucidate the botanical origins of the commercial "Qian-hu". The nucleotide signature specific to Peucedani Radix was subsequently developed by analyzing the polymorphic sites within the aligned ITS2 sequences. RESULTS: The results demonstrated a success rate of 100% and 93.3% for DNA extraction and PCR amplification, respectively. Forty-five samples were authentic "Qian-hu", while the remaining samples were all adulterants, originating from nine distinct species. Peucedani Radix, its substitutes, and adulterants were successfully identified based on the neighbor-joining tree. The 24-bp nucleotide signature (5'-ATTGTCGTACGAATCCTCGTCGTC-3') revealed distinct differences between Peucedani Radix and its common substitutes and adulterants. The newly designed specific primers (PR-F/PR-R) can amplify the nucleotide signature region from commercial samples and processed materials with severe DNA degradation. CONCLUSIONS: We advocate for the utilization of ITS2 and nucleotide signature for the rapid and precise identification of herbal medicines and their adulterants to regulate the Chinese herbal medicine industry.

Identification of Angelica acutiloba, A. sinensis, and other Chinese medicinal Apiaceae plants by DNA barcoding.

Crude drug Angelicae acutilobae radix is one of the most important crude drugs in Japanese traditional medicine and is used mainly for the treatment of gynecological disorders. In the listing in the Japanese Pharmacopoeia XVIII, Angelicae acutilobae radix is defined as the root of Angelica acutiloba (Apiaceae), which has long been produced on an industrial scale in Japan. With the aging of farmers and depopulation of production areas, the domestic supply has recently declined and the majority of the supply is now imported from China. Due to having only slightly different morphological and chemical characteristics for the Apiaceae roots used to produce dried roots for Chinese medicines, the plant species originating the crude drug Apiaceae roots may be incorrectly identified. In particular, Angelicae sinensis radix, which is widely used in China, and Angelicae acutilobae radix are difficult to accurately identify by morphology and chemical profiles. Thus, in order to differentiate among Angelicae acutilobae radix and other radixes originated from Chinese medicinal Apiaceae plants, we established DNA markers. Using DNA sequences for the chloroplast psbA-trnH intergenic spacer and nuclear internal transcribed spacer regions, Angelicae acutilobae radix and other Chinese Apiaceae roots, including Angelicae sinensis radix, can be definitively identified.

The complete plastomes of seven Peucedanum plants: comparative and phylogenetic analyses for the Peucedanum genus.

BACKGROUND: The Peucedanum genus is the backbone member of Apiaceae, with many economically and medically important plants. Although the previous studies on Peucedanum provide us with a good research basis, there are still unclear phylogenetic relationships and many taxonomic problems in Peucedanum, and a robust phylogenetic framework of this genus still has not been obtained, which severely hampers the improvement and revision of taxonomic system for this genus. The plastid genomes possessing more variable characters have potential for reconstructing a robust phylogeny in plants. RESULTS: In the current study, we newly sequenced and assembled seven Peucedanum plastid genomes. Together with five previously published plastid genomes of Peucedanum, we performed a comprehensively comparative analyses for this genus. Twelve Peucedanum plastomes were similar in terms of genome structure, codon bias, RNA editing sites, and SSRs, but varied in genome size, gene content and arrangement, and border of SC/IR. Fifteen mutation hotspot regions were identified among plastid genomes that can serve as candidate DNA barcodes for species identification in Peucedanum. Our phylogenetic analyses based on plastid genomes generated a phylogeny with high supports and resolutions for Peucedanum that robustly supported the non-monophyly of genus Peucedanum. CONCLUSION: The plastid genomes of Peucedanum showed both conservation and diversity. The plastid genome data were efficient and powerful for improving the supports and resolutions of phylogeny for the complex Peucedanum genus. In summary, our study provides new sights into the plastid genome evolution, taxonomy, and phylogeny for Peucedanum species.

Identifying the compounds that can distinguish between Saposhnikovia root and its substitute, Peucedanum ledebourielloides root, using LC-HR/MS metabolomics.

Previously, we established a 1H NMR metabolomics method using reversed-phase solid-phase extraction column (RP-SPEC), and succeeded in distinguishing wild from cultivated samples of Saposhnikoviae radix (SR), and between SR and its substitute, Peucedanum ledebourielloides root (PR). Herein, we performed LC-HR/MS metabolomics using fractions obtained via RP-SPEC to identify characteristic components of SR and PR. One and three characteristic components were respectively found for SR and PR; these components were isolated with their m/z values and retention times as a guide. The characteristic component of SR was identified as 4'-O-ß-D-glucosyl-5-O-methylvisamminol (1), an indicator component used to identify SR in the Japanese Pharmacopoeia. In contrast, the characteristic components of PR were identified as xanthalin (2), 4'-O-ß-D-apiosyl (1 → 6)-ß-D-glucosyl-5-O-methylvisamminol (3), and 3'-O-ß-D-apiosyl (1 → 6)-ß-D-glucosylhamaudol (4) based on spectroscopic data such as 1D- and 2D-NMR, MS, and specific optical rotation. Among them, 4 is a novel compound. For the correlation between the NMR metabolomics results in the present and our previous report, only 1 and 2 were found to correlate with the chemical shifts, and the other compounds had no correlation. As the chemical shifts for compounds 1, 3, and 4 were similar to each other, especially for the aglycone moiety, they could not be distinguished because of the sensitivity and resolution of 1H NMR. Accordingly, combining NMR and LC/MS metabolomics with their different advantages is considered useful for metabolomics of natural products. The series of methods used in our reports could aid in quality evaluations of natural products and surveying of marker components.

Mining genes associated with furanocoumarin biosynthesis in an endangered medicinal plant, Glehnia littoralis.

The endangered medicinal plant Glehnia littoralis is one of the important natural source of furanocoumarin, which has been used as mucolytic, antitussive, antitumour and antibacterial. However, the genetic information of furanocoumarin biosynthesis in G. littoralis is scarce at present. The objective of this study was to mine the putative candidate genes involved in the biosynthesis pathwayof furanocoumarin and provide references for gene identification, and functional genomics of G. littoralis. We carried out the transcriptome analysis of leaves and roots in G. littoralis, which provided a dataset for gene mining. Psoralen, imperatorin and isoimperatorin were detected in G. littoralis by high performance liquid chromatography analysis. Candidate key genes were mined based on the annotations and local BLAST with homologous sequences using BioEdit software. The relative expression of genes was analysed using quantitative real-time polymerase chain reaction. Further, the CYP450 genes were mined using phylogenetic analyses using MEGA 6.0 software. Atotal of 156,949 unigenes were generated, of which 9021 were differentially-expressed between leaves and roots. A total of 82 unigenes encoding eight enzymes in furanocoumarin biosynthetic pathway were first obtained. Seven genes that encoded key enzymes in the downstream furanocoumarin biosynthetic pathway and expressed more in roots than leaves were screened. Twenty-six candidate CYP450 unigenes expressed abundantly in roots and were chiefly concentrated in CYP71, CYP85 and CYP72 clans. Finally, we filtered 102 differentially expressed transcription factors (TFs) unigenes. The transcriptome of G. littoralis was characterized which would help to elucidate the furanocoumarin biosynthetic pathway in G. littoralis and provide an invaluable resource for further study of furanocoumarin.

Rapid and intelligent discrimination of Notopterygium incisum and Notopterygium franchetii by infrared spectroscopic fingerprints and electronic olfactory fingerprints.

This preliminary research evaluated mid-infrared (MIR) spectroscopy, near-infrared (NIR) spectroscopy and electronic nose (E-nose) for the rapid identification of Notopterygium incisum and Notopterygium franchetii, which were both approved sources of Notopterygii Rhizoma et Radix (Chinese Pharmacopoeia, 2015) but possessed different chemical compositions and pharmacological activities. At the level of single variables, MIR showed quite a few discriminating peaks in the regions of 3000-2800 cm-1 (the stretching bands of CH), 1770-1670 cm-1 (the stretching bands of CO), and 1400-1200 cm-1 (the bending bands of CH and the stretching bands of CO). Meanwhile, NIR only showed an intuitive discriminating peak near 4736 cm-1 (the combination band of OH and CO stretching modes). E-nose response signals of N. incisum and N. franchetii were significant different (p < 0.05) on four sensors, i.e., LY2/LG, LY2/GH, LY2/gCT and LY2/gCTI. Using the infrared spectra or E-nose sensor responses as fingerprints, support vector machine (SVM) models can provide good recognition accuracy (100% for MIR and NIR models, 92.9% for E-nose model). This research indicated the feasibility of MIR, NIR and E-nose for the accurate, rapid, cheap and green identification of N. incisum and N. franchetii, which was desirable to assure the authenticity, efficacy and safety of related herb materials and products.

Chemical constituents from Glehnia littoralis and their chemotaxonomic significance.

Phytochemical investigation of the roots of Glehnia littoralis Fr. Schmidt. ex Miq. led to the isolation of 16 known compounds, including three ß-carboline alkaloids (1-3), four phenylpropanoids (4-7), five phenolic acids (8-12), three polyacetylenes (13-15) and one fatty acid (16). The structures of these compounds were elucidated on the basis of spectral analysis and comparison with those reported in literatures. To the best of knowledge, the report of the first ß-carboline alkaloid in the Umbelliferae family. Additionally, compounds 1-5, 9, 10 and 16 have not been reported from any species in Umbelliferae family, compounds 7, 8 and 12 were isolated from the genus Glehnia for the first time and could be of the chemotaxinomic significance and serve as valuable chemotaxonomic makers for G. littoralis. The chemotaxonomic significance of the isolated compounds was summarised.

A Single Standard to Determine Multi-Components Method Coupled with Chemometric Methods for the Quantification, Evaluation and Classification of Notopterygii Rhizoma et Radix from Different Regions.

An ultra-high-performance liquid chromatography-quadrupole/time of flight mass spectrometry is used to identify 33 compounds in Notopterygii rhizoma and radix, after which a single standard to determine multi-components method is established for the simultaneous determination of 19 compounds in Notopterygii rhizoma and radix using chlorogenic acid and notopterol as the internal standard. To screen the potential chemical markers among Notopterygii rhizoma and radix planted in its natural germination area and in others, the quantitative data of 19 compounds are analyzed via partial least-squares discriminant analysis (PLS-DA). Depending on the variable importance parameters (VIP) value of PLS-DA, six compounds are selected to be the potential chemical markers for the discrimination of Notopterygii rhizoma and radix planted in the different regions. Furthermore, the Fisher's discriminant analysis is used to build the models that are used to classify Notopterygii rhizoma and radix from the different regions based on the six chemical markers. Experimental results indicate that Notopterygii rhizoma and radix planted in the Sichuan province are distinguished successfully from those in other regions, reaching a 96.0% accuracy rating. Therefore, a single standard to determine multi-components method combined with a chemometrics method, which contains the advantages such as simple, rapid, economical and accurate identification, offers a new perspective for the quantification, evaluation and classification of Notopterygii rhizoma and radix from the different regions.

New insights into the chemical profiling, cytotoxicity and bioactivity of four Bunium species.

Bunium species have been reported to be used both as food and in traditional medicines. The scientific community has attempted to probe into the pharmacological and chemical profiles of this genus. Nonetheless, many species have not been investigated fully to date. In this study, we determined the phenolic components, antimicrobial, antioxidant, and enzyme inhibitory activities of aerial parts of four Bunium species (B. sayai, B. pinnatifolium, B. brachyactis and B. macrocarpum). Results showed that B. microcarpum and B. pinnatifolium were strong antioxidants as evidenced in the DPPH, ABTS, CUPRAC, and FRAP assays. B. brachyactis was the most effective metal chelator, and displayed high enzyme inhibition against cholinesterase, tyrosinase, amylase, glucosidase, and lipase. The four species showed varied antimicrobial activity against each microorganism. Overall, they showed high activity against P. mirabilis and E. coli (MIC and MBC <1 mg mL-1). B. brachyactis was more effective against Aspergillus versicolor compared to the standard drug ketoconazole. B. brachyactis was also more effective than both ketoconazole and bifonazole against Trichoderma viride. B. sayai was more effective than ketoconazole in inhibiting A. fumigatus. B. sayai was most non-toxic to HEK 293 (cellular viability = 117%) and HepG2 (cellular viability = 104%). The highest level of TPC was observed in B. pinnatifolium (35.94 mg GAE g-1) while B. microcarpum possessed the highest TFC (39.21 mg RE g-1). Seventy four compounds were detected in B. microcarpum, 70 in B. brachyactis, 66 in B. sayai, and 51 in B. pinnatifolium. Quinic acid, chlorogenic acid, pantothenic acid, esculin, isoquercitrin, rutin, apigenin, and scopoletin were present in all the four species. This study showed that the four Bunium species are good sources of biologically active compounds with pharmaceutical and nutraceutical potential.

Dynamic Chloroplast Genome Rearrangement and DNA Barcoding for Three Apiaceae Species Known as the Medicinal Herb "Bang-Poong".

Three Apiaceae species Ledebouriella seseloides, Peucedanum japonicum, and Glehnia littoralis are used as Asian herbal medicines, with the confusingly similar common name "Bang-poong". We characterized the complete chloroplast (cp) genomes and 45S nuclear ribosomal DNA (45S nrDNA) sequences of two accessions for each species. The complete cp genomes of G. littoralis, L. seseloides, and P. japonicum were 147,467, 147,830, and 164,633 bp, respectively. Compared to the other species, the P. japonicum cp genome had a huge inverted repeat expansion and a segmental inversion. The 45S nrDNA cistron sequences of the three species were almost identical in size and structure. Despite the structural variation in the P. japonicum cp genome, phylogenetic analysis revealed that G. littoralis diverged 5-6 million years ago (Mya), while P. japonicum diverged from L. seseloides only 2-3 Mya. Abundant copy number variations including tandem repeats, insertion/deletions, and single nucleotide polymorphisms, were found at the interspecies level. Intraspecies-level polymorphism was also found for L. seseloides and G. littoralis. We developed nine PCR barcode markers to authenticate all three species. This study characterizes the genomic differences between L. seseloides, P. japonicum, and G. littoralis; provides a method of species identification; and sheds light on the evolutionary history of these three species.

Investigating Schizocarp morphology as a taxonomic tool in study of Apiaceae family by utilizing LM and SEM techniques.

In present study, the schizocarp morphology of 14 species belonging to Apiaceae family has been investigated. Light microscopy (LM) and scanning electron microscopy (SEM) have been utilized to highlight qualitative and quantitative features of studied species. Variations have been observed in macro- and micro-morphological features such as color, shape, symmetry, length, width, apex, epicuticular projections, surface patterns, anticlinal, and periclinal wall patterns. Schizocarp shapes observed were oval, round, triangular, linear, elliptic, and globose. Fruit was either homomorphic or heteromorphic. Crystalloids, stellate hair, multicellular spines, and platelets were mostly observed epicuticular projections. Surface patterns on the fruit surface were striate, rugulate-striate, reticulate, and striato-knotted. Both macro- and micro-morphological characters can serve as an important tool in classifying Apiaceae family at various taxonomic ranks. Substantial variations observed can assist as useful constraints at various taxonomic levels as they provide reliable and constant details. Disparities observed in schizocarp features can pave a path for Apiaceae family classification based on phylogenetic and molecular studies.

[Collection and sorting of medicinal plants in Chinese Apiaceae( Umbelliferae)].

The family Apiaceae( Umbelliferae) includes some of the world's most important medicinal plants,with more than 100 species recorded in the traditional Chinese medicine,of which more than ten species are commonly used medicinal materials. However,due to morphological similarities,high market demands and regional factors,substitutes and adulterants are often mixed with genuine in the medicinal market. Therefore,a comprehensive sorting for these poorly known plants has been done in this study by combining market survey with literature review,including its species,distribution,price and substitutes. According to the statistics,there are 65 genera and 262 species of medicinal plants of Apiaceae in China,with medicinal part mostly from radix and rhizoma. Sichuan province is the most abundant in distribution and planting resources,with about 137 species,followed by Yunnan,Hubei and Gansu provinces.Furthermore,we summarized the genuine and substitutes of 11 medicinal plants,e. g. Bupleurum,Angelica and Peucedanum etc.,which found that the medicinal plants of Apiaceae were substituted or mixed in different taxonomic ranks. This study would contribute to reduce the risk of medicine misuse,as well as explore other plants of Apiaceae with potential medicinal value,to achieve sustainable development of related industries.

Isolation and cross-amplification of the first set of polymorphic microsatellite markers of two high-Andean cushion plants.

In the southern Andes mountains (27-39◦S) Azorella madreporica and Laretia acaulis, two Apiaceae cushion plant species commonly known as yaretas, conform a well-established altitudinal vegetation belt along the lower Andean zone. These species have been considered as fundamental components of several ecological dynamics within their communities; however, high mountain ecosystems are increasingly threatened worldwide by natural and anthropogenic pressures and the southern Andes are not the exception. Recognizing that genetic information is crucial for the success of any conservation or restoration initiative inwild populations, we developed and cross-amplified 28 specifically designed microsatellite markers (14 in A. madreporica and 14 in L. acaulis), and also tested the cross amplification of 25 markers from the related species Azorella selago. In a region which is particularly vulnerable to global change trends, this new polymorphic microsatellite loci will be useful in the study of the genetic diversity of these high-mountain cushion plants, which are pivotal in the structuring of their native ecosystems.

[Study on characteristics and microscopic identification of fruits of Notopterygium franchetii and N. forrestii].

Rhizoma et Radix Notopterygii is a rare and endangered Chinese medicine. In the collection of Notopterygium franchetii fruits, we collected a sample of N. forrestii , which is a spurious breed. Fruits of N. franchetii and N. forrestii are very similar in morphology and can be easily confused. Until now the morphological identification of the fruits of Notopterygium has not been reported. To provide a scientific basis for the identification of N. franchetii and N. forrestii fruits, the morphology and microscopic identification were studied in this paper. In this study, stereomicroscope and paraffin sections were used to compare the morphological characteristics and microscopic characteristics of these two fruits. Our results showed that these two fruits were different in size, surface texture and the number of vertical edges on the back. These traits can be used as diagnostic characteristic of these two fruits. The difference between the number of tubing and the endosperm cell contents can be used as microscopic identification features. The above discriminative characteristics can distinguish the two fruits and provide scientific basis for the identification and germplasm evaluation of Notopterygium fruits.

Rapid discrimination of different Apiaceae species based on HPTLC fingerprints and targeted flavonoids determination using multivariate image analysis.

INTRODUCTION: Species of Apiaceae are used in folk medicine as spices and in officinal medicinal preparations of drugs. They are an excellent source of phenolics exhibiting antioxidant activity, which are of great benefit to human health. Discrimination among Apiaceae medicinal herbs remains an intricate challenge due to their morphological similarity. OBJECTIVE: In this study, a combined "untargeted" and "targeted" approach to investigate different Apiaceae plants species was proposed by using the merging of high-performance thin layer chromatography (HPTLC)-image analysis and pattern recognition methods which were used for fingerprinting and classification of 42 different Apiaceae samples collected from Egypt. METHODOLOGY: Software for image processing was applied for fingerprinting and data acquisition. HPTLC fingerprint assisted by principal component analysis (PCA) and hierarchical cluster analysis (HCA)-heat maps resulted in a reliable untargeted approach for discrimination and classification of different samples. The "targeted" approach was performed by developing and validating an HPTLC method allowing the quantification of eight flavonoids. RESULTS: The combination of quantitative data with PCA and HCA-heat-maps allowed the different samples to be discriminated from each other. CONCLUSION: The use of chemometrics tools for evaluation of fingerprints reduced expense and analysis time. The proposed method can be adopted for routine discrimination and evaluation of the phytochemical variability in different Apiaceae species extracts.

Genome-wide searches and molecular analyses highlight the unique evolutionary path of flavone synthase I (FNSI) in Apiaceae.

Flavone synthase is a key enzyme for flavone biosynthesis and is encoded by two gene families: flavone synthase I (FNSI) and flavone synthase II (FNSII). FNSII is widely distributed in plants, while FNSI has been reported in rice (Oryza sativa) and seven species of Apiaceae. FNSI has likely evolved from the duplication of flavanone 3ß-hydroxylase (F3H). In this study, we used multiple bioinformatics tools to identify putative FNSI and F3H genes from 42 publicly available genome and transcriptome datasets. Results showed that rice FNSI does not share a common ancestral sequence with other known FNSI genes and that FNSI is absent from species outside of Apiaceae. Positive selection site identification analysis revealed that four sites within the FNSI tree branches of Apiaceae evolved under significant positive selection. The putative F3H genes identified in this study provide a valuable resource for further function analysis of flavone synthase.

Rampant polyphyly in the Arracacia clade (Apiaceae) and an assessment of the phylogenetic utility of 20 noncoding plastid loci.

The Arracacia clade (Apiaceae, Apioideae) is a heterogeneous assemblage of 12 genera, comprising 111 known species distributed in high montane temperate and sub-alpine habitats of meso- and South America. Previous studies have indicated that the genera Arracacia, Coulterophytum, and Prionosciadium are polyphyletic, but for the most part relationships among the members of the clade are largely unknown. Initially, cladistic analyses of nrDNA ITS sequences were carried out on 212 accessions (122 taxa), representing 92 species of the Arracacia clade and outgroups from the closely-related páramo genera Cotopaxia, Niphogeton, and Perissocoeleum and members of the Perennial Endemic North American clade and its allies. Using the ITS results to inform sampling of a small subset of taxa, a pilot study examining the phylogenetic utility of 20 noncoding chloroplast loci was subsequently performed to identify those regions most useful at resolving relationships. A cost-benefit analysis determined that five loci (trnQ-5'rps16, trnD-trnT, rpl32-trnL, psbD-trnT, ndhA intron) would maximize resolution and branch support in the clade. Cladistic analyses of four of these loci (trnQ-5'rps16, trnD-trnT, rpl32-trnL, ndhA intron) and the ITS region, separately and combined, revealed that Arracacia, Coaxana, Coulterophytum, Prionosciadium, and Rhodosciadium are each polyphyletic and that Donnellsmithia and Myrrhidendron are each monophyletic. Although most relationships in the Arracacia clade and among the closely-related genera Cotopaxia, Niphogeton, and Perissocoeleum are poorly resolved and supported, ten groups are recognized for future revisionary studies. Polyploidy and rapid species radiation have likely confounded generic circumscriptions and interpretation of relationships.

Chloroplast and ITS phylogenies to understand the evolutionary history of southern South American Azorella, Laretia and Mulinum (Azorelloideae, Apiaceae).

Azorella, Laretia and Mulinum are taxonomically complex, and good candidates to study evolutionary radiations in the Andes and the importance of hybridizations. Previous phylogenetic studies of subfamily Azorelloideae agree that Azorella and Mulinum as currently conceived are not monophyletic, and hence a revision of their circumscription is necessary. However, these phylogenies were based only on chloroplast DNA sequence data. Here, phylogenetic relationships within Azorelloideae were inferred using sequence data from five chloroplast DNA (rps16 intron, trnQ-rps16, rps16-trnKUUU 5' -exon, trnGGCC-trnSGCU and rpL32-trnLUAG), and from nuclear rDNA ITS regions to assess the monophyly of Azorella and Mulinum and discuss generic re-circumscriptions, determine hybridization and radiation events, identify and characterize important lineages, and propose hypotheses on evolution of key morphological characters. In total, 121 accessions of Azorelloideae were analyzed. Phylogenetic analyses of the different genomes were conducted separately and combined, with and without indels, using maximum parsimony, maximum likelihood, and Bayesian methods. To analyze the incongruence between plastid and nuclear-derived trees a consensus network from strongly supported nodes from cpDNA and ITS trees was constructed. Internode certainty values were calculated to evaluate the reliability of the relationships estimated from the individual cpDNA and ITS data sets and to examine the degree of conflict within the total evidence data set. Azorella and Mulinum were confirmed as not monophyletic. Except three Azorella species, the remaining azorellas, all species of Mulinum, and Laretia form a monophyletic group, designated here as Andean-Patagonian. The three species of Azorella that are not part of the Andean-Patagonian lineage are grouped together with Huanaca and Schizeilema in another lineage, designated here as Austral. Within the Andean-Patagonian clade, three major lineages can be recognized: Diversifolia, Trifurcata, and Spinosum. Each of these lineages have different leaf morpho-anatomies, Diversifolia species being more mesomorphic compared to species of Trifurcata, and species of Spinosum being the most xeromorphic. Hybridizations have been important in the evolution of the group, especially within Diversifolia, with at least six reticulation events resulting in putative homoploid and allopolyploid hybrid species. Evidence from branch lengths and low sequence divergences suggest a rapid radiation in the Spinosum group, probably associated with the acquisition of wings in the fruits.

Phylogeography of Libanotis buchtormensis (Umbelliferae) in Disjunct Populations along the Deserts in Northwest China.

In Northwest China, aridification and desert expansion play significant roles in promoting desert plant diversification and speciation. However, to date, little is known about the effects of the desert barrier on the population structure of montane, non-desert species in the area. In this study, we sequenced chloroplast DNA regions (trnL-trnF and trnS-trnG) and a nuclear gene (rpb2) to investigate the population differentiation and phylogeographical history of Libanotis buchtormensis, a perennial montane species possessing a disjunct distribution at the periphery of the central desert. In total, 23 chloroplast haplotypes and 24 nuclear haplotypes were recovered from the 21 natural populations and six hebarium specimens. Phylogenetic analysis based on the combined plastid and nuclear dataset revealed two distinct lineages of L. buchtormensis, which inhabit the disjunct areas on both sides of the desert zone. The molecular dating analysis indicated that the divergence between the southeastern and the northwestern populations occurred in the middle Pleistocene, concomitantly with the desert expansion. The geographical vicariance likely contributed to the present disjunct distribution of L. buchtormensis across the deserts in Northwest China. Populations in the southeastern region may have migrated from the northwestern region, and seem to be a peripheral distribution of L. buchtormensis.

Monoterpene isolated from the essential oil of Trachyspermum ammi is cytotoxic to multidrug-resistant Pseudomonas aeruginosa and Staphylococcus aureus strains.

INTRODUCTION: The aim of this study was to determine whether an herbal extract containing monoterpene exhibited activity against multidrug-resistant Staphylococcus aureus and Pseudomonas aeruginosa isolated from clinical infection samples. METHODS: The essential oil of Trachyspermum ammi (L.) Sprague ex Turrill (Apiaceae) fruit was extracted by hydrodistillation. Fruit residues were treated with hydrochloric acid and re-hydrodistilled to obtain volatile compounds. Compounds in the distilled oil were identified using gas-chromatography (GC) and GC-mass spectrometry (MS). The antibiotic susceptibility of all bacterial isolates was analyzed using both the disc diffusion method and determination of the minimum inhibitory concentration (MIC). The sensitivity of antibiotic-resistant isolates to essential oil was also determined by using the disc diffusion method and MIC determination. RESULTS: Of 26 clinical isolates, 92% were multidrug-resistant (MDR). Aromatic monoterpenes (thymol, paracymene, and gamma-terpinene) were the major (90%) components of the oil. Growth of S. aureus strains was successfully inhibited by the oil, with an inhibitory zone diameter (IZD) between 30-60mm and MIC <0.02µL/mL. The oil had no antimicrobial activity against clinical isolates of P. aeruginosa; rather, it prevented pigment production in these isolates. CONCLUSIONS: This study revealed that the essential oil of Trachyspermum ammi, which contains monoterpene, has good antibacterial potency. Monoterpenes could thus be incorporated into antimicrobial ointment formulas in order to treat highly drug-resistant S. aureus infections. Our findings also underscore the utility of research on natural products in order to combat bacterial multidrug resistance.