Young-Dong Kim

A DNA barcode for land plants

DNA barcoding involves sequencing a standard region of DNA as a tool for species identification. However, there has been no agreement on which region(s) should be used for barcoding land plants. To provide a community recommendation on a standard plant barcode, we have compared the performance of 7 leading candidate plastid DNA regions (atpF–atpH spacer, matK gene, rbcL gene, rpoB gene, rpoC1 gene, psbK–psbI spacer, and trnH–psbA spacer). Based on assessments of recoverability, sequence quality, and levels of species discrimination, we recommend the 2-locus combination of rbcL+matK as the plant barcode. This core 2-locus barcode will provide a universal framework for the routine use of DNA sequence data to identify specimens and contribute toward the discovery of overlooked species of land plants.

Phylogenetic relationships in family Magnoliaceae inferred from ndhF sequences

The ndhF sequences of 99 taxa, representing all sections in extant Magnoliaceae, were analyzed to address phylogenetic questions in the family. Magnolia macrophylla and M. dealbata, North American species of Magnolia section Rytidospermum, are placed at the base in the subfamily Magnolioideae although its supporting value is low. In the remaining taxa, several distinctive lineages are recognized: (1) Magnolia, the biggest genus in the family, is not monophyletic; (2) Michelia, including section Maingola of Magnolia subgenus Magnolia, is closely related with Elmerrillia and sections Alcimandra and Aromadendron of Magnolia subgenus Magnolia; (3) the associates of Michelia are grouped with Magnolia subgenus Yulania and section Gynopodium of Magnolia subgenus Magnolia; (4) Pachylarnax forms a clade with sections Manglietiastrum and Gynopodium of Magnolia; (5) a well-supported Manglietia clade is recognized; (6) Caribbean species of section Theorhodon of Magnolia subgenus Magnolia, which are section Splendentes sensu Vázquez-Garcia, are closely allied with New World members of Magnolia subgenus Talauma; and (7) section Rytidospermum of Magnolia subgenus Magnolia and subgenus Talauma are polyphyletic. The separated clades in the molecular tree are considerably different from traditional taxonomic dispositions in the family. The molecular data strongly suggest that a taxonomic realignment of infrafamilial delimitations and compositions should be considered.

Complete chloroplast and ribosomal sequences for 30 accessions elucidate evolution of Oryza AA genome species.

Cytoplasmic chloroplast (cp) genomes and nuclear ribosomal DNA (nR) are the primary sequences used to understand plant diversity and evolution. We introduce a high-throughput method to simultaneously obtain complete cp and nR sequences using Illumina platform whole-genome sequence. We applied the method to 30 rice specimens belonging to nine Oryza species. Concurrent phylogenomic analysis using cp and nR of several of specimens of the same Oryza AA genome species provides insight into the evolution and domestication of cultivated rice, clarifying three ambiguous but important issues in the evolution of wild Oryza species. First, cp-based trees clearly classify each lineage but can be biased by inter-subspecies cross-hybridization events during speciation. Second, O. glumaepatula, a South American wild rice, includes two cytoplasm types, one of which is derived from a recent interspecies hybridization with O. longistminata. Third, the Australian O. rufipogan-type rice is a perennial form of O. meridionalis.

Chloroplast DNA restriction site variation and phylogeny of the Berberidaceae

Comparative restriction site mapping of the chloroplast genome was performed to examine phylogenetic relationships among 27 species representing 16 genera of the Berberidaceae and two outgroups. Chloroplast genomes of the species included in this study showed no major structural rearrangements (i.e., they are collinear to tobacco cpDNA) except for the extension of the inverted repeat in species of Berberis and Mahonia. Excluding several regions that exhibited severe length variation, a total of 501 phylogenetically informative sites was mapped for ten restriction enzymes. The strict consensus tree of 14 equally parsimonious trees indicated that some berberidaceous genera (Berberis, Mahonia, Diphylleia) are not monophyletic. To explore phylogenetic utility of different parsimony methods phylogenetic trees were generated using Wagner, Dollo, and weighted parsimony for a reduced data set that included 18 species. One of the most significant results was the recognition of the four chromosomal groups, which were strongly supported regardless of the parsimony method used. The most notable difference among the trees produced by the three parsimony methods was the relationships among the four chromosomal groups. The cpDNA trees also strongly supported a close relationship of several generic pairs (e.g., Berberis-Mahonia, Epimedium-Vancouveria, etc.). Maximum likelihood values were computed for the four different tree topologies of the chromosomal groups, two Wagner, one Dollo, and one weighted topology. The results indicate that the weighted tree has the highest likelihood value. The lowest likelihood value was obtained for the Dollo tree, which had the highest bootstrap and decay values. Separate analyses using only the Inverted Repeat (IR) region resulted in a tree that is identical to the weighted tree. Poor resolution and/or support for the relationships among the four chromosomal lineages of the Berberidaceae indicate that they may have radiated from an ancestral stock in a relatively short evolutionary time.

Phylogeny of Weigela and Diervilla (Caprifoliaceae) Based on Nuclear rDNA ITS Sequences: Biogeographic and Taxonomic Implications

Weigela and three species of Diervilla were inferred from nucleotide sequence variation in the internal transcribed spacer (ITS) regions of 18-26S nuclear ribosomal DNA. Phylogenetic trees were obtained using parsimony, distance, and maximum likelihood methods. The ITS phylogenies did not support the monophyly of Weigela, which comprises three major clades. The first clade includes a single species, W. middendorffiana, which is more closely related with American genus, Diervilla. The second clade consisted of a single species, W. maximowiczii, but its relationship to other Weigela species is equivocal. The third one is a strongly supported core clade which contains the remaining Weigela species. Within the core group, W. hortensis exhibits the most distinct ITS sequence type. The hybrid origin of the species from the putative parents, W. florida and W. subsessilis, is not supported. The ITS sequence evidence favors the taxonomic hypothesis proposed by Nakai who recognized four distinct evolutionary lines (Diervilla, Macrodiervilla, Weigelastrum and Weigela). However, recognition of the two sections, Calysphyrum and Weigela was not supported in the ITS phylogenies. Substantial morphological differences between Diervilla and Weigela middendorffiana do not support the morphological stasis concept which has been considered as a general evolutionary mode among the Asian and North American disjunct taxa.

Phylogenetic Implications of rbcL and ITS Sequence Variation in the Berberidaceae

In spite of the use of chromosome number, floral anatomy, pollen morphology, and serology, substantial disagreement remains regarding the systematics of the Berberidaceae. To test competing hypoth- eses about intergeneric relationships in the family, sequences of the chloroplast encoded rbcL gene and ITS regions of the nuclear ribosomal repeat were examined. Analysis of rbcL sequences does not support fragmentation of the Berberidaceae into smaller families, especially the separation of Nandina as a distinct family. The large woody genera, Berberis and Mahonia, are related to the monotypic herbaceous genus Ranzania. Bongardia is nested within a clade characterized by a basic chromosome number of x = 6, and has a remote relationship to the Leontice group with which the genus has been placed in most previous classifica- tions. The x = 6 group, the largest in the Berberidaceae, comprises two distinct lineages: Jeffersonia and the rest of the core genera (Diphylleia, Bongardia, Achlys, and Epimedium). Among the core genera, Diphylleia is sister to Epimedium/Achlys and their sister genus Bongardia. ITS sequence data provided additional support for the phylogenetic relationships of the x = 6 group observed in the rbcL gene tree. Berberidaceae (s.l.) is a heterogeneous assem-

The origin and relationships of the pepino,Solanum muricatum (solanaceae): DNA restriction fragment evidence

The pepino (or pepino dulce:Solanum muricatum) is a domesticate, of interest because of its close relationship to tomatoes and potatoes, because it is enjoying increasing exposure in the international market, and because it is a cultigen with no known wild ancestor. Morphologically this South American native is a member of the Solanum sect. Basarthrum, and as such, is allied to a number of Andean wild species. Data from other studies are combined with results from restriction site analysis of chloroplast and nuclear ribosomal DNA to assay relationships and the potential origin of the pepino. The pepino may have existed in the wild previously and may be represented today only by the cultigen. However, if its ancestors are extant, three wild species—Solanum basendopogon (Perú),S. caripense (Costa Rica through Perú), S. tabanoense (Colombia and Ecuador)—emerge as most likely progenitors. Phylogenetic analyses of 61 accessions, including 27 of the pepino, dependent on chloroplast DNA (cpDNA) and nuclear ribosomal (rDNA) restriction site data show the pepino to be polymorphic, suggest independent origins for some of the cultivars, and most strongly supportS. tabanoense as the progenitor of the cultigen.Solanum caripense also may have been a direct ancestor of the pepino, or may have hybridized subsequent to its origin with the pepino to yield some of the haplotype variation. Similarly, S.cochoae may have hybridized with the pepino. There are no DNA characters supporting the involvement ofS. basendopogon in the origin.ResumenAnálisis de Sitios de Restricción en el Adn.El ‘pepino’ (o ‘pepino dulce’:Solanum muricatum) es una planta domesticada de gran interés por estar estrechamente emparentada con los tomates y papas, porque su dispersión en el mercado internacional está incrementando y porque es un cultigen con ancestro silvestre desconocido. Morfológicamente esta planta natural de Sudamérica es un meimbro del géneroSolanum sect.Basarthrum, y como tal, está ligada a varias especies silvestres de la región andina. Datos provenientes de estudios previos son combinados aquí con los resultados del análisis de sitios de restricción en el ADN ribosómico nuclear y plastidial para investigar el parentesco y el origen potencial del pepino. Este último pudo haber existido previamente en estado silvestre y hoy en día estar probablemente representado sólo por el cultigen. Sin embargo, si sus ancestros están extintos, tres expecies silvestres—Solanum basendopogon (Perú),S. caripense (de Costa Rica hasta Perú),S. tabanoense (Colombia y Ecuador) –aparecen como los progenitores más probables. Análisis filogenéticos de 61 ancestros incluyendo 27 del pepino, localizados en los sitios de restricción del ADN plastidial (cpDNA) y ADN ribosómico (rDNA), muestran que el pepino es polimórfico, sugiriendo al mismo tiempo orígenes independientes para algunos de los cultivares, y sustentan fuertemente aS. tabanoense como progenitor del cultigen. Solanum caripense también pudo haber sido ancestro directo del pepino, o podría haberse hibridizado después de su origen con algunas de las accesiones del pepino para eliminar así algo de la variación haplotípica. De la misma manera,S. cochoae pudo haberse hibridizado con el pepino. No existen caracteres del ADN que sugieran la implicación de S. basendopogon en el origen.

Myrsine seguinii ethanolic extract and its active component quercetin inhibit macrophage activation and peritonitis induced by LPS by targeting to Syk/Src/IRAK-1.

ETHNOPHARMACOLOGICAL RELEVANCE Myrsine seguinii H. LÉVEILLÉ (syn. Rapanea neriifolia) (Myrsinaceae) is a medicinal plants traditionally used in Myanmar to treat infectious and inflammatory diseases. Since none of reports have systematically demonstrated the anti-inflammatory activity of this plant, we aimed to mechanistically understand the regulatory roles of the plant in inflammatory responses using the ethanolic extract of Myrsine seguinii (Ms-EE). MATERIALS AND METHODS Activated macrophages and peritonitis symptoms induced by lipopolysaccharide (LPS) were employed. HPLC analysis was used to identify active components. To characterize direct target enzymes, kinase assay was established. RESULTS Ms-EE inhibited the production of nitric oxide (NO) and prostaglandin (PG)E2 in RAW264.7 cells and peritoneal macrophages stimulated by LPS. This extract suppressed the mRNA expression of the inducible NO synthase (iNOS) and cyclooxygenase (COX)-2 genes by down-regulating the activation of nuclear factor (NF)-κB and activator protein (AP-1). Interestingly, it was found that Ms-EE can directly suppress the enzyme activities of Syk, Src, and interleukin-1 receptor-associated kinase-1 (IRAK-1). Similarly, orally administered Ms-EE inhibited the phosphorylation of Src and Syk in peritoneal exudate-derived cells prepared from peritonitis. Finally, HPLC analysis clearly demonstrated that quercetin is a major active component with suppressing activity on the release of inflammatory mediators (NO and PGE2), and the enzyme activities of Src, Syk, and IRAK-1. CONCLUSION Ms-EE containing quercetin negatively modulates macrophage-mediated in vitro inflammatory responses and LPS-induced peritonitis by blocking the Src/Syk/NF-κB and IRAK-1/AP-1 pathways, which contributes to its major ethnopharmacological use as an anti-inflammatory herbal medicine.

Quantitative ethnobotanical study of the medicinal plants used by the Ati Negrito indigenous group in Guimaras island, Philippines

ETHNOPHARMACOLOGICAL RELEVANCE This study is (1) a documentation of medicinal plant use in traditional therapies, and (2) an evaluation of the medicinal plant knowledge and practices of the Ati Negrito indigenous people in Guimaras Island, Philippines. MATERIALS AND METHODS A semi-structured interview was conducted to 65 informants in order to determine the medicinal plants and their uses in traditional therapies. The plants were collected, identified and deposited as voucher specimens. Plant importance was determined using quantitative ethnobotanical indices such as Use Value (UV), Fidelity Level (FL) and Informant Consensus Factor (ICF). Descriptive and the inferential statistics Mann-Whitney U and Kruskal-Wallis tests were used to measure and compare the informants׳ medicinal plant use knowledge and practices. RESULTS This study was able to identify 142 medicinal plant taxa in 55 families used in 16 categories of diseases. Plants with the highest recorded UVs were Psidium guajava (2.52), Blumea balsamifera (2.15) and Cocos nucifera (2.06). A total of 24 species were found to have 100% FL values with Senna alata, Hibiscus rosa-sinensis and Breynia vitis-idaea recording the highest number of use-mentions (48) for treating white spot, boils, and child sleeplessness, malaise and fatigue, respectively. The highest ICF value (1.00) was cited for Category 6 (Diseases of the ear) and Category 16 (Factors that influence health status and services). Finally, significant differences in medicinal plant use knowledge were recorded when informants were grouped according to location, educational level, gender and age. CONCLUSIONS This documentation of medicinal plants and their uses shows the rich tradition in ethno-medicinal knowledge of the Ati Negrito indigenous people although results might also imply that knowledge is eroding. Nevertheless, this study could open an avenue for pharmacological research works, or serve as reference for future quantitative ethnobotanical investigations.

Characterization and phylogenetic distribution of a chloroplast DNA rearrangement in theBerberidaceae

Restriction site maps and a clone bank of chloroplast DNA (cpDNA) ofMahonia higginsae (Munz)Ahrendt (Berberidaceae) were constructed. The size ofMahonia cpDNA was about 167 kb. Precise mapping using gene probes revealed that cpDNA ofM. higginsae has an inverted repeat (IR) 11.5 kb larger than the tobacco IR. The expansion of the IR into the large single copy region has resulted in the duplication of at least ten genes includingpsbB. The phylogenetic distribution of the expanded IR was examined in twenty-five species ofBerberis andMahonia, twenty species representing the fifteen remaining genera of theBerberidaceae, and four species from four allied families. Our survey indicates that only the species of the closely related generaBerberis andMahonia share the 11.5kb expansion of IR. This result supports their close phylogenetic relationship, which has been suggested previously by chromosomal, morphological, and serological data.