Comprehensive phylogenetic analyses of the Ruppia maritima complex focusing on taxa from the Mediterranean.

Recent molecular phylogenetic studies reported high diversity of Ruppia species in the Mediterranean. Multiple taxa, including apparent endemics, are known from that region, however, they have thus far not been exposed to phylogenetic analyses aimed at studying their relationships to taxa from other parts of the world. Here we present a comprehensive phylogenetic analyses of the R. maritima complex using data sets composed of DNA sequences of the plastid genome, the multi-copy nuclear ITS region, and the low-copy nuclear phyB gene with a primary focus on the Mediterranean representatives of the complex. As a result, a new lineage, "Drepanensis", was identified as the seventh entity of the complex. This lineage is endemic to the Mediterranean. The accessions included in the former "Tetraploid" entity were reclassified into two entities: an Asia-Australia-Europe disjunct "Tetraploid_α" with a paternal "Diploid" origin, and a European "Tetraploid_γ" originating from a maternal "Drepanensis" lineage. Another entity, "Tetraploid_ß", is likely to have been originated as a result of chloroplast capture through backcrossing hybridization between paternal "Tetraploid_α" and maternal "Tetraploid_γ". Additional discovery of multiple tetraploidizations as well as hybridization and chloroplast capture at the tetraploid level indicated that hybridization has been a significant factor in the diversification of Ruppia.

Morphology and phylogenetics of two holoparasitic plants, Balanophora japonica and Balanophora yakushimensis (Balanophoraceae), and their hosts in Taiwan and Japan.

Balanophora japonica and B. yakushimensis are two putatively agamospermic taxa previously reported from southern Japan. Their inflorescences superficially represent those of B. laxiflora and B. fungosa. In this study we confirmed their presence in Taiwan by morphological and phylogenetic analysis using nuclear 18S rDNA and nrITS sequences with related taxa. B. japonica, B. yakushimensis, and B. laxiflora formed a well-supported clade that is distinct from other Balanophora. All three taxa also show considerable differences on morphological and nucleotide sequence differences, therefore the name of B. yakushimensis is retained. The results provide new insights on the intrageneric classification of Balanophora and suggest the positioning of female flowers should be down-weighted. We also successfully identify the hosts of B. japonica and B. yakushimensis by amplifying chloroplast matK sequences from the connected root tissues. The results showed that B. japonica parasitizes on Symplocos species, and that B. yakushimensis parasitizes on Distylium racemosum in Japan and Schima superba in Taiwan's population.

Novel zierane- and guaiane-type sesquiterpenes from the root of Melicope denhamii.

Two novel zierane-type sesquiterpenes, named melicodenones A and B (1 and 2, resp.), and three new guaiane-type sesquiterpenes, named melicodenones C-E (3-5), were isolated from the root of Melicope denhamii (Seem.) T. G. Hartley together with zierone (6). Their structures were established by extensive NMR-spectroscopic analyses. Compounds 1-6 were tested for cytotoxicity using human colon cancer DLD-1 cells, and melicodenone A (1) was found to exhibit moderate activity.

Objective display and discrimination of floral odors from Amorphophallus titanum, bloomed on different dates and at different locations, using an electronic nose.

As olfactory perceptions vary from person to person, it is difficult to describe smells objectively. In contrast, electronic noses also detect smells with their sensors, but in addition describe those using electronic signals. Here we showed a virtual connection method between a human nose perceptions and electronic nose responses with the smell of standard gases. In this method, Amorphophallus titanum flowers, which emit a strong carrion smell, could objectively be described using an electronic nose, in a way resembling the skill of sommeliers. We could describe the flower smell to be close to that of a mixture of methyl mercaptan and propionic acid, by calculation of the dilution index from electronic resistances. In other words, the smell resembled that of "decayed cabbage, garlic and pungent sour" with possible descriptors. Additionally, we compared the smells of flowers which bloomed on different dates and at different locations and showed the similarity of odor intensities visually, in standard gas categories. We anticipate our assay to be a starting point for a perceptive connection between our noses and electronic noses.

Three new constituents from the roots of Erythrina variegata and their antibacterial activity against methicillin-resistant Staphylococcus aureus.

Two new isoflavonoids, eryvarins V and W (1 and 2, resp.), and a new chromen-4-one derivative, eryvarin X (3), along with three known isoflavonoids, 4-6, were isolated from the roots of Erythrina variegata. Their structures were established by spectroscopic analyses. Compound 1 is a rare naturally occurring isoflavanone which possesses a OH group at C(3). Among the new compounds 1-3, 2 exhibited a potent antibacterial activity against methicillin-resistant Staphylococcus aureus (MRSA) strains.

Chemical identity of a rotting animal-like odor emitted from the inflorescence of the titan arum (Amorphophallus titanum).

The titan arum, Amorphophallus titanum, is a flowering plant with the largest inflorescence in the world. The flower emits a unique rotting animal-like odor that attracts insects for pollination. To determine the chemical identity of this characteristic odor, we performed gas chromatography-mass spectrometry-olfactometry analysis of volatiles derived from the inflorescence. The main odorant causing the smell during the flower-opening phase was identified as dimethyl trisulfide, a compound with a sulfury odor that has been found to be emitted from some vegetables, microorganisms, and cancerous wounds.

New phenylpropanoid glycosides from Juniperus communis var. depressa.

Two new phenylpropanoid glycosides were isolated from the leaves and stems of Juniperus communis var. depressa (Cupressaceae) along with 14 known compounds. Their structures were determined by spectral analyses, in particular by 2D-NMR spectral evidence.

Molecular phylogenetic study of the tribe Tropidieae (Orchidaceae, Epidendroideae) with taxonomic and evolutionary implications.

The orchid tribe Tropidieae comprises three genera, Tropidia, Corymborkis and Kalimantanorchis. There are three fully mycoheterotrophic species within Tropidieae: Tropidia saprophytica, T. connata and Kalimantanorchis nagamasui. A previous phylogenetic study of K. nagamasui, based only on plastid matK data, placed K. nagamasui outside the clade of Tropidia and Corymborkis without support. In this study, we performed phylogenetic analyses using a nuclear ribosomal DNA spacer (ITS1-5.8S-ITS2), a low-copy nuclear coding gene (Xdh) and a mitochondrial intron (nad1b-c intron) to study the phylogenetic relationships within Tropidieae. We included six photosynthetic and all three fully mycoheterotrophic Tropidieae species. The resulting phylogenetic trees placed these fully mycoheterotrophic species inside the Tropidia clade with high support. In our trees, these three species do not form a monophyletic group together, because the photosynthetic T. graminea is nested amongst them. Our results also suggest that the loss of photosynthetic ability occurred at least twice in Tropidia.

Endangered island endemic plants have vulnerable genomes.

Loss of genetic diversity is known to decrease the fitness of species and is a critical factor that increases extinction risk. However, there is little evidence for higher vulnerability and extinction risk in endangered species based on genomic differences between endangered and non-endangered species. This is true even in the case of functional loci, which are more likely to relate to the fitness of species than neutral loci. Here, we compared the genome-wide genetic diversity, proportion of duplicated genes (PD), and accumulation of deleterious variations of endangered island endemic (EIE) plants from four genera with those of their non-endangered (NE) widespread congeners. We focused on exhaustive sequences of expressed genes obtained by RNA sequencing. Most EIE species exhibited significantly lower genetic diversity and PD than NE species. Additionally, all endangered species accumulated deleterious variations. Our findings provide new insights into the genomic traits of EIE species.

Evolutionary origin of a periodical mass-flowering plant.

The evolutionary origin of periodical mass-flowering plants (shortly periodical plants), exhibiting periodical mass flowering and death immediately after flowering, has not been demonstrated. Within the genus Strobilanthes (Acanthaceae), which includes more than 50 periodical species, Strobilanthes flexicaulis on Okinawa Island, Japan, flowers gregariously every 6 years. We investigated the life history of S. flexicaulis in other regions and that of closely related species together with their molecular phylogeny to reveal the evolutionary origin of periodical mass flowering. S. flexicaulis on Taiwan Island was found to be a polycarpic perennial with no mass flowering and, in the Yaeyama Islands, Japan, a monocarpic perennial with no mass flowering. Molecular phylogenetic analyses indicated that a polycarpic perennial was the ancestral state in this whole group including S. flexicaulis and the closely related species. No distinctive genetic differentiation was found in S. flexicaulis among all three life histories (polycarpic perennial, monocarpic perennial, and periodical plant). These results suggest that among S. flexicaulis, the periodical mass flowering on Okinawa Island had evolved from the polycarpic perennial on Taiwan Island via the monocarpic perennial in the Yaeyama Islands. Thus, the evolution of life histories could have taken at the level of local populations within a species.

Gene flow and population subdivision in a pantropical plant with sea-drifted seeds Hibiscus tiliaceus and its allied species: evidence from microsatellite analyses.

The genetic differentiation and structure of Hibiscus tiliaceus, a pantropical plant with sea-drifted seeds, and four allied species were studied using six microsatellite markers. A low level of genetic differentiation was observed among H. tiliaceus populations in the Pacific and Indian Ocean regions, similar to the results of a previous chloroplast DNA (cpDNA) study. Frequent gene flow by long-distance seed dispersal is responsible for species integration of H. tiliaceus in the wide distribution range. On the other hand, highly differentiated populations of H. tiliaceus were detected in West Africa, as well as of Hibiscus pernambucensis in southern Brazil. In the former populations, the African continent may be a geographical barrier that prevents gene flow by sea-drifted seeds. In the latter populations, although there are no known land barriers, the bifurcating South Equatorial Current at the north-eastern horn of Brazil can be a potential barrier to gene flow and may promote the genetic differentiation of these populations. Our results also suggest clear species segregation between H. tiliaceus and H. pernambucensis, which confirms the introgression scenario between these two species that was suggested by a previous cpDNA study. Our results also provide good evidence for recent transatlantic long-distance seed dispersal by sea current. Despite the distinct geographical structure observed in the cpDNA haplotypes, a low level of genetic differentiation was found between Pacific and Atlantic populations of H. pernambucensis, which could be caused by transisthmian gene flow.

Loss of the rpl32 gene from the chloroplast genome and subsequent acquisition of a preexisting transit peptide within the nuclear gene in Populus.

Gene transfer events from organelle genomes (mitochondria and chloroplasts in plants) to the nuclear genome are important processes in the evolution of the eukaryotic cell. It is highly likely that the gene transfer event is still an ongoing process in higher plant mitochondria and chloroplasts. The number and order of genes encoded in the chloroplast genome of higher plants are highly conserved. Recently, several exceptional cases of gene loss from the chloroplast genome have been discovered as the number of complete chloroplast genome sequences has increased. The Populus chloroplast genome has lost the rpl32 gene, while the corresponding the chloroplast rpl32 (cp rpl32) gene has been identified in the nuclear genome. Nuclear genes transferred from the chloroplast genome need to gain a sequence that encodes a transit peptide. Here, we revealed that the nuclear cp rpl32 gene has acquired the exon sequence, which is highly homologous to a transit peptide derived from the chloroplast Cu-Zn superoxide dismutase (cp sod-1) gene. The cp rpl32 gene has acquired the sequence that encodes not only for the transit peptide, but also for the conserved N-terminal portion of the mature SOD protein from the cp sod-1 gene, suggesting the occurrence of DNA sequence duplication. Unlike cp SOD-1, cp RPL32 did not show biased localization in the chloroplasts. This difference may be caused by mutations accumulated in the sequence of the SOD domain on the cp rpl32 gene. We provide new insight into the fate of the inherent sequence derived from a transit peptide.

A new flavone xyloside and two new flavan-3-ol glucosides from Juniperus communis var. depressa.

A new flavone xyloside, 1, and two new flavan-3-ol glucosides, 3 and 4, were isolated together with three known flavones, 2, 11, and 12, five known flavans, 5-9, and a known dihydrochalcone, 10, from the stems and leaves of Juniperus communis var. depressa (Cupressaceae) collected in Oregon, U.S.A., and their structures were determined on the basis of spectral evidence. A novel flavone nucleus such as that in 1 is seldom found in nature today, and new methylcatechin glucosides 3 and 4 are also rare in nature. In addition, we investigated the inhibitory activity of individual components, i.e., 8-11, and others, that were abundantly isolated from the same plant material for the Maillard reaction.

Twenty novel polymorphic microsatellite primers in the critically endangered Melastoma tetramerum var. tetramerum (Melastomataceae).

PREMISE OF THE STUDY: Microsatellite markers were identified for Melastoma tetramerum var. tetramerum (Melastomataceae), a critically endangered shrub endemic to the Bonin Islands, to reveal genetic characteristics in wild and restored populations. METHODS AND RESULTS: Using next-generation sequencing, 27 microsatellite markers were identified. Twenty of these markers were polymorphic in M. tetramerum var. tetramerum, with two to nine alleles per locus and expected heterozygosity ranging from 0.10 to 0.71. Among the 20 polymorphic markers, 15 were applicable to other closely related taxa, namely M. tetramerum var. pentapetalum, M. candidum var. candidum, and M. candidum var. alessandrense. CONCLUSIONS: These markers can be potentially useful to investigate the genetic diversity, population genetic structure, and reproductive ecology of M. tetramerum var. tetramerum as well as of the three related taxa to provide appropriate genetic information for conservation.

New resveratrol tetramers from the stem bark of Upuna borneensis.

Five new resveratrol (=5-[(E)-2-(4-hydroxyphenyl)ethenyl]benzene-1,3-diol) tetramers, upunaphenols H-J (1-3) and trans-(4) and cis-upunaphenol K (5), were isolated from the stem of Upuna borneensis (Dipterocarpaceae). Their structures were elucidated on the basis of 1D- and 2D-NMR as well as FAB-MS data. Compounds 1-3 bear a rare biphenyl bond in their frameworks. Compounds 1 and 2 have an unprecedented nonacyclic fused ring system, and compounds 2 and 3 have symmetrical structures.

Two novel trimeric resveratrol derivatives from Cotylelobium lanceolatum.

Two new resveratrol (= 5-[(E)-2-(4-hydroxyphenyl)ethenyl]benzene-1,3-diol) trimers, cotylelophenols A (1) and B (2), were isolated from the stem of Cotylelobium lanceolatum (Dipterocarpaceae), together with ten known resveratrol oligomers (3-12). The structures of the isolates were established on the basis of spectroscopic analyses, including a detailed NMR spectroscopic investigation of 1 under different conditions. Compound 1 is the first resveratrol trimer with a rearranged 4-hydroxyphenyl group. Four possible biogenetic pathways towards resveratrol oligomers are proposed (Scheme).

Acetophenone C-glucosides and stilbene O-glucosides in Upuna borneensis.

Three acetophenone C-glycosides; 2,4,6-trihydroxyacetophenone 3-C-beta-(2'-O-p-hydroxybenzoyl)-glucopyranoside, 2,4,6-trihydroxyacetophenone 3-C-beta-(2'-O-E-coumaroyl)-glucopyranoside, 2,4,6-trihydroxyacetophenone 3-C-beta-(2'-O-E-cinnamoyl)-glucopyranoside, and two resveratrol O-glycosides; piceid 2'-O-p-hydroxybenzoate and, piceid 2'-O-E-ferulate, together with three known compounds were isolated from the acetone soluble part of stem of Upuna borneensis (Dipterocarpaceae). The structures of isolates were determined by spectral analysis including extensive 2D-NMR spectral analyses.

Isoflavonoids from roots of Erythrina zeyheri.

Five isoflavonoids, (+/-)-7,2',4'-trihydroxy-8,3'-di(gamma,gamma-dimethylallyl)isoflavanone, (3R)-7,4'-dihydroxy-2'-methoxy-6,8-di(gamma,gamma-dimethylallyl)isoflavanone, (3R)-7,2',4'-trihydroxy-6,8-di(gamma,gamma-dimethylallyl)isoflavan, 2',4'-dihydroxy-8-gamma,gamma-dimethylallyl-2",2"-dimethylpyrano-[5,6:6,7]isoflavan and (6aS, 11aS)-3,6a-dihydroxy-9-methoxy-4,10-di(gamma,gamma-dimethylallyl)pterocarpan, along with five known compounds, were isolated from the roots of Erythrina zeyheri. Their structures were established on the basis of spectroscopic evidence, and their antibacterial activities against methicillin-resistant Staphylococcus aureus (MRSA) were estimated by determining minimum inhibitory concentrations.

Neolignan and flavonoid glycosides in Juniperus communis var. depressa.

Two neolignan glycosides (junipercomnosides A and B) were isolated from aerial parts of Juniperus communis var. depressa along with two known neolignan glycosides and seven flavonoid glycosides. The structures of the isolated compounds were determined by spectral analysis, in particular by 2D-NMR analysis. The significance of distribution of flavonoids in the chemotaxonomy of genus Juniperus was also discussed.

Stilbenoids from the stem of Gnetum latifolium (Gnetaceae).

An acetone extract of the stem of Gnetum latifolium Blume afforded the stilbene trimer (latifolol) together with five known stilbenoids (gnetin E, gnetin D, gnetin C, (-)epsilon -viniferin and resveratrol). Their structures were elucidated on the basis of spectral evidence, in particular by using 2D NMR methods.