Inferring complex evolutionary history of the closely related East Asian wild roses in Rosa sect. Synstylae (Rosaceae) based on genomic evidence from conserved orthologs.

BACKGROUND AND AIMS: The section Synstylae in genus Rosa (Rosaceae) comprises 25-36 species and includes several major progenitors of modern rose cultivars. East Asian Synstylae species have recently diverged and are closely related, and their phylogenetic relationships remain unclear. In the present study, we employed a conserved ortholog set (COS) markers and genome-wide nuclear orthologs to elucidate their phylogenetic relationships and unravel their complex evolutionary history. METHODS: Utilising on eight Rosaceae COS (RosCOS) markers, we analysed a total of 137 accessions representing 15 East Asian Synstylae taxa to establish a robust phylogenetic framework and reconstruct ancestral areas. Furthermore, we constructed the species tree for eight representative species and estimated their divergence times based on 1,683 genome-wide orthologs. The species tree-gene tree coalescence time comparison, Patterson's D, f4-ratio, and f-branch statistics were analysed to identify incomplete lineage sorting (ILS), genetic introgression, and reticulation events using conserved ortholog data. KEY RESULTS: RosCOS markers and genome-wide orthologs effectively resolved the robust phylogeny of East Asian Rosa sect. Synstylae. Species divergence times estimated with genome-wide orthologs indicated that East Asian Synstylae species have recently diverged, with an estimated crown age of approximately 2 Mya. The rampant gene tree discordance indicated the possibility of ILS and/or genetic introgression. In the section Synstylae, deeper coalescence in the gene trees compared to the species tree suggested ILS as a source of gene tree discordance. Further, Patterson's D and f-branch statistics indicated that several lineages in the section were involved in genetic introgression. CONCLUSIONS: We have unravelled the complex evolutionary history of East Asian Rosa sect. Synstylae, including recent species divergences, ILS, and genetic introgression. Coupled with the geographical and ecological complexity of East Asia, ILS and genetic introgression may have contributed to the rapid diversification of East Asian Synstylae species by permitting adaptation to diverse environments.

Comparative chemical profiling of leaf essential oils from Cinnamomum kanehirae and related species using steam distillation and solvent extraction: Implications for plant-based classification.

Cinnamomum kanehirae Hayata, belonging to Lauraceae family, is an indigenous and endangered species of considerable economic importance in Taiwan. It plays a crucial role as the host for the economically valuable saprotrophic fungus, Taiwanofungus camphorates. However, accurate species identification poses a challenge due to the similarity in morphological features and frequent natural hybridization with closely related species. Acquiring high-quality and pure leaf oils becomes imperative for precise species identification and producing superior goods. In this study, our objective was to establish methodologies for analyzing the chemical composition of leaf essential oils and subsequently apply this knowledge to differentiate among three Cinnamomum species. Gas chromatography-mass spectrometry (GC/MS) was employed to scrutinize the chemical makeup of leaf essential oils from three closely related species: C. kanehirae, C. micranthum, and C. camphora. We utilized Steam Distillation (SD) and steam distillation-solvent extraction (SDSE) methods, with the SDSE-Hexane approach chosen for optimization, enhancing extraction efficiency and ensuring essential oil purity. Through the SDSE-Hexane method, we identified seventy-four compounds distributed across three major classes: monoterpenes hydrocarbons (0.0-7.0 %), oxygenated monoterpenes (3.8-90.9 %), sesquiterpenes hydrocarbons (0.0-28.3 %), and oxygenated sesquiterpenes (1.6-88.1 %). Our findings indicated the presence of more than one chemotype in both C. kanehirae and C. camphora, whereas no specific chemotype could be discerned in C. micranthum. Furthermore, clustering based on chemotypes allowed for the differentiation of samples from the three species. Notably, we demonstrated that the chemical compositions of grafted C. kanehirae remained largely unaffected by the rootstock. Conversely, natural hybrids between C. kanehirae and C. camphora exhibited profiles more closely aligned with C. kanehirae. The optimized extraction method and the chemotype-based classification system established in this study present valuable tools for essential oil preparation, species identification, and further exploration into the genetic variation of Cinnamomum.

Genetic diversities in wild and cultivated populations of the two closely-related medical plants species, Tripterygium Wilfordii and T. Hypoglaucum (Celastraceae).

BACKGROUND: The sustainable supply of medicinal plants is important, and cultivating and domesticating them has been suggested as an optimal strategy. However, this can lead to a loss of genetic diversity. Tripterygium wilfordii Hook. f. is a medicinal plant commonly used in traditional Chinese medicine, but its wild populations are dwindling due to excessive harvesting. To protect the species and meet the increasing demand, it is urgent to cultivate it on a large scale. However, distinguishing between T. wilfordii and T. hypoglaucum, two similar species with different medicinal properties, is challenging. Therefore, it is crucial to understand the genetic diversity and population structure of these species for their sustainable utilization. RESULTS: In this study, we investigated the genetic diversity and population structure of the two traditional medicinal semiwoody vines plant species, Tripterygium wilfordii and T. hypoglaucum, including wild and cultivated populations using chloroplast DNA (cpDNA) sequences and microsatellite loci. Our results indicated that the two species maintain a high level of genetic divergence, indicating possible genetic bases for the different contents of bioactive compounds of the two species. T. wilfordii showed lower genetic diversity and less subdivided population structures of both markers than T. hypoglaucum. The potential factors in shaping these interesting differences might be differentiated pollen-to-seed migration rates, interbreeding, and history of population divergence. Analyses of cpDNA and microsatellite loci supported that the two species are genetically distinct entities. In addition, a significant reduction of genetic diversity was observed for cultivated populations of the two species, which mainly resulted from the small initial population size and propagated vegetative practice during their cultivation. CONCLUSION: Our findings indicate significant genetic divergence between T. wilfordii and T. hypoglaucum. The genetic diversity and population structure analyses provide important insights into the sustainable cultivation and utilization of these medicinal plants. Accurate identification and conservation efforts are necessary for both species to ensure the safety and effectiveness of crude drug use. Our study also highlighted the importance of combined analyses of different DNA markers in addressing population genetics of medicinal plants because of the contrasts of inheritance and rates of gene flow. Large-scale cultivation programs should consider preserving genetic diversity to enhance the long-term sustainability of T. wilfordii and T. hypoglaucum. Our study proposed that some populations showed higher genetic diversity and distinctness, which can be considered with priority for conservation and as the sources for future breeding and genetic improvement.

Unveiling Cryptic Species Diversity and Genetic Variation of Lasiodiplodia (Botryosphaeriaceae, Botryosphaeriales) Infecting Fruit Crops in Taiwan.

The genus Lasiodiplodia, a member of the family Botryosphaeriaceae, is an important fungal disease genus in agriculture. However, the Lasiodiplodia species survey and genetic diversity in Taiwan remain unclear. This study aimed to investigate the Lasiodiplodia species associated with various fruit species to explore the cryptic Lasiodiplodia species diversity, validate species delimitation, and unveil cryptic genetic diversity. Overall, six Lasiodiplodia species were identified, with several new records of infection identified. Additionally, phylogenetic analyses indicated that the relations of all isolates of L. theobromae might be paraphyletic. They were grouped with L. brasiliense based on Automatic Barcode Gap Discovery (ABGD), Automatic Partitioning (ASAP) and structure-based clustering analyses. These analyses did not provide conclusive evidence for L. brasiliensis as a stable species. It may be necessary to gather more information to clarify the species delineation. The multiple new records of Lasiodiplodia species with high genetic diversity and differentiation revealed that the diversity of Lasiodiplodia in Taiwan was underestimated in the past. We found that L. theobromae has the highest number of haplotypes but the lowest number of haplotype and nucleotide diversities, indicating a recent population expansion. This was supported by the significant negative Tajima's D and Fu and Li's D* tests. The high genetic diversity, low gene flow, and host-associated differentiation of Lasiodiplodia species indicate that they might harbour powerful evolutionary potential in Taiwan. This study provided critical insights into genetic variation, host-associated differentiation, and demography of Lasiodiplodia species, which would be helpful for disease management of related pathogens.

Deep structured populations of geographically isolated nipa (Nypa fruticans Wurmb.) in the Indo-West Pacific revealed using microsatellite markers.

Nipa (Nypa fruticans Wurmb.) is an important mangrove palm species, but it is understudied due to lack of information on genetic patterns within its distribution range. In this study, we identified 18 informative microsatellite markers to assess genetic variations among local populations in the Indo-West Pacific (IWP). Results showed population stratification based on high genetic differentiation (FST = 0.22131) with the Mantel test indicating significance to isolation-by-distance. We found a pronounced differentiation between the west populations in Sri Lanka and east populations in Southeast Asia. The east populations around the South China Sea were more genetically similar than those along the Malacca Strait and Java Sea. These genetic clines were shaped by ocean circulations and seasonal monsoon reversals as plausible factors. The Malacca Strait was confirmed as both a genetic and a geographic barrier rather than a corridor according to the Monmonier plot. Simulations of directional migration indicated a statistically strong contemporary genetic connectivity from west to east where Sri Lankan immigrants were detected as far as central Philippines via long-distance dispersal. This is the first report on the recent migration patterns of nipa using microsatellites. Assignment of first-generation (F0) immigrants suggested Mainland Southeast Asia as a melting pot due to the admixture associated with excess of homozygosity. The western populations were recent expansions that emerged in rapid succession based on a phylogram as supported by footprints of genetic drift based on bottleneck tests.

Structured Populations of Critically Endangered Yellow Water Lily (Nuphar shimadai Hayata, Nymphaeaceae).

Yellow water lily (Nuphar shimadai Hayata) is a critically endangered species in Taiwan. Here, we examined genetic structures of four extant populations, WP, GPa, GPb and GPn, using 39 simple sequence repeat (SSR) markers. Positive genetic correlation was observed within 50 m, beyond which no correlation was detected due to isolation by distance according to Mantel correlogram. This suggests a significant genetic structuring of the species. Besides, multilocus genotype (MLG) analysis revealed that GPa was a panmictic population and the species' putative center of origin. Genetic exchange was observed between GPa and GPb populations, which likely resulted from their geographic proximity. Nevertheless, there was a strong asymmetric migration detected from GPa to WP, but a recent genetic barrier prevented dispersal further northward (WP). Geneland estimated the best number of clusters as K = 2, where WP distinctly separated from the rest of the populations. In STRUCTURE output of K = 3, a third cluster was abundant only in WP. We suggest to consider GPn and WP as separate conservation units, being far from GPa. There is indeed a need to investigate these populations; as predicted, Ne = 1.6 to 3.0 is considered low and that may put the species at risk of extinction.

Hinokitiol impedes tumor drug resistance by suppressing protein kinase B/mammalian targets of rapamycin axis.

Chemotherapy is a treatment method commonly used for cancer and that patients showing low to no response to the treatment often developed drug resistance via multiple mechanisms. Natural products have been shown to reduce tumor drug resistance. Hinokitiol, a natural tropolone derivative, has potential as an antitumor agent. To improve the efficacy and safety of hinokitiol, a further understanding of hinokitiol interactions with the tumor microenvironment is necessary. The presence of plasma membrane multidrug resistance protein P-glycoprotein (P-gp) is favorable for tumor cells to elicit chemotherapeutic resistance. Here, we showed that hinokitiol dose-dependently decreased P-gp expression and suppressed the P-gp-driven efflux activity based on Rhodamine 123 assay. The protein expression levels of phosph-protein kinase B (P-AKT), phosph-mammalian targets of rapamycin (P-mTOR), and phosph-p70 ribosomal s6 kinase (P-p70s6K) in tumor cells were likewise reduced after hinokitiol treatment. The transfection of cells with active P-AKT rescued hinokitiol-induced downregulation of P-gp, suggesting the involvement of Akt/mTOR/p70s6K signaling in P-gp expression. Our results showed that hinokitiol can chemosensitize cancer cells. These findings indicate that hinokitiol could enhance 5-Fluorouracil therapeutic effects in murine B16F10 and CT26 tumor cells via downregulation of the AKT/mTOR pathway.

Biopiracy: Abolish Corporate Hijacking of Indigenous Medicinal Entities.

Biopiracy as "a silent disease" is hardly detectable because it does not leave traces frequently. The corporate hijacking of food is the most important health hazard in this era; giant commercial enterprises are using intellectual property rights to patent indigenous medicinal plants, seeds, genetic resources, and traditional medicines. The new era of biotechnology relies on the genes of living organisms as raw materials. The "Gene Rush" has thus become similar to that of the old "Gold Rush." Sri Lanka has been spotted in the top 34 biodiversity hotspots globally. Moreover, localized in the tropics, human generations in Sri Lanka have utilized the array of plant species for herbal treatments and treatment of diseases. Sri Lanka after its 30-year civil war is moving towards a solid growth and conservation of the environment which is a major component in a sustainable development where the conservation of biodiversity plays a significant role. In this paper, we present an overview of typical cases of global biopiracy, bioprospecting via introduction of cost-effective deoxyribonucleic acid (DNA) fingerprinting and international protocol with Private-Public-People Partnership concept as excellent forms of utilization of natural resources. We propose certain perspectives as scientists towards abolishing biopiracy and also to foster the fair utilization of natural resources; since the economy of most developing countries is agriculture based, the gross domestic product of the developing countries could be increased by enhanced bioprospecting via introduction of cost-effective DNA fingerprinting technologies and thus not being a pray of corporate hijacking."Biopiracy is biological theft; illegal collection of indigenous plants by corporations who patent them for their own use" (Vandana Shiva).

Correlation Between Pathogenic Determinants Associated with Clinically Isolated Non-Typhoidal Salmonella.

Non-typhoidal and Typhoidal Salmonella are bacterial pathogens source of worldwide and major disease burden. Virulent determinants of specific serovars belonging to non-typhoidal Salmonella have been extensively studied in different models, yet the pathogenesis of this group of bacteria and the development of clinical symptoms globally remains underexplored. Herein, we implemented microbiological and molecular procedures to investigate isolate virulence traits and molecular diversity, likely in association with disease severity. Our results show that selected clinical isolates from a tertiary referring hospital, depending on the richness of the environment and isolate serotypes, exhibited different, and sometimes controversial, virulence properties. The tested strains were susceptible to Ceftriaxone (90%) with decreasing reactivity to Trimethoprim-Sulfamethoxazole (72%), Chloramphenicol (64%), Ampicillin (48%), Gentamicin (44%), and Ciprofloxacin (2%). Disc susceptibility results partially correlated with minimum inhibitory concentration (MIC); however, special attention must be given to antimicrobial treatment, as a rise in multi-resistant isolates to Trimethoprim-Sulfamethoxazole (2/38 µg/mL), Minocycline (8 µg/mL) and Ampicillin (16 µg/mL) has been noticed, with two isolates resistant to Ceftazidime (16 µg/mL). By comparison to previous molecular epidemiology studies, the variation in the gene profiles of endemic pathogens supports the need for continuous and up-to-date microbiological and molecular reports.

Characterization and comparative analysis among plastome sequences of eight endemic Rubus (Rosaceae) species in Taiwan.

Genus Rubus represents the second largest genus of the family Rosaceae in Taiwan, with 41 currently recognized species across three subgenera (Chamaebatus, Idaoeobatus, and Malochobatus). Despite previous morphological and cytological studies, little is known regarding the overall phylogenetic relationships among the Rubus species in Taiwan, and their relationships to congeneric species in continental China. We characterized eight complete plastomes of Taiwan endemic Rubus species: subg. Idaeobatus (R. glandulosopunctatus, R. incanus, R. parviaraliifolius, R rubroangustifolius, R. taitoensis, and R. taiwanicolus) and subg. Malachobatus (R. kawakamii and R. laciniastostipulatus) to determine their phylogenetic relationships. The plastomes were highly conserved and the size of the complete plastome sequences ranged from 155,566 to 156,236 bp. The overall GC content ranged from 37.0 to 37.3%. The frequency of codon usage showed similar patterns among species, and 29 of the 73 common protein-coding genes were positively selected. The comparative phylogenomic analysis identified four highly variable intergenic regions (rps16/trnQ, petA/psbJ, rpl32/trnL-UAG, and trnT-UGU/trnL-UAA). Phylogenetic analysis of 31 representative complete plastomes within the family Rosaceae revealed three major lineages within Rubus in Taiwan. However, overall phylogenetic relationships among endemic species require broader taxon sampling to gain new insights into infrageneric relationships and their plastome evolution.

Ecological Factors Generally Not Altitude Related Played Main Roles in Driving Potential Adaptive Evolution at Elevational Range Margin Populations of Taiwan Incense Cedar (Calocedrus formosana).

Population diversification can be shaped by a combination of environmental factors as well as geographic isolation interacting with gene flow. We surveyed genetic variation of 243 samples from 12 populations of Calocedrus formosana using amplified fragment length polymorphism (AFLP) and scored a total of 437 AFLP fragments using 11 selective amplification primer pairs. The AFLP variation was used to assess the role of gene flow on the pattern of genetic diversity and to test environments in driving population adaptive evolution. This study found the relatively lower level of genetic diversity and the higher level of population differentiation in C. formosana compared with those estimated in previous studies of conifers including Cunninghamia konishii, Keteleeria davidiana var. formosana, and Taiwania cryptomerioides occurring in Taiwan. BAYESCAN detected 26 F ST outlier loci that were found to be associated strongly with various environmental variables using multiple univariate logistic regression, latent factor mixed model, and Bayesian logistic regression. We found several environmentally dependent adaptive loci with high frequencies in low- or high-elevation populations, suggesting their involvement in local adaptation. Ecological factors, including relative humidity and sunshine hours, that are generally not altitude related could have been the most important selective drivers for population divergent evolution in C. formosana. The present study provides fundamental information in relation to adaptive evolution and can be useful for assisted migration program of C. formosana in the future conservation of this species.

The genetic relationships of Indian jujube (Ziziphus mauritiana Lam.) cultivars using SSR markers.

The genetic relationships among 24 Indian jujube cultivars (Ziziphus mauritiana Lam.) were evaluated by genotyping the microsatellite loci using simple sequence repeat (SSR) markers. The SSR loci were scored by fluorescent labelling and automated detection systems for the high-throughput capillary electrophoresis and high-resolution gel electrophoresis. Out of the 29 newly characterized SSR loci, 26 were considered as polymorphic with a total of 181 alleles obtained. The number of alleles ranged from 2-12, while the polymorphism information content ranged from 0.08-0.83, and the expected and observed heterozygosity were 0.04-0.83 and 0.04-0.82, respectively. The allele pattern of Indian jujube for all SSR loci confirmed its karyotype as tetraploid. Similarity coefficients and UPGMA dendrogram revealed that the Taiwanese cultivars consisted of a large 'A' clade, which is further divided into 'A1' and 'A2' groups, and the 'B' clade where both are rooted by the wild accession, 'Chad native'. These four genetic clusters were supported by the results of PCoA and the assignment test. The excess of heterozygotes based on F-statistics was attributed to its mating system as outcrossing and self-incompatible, and the introgression of the presumed mutation-derived cultivars with genetic admixture. Based on this study, SSR markers offer valuable information on the genetic relationship of this tropical fruit tree which is basically in agreement with the genealogy of its breeding history.

Cloning and promoter analysis of palladin 90-kDa, 140-kDa, and 200-kDa isoforms involved in skeletal muscle cell maturation.

OBJECTIVE: Palladin is a ubiquitous phosphoprotein expressed in vertebrate cells that works as a scaffolding protein. Several isoforms deriving from alternative splicing are originated from the palladin gene and involved in mesenchymal and muscle cells formation, maturation, migration, and contraction. Recent studies have linked palladin to the invasive spread of cancer and myogenesis. However, since its discovery, the promoter region of the palladin gene has never been studied. The objective of this study was to predict, identify, and measure the activity of the promoter regions of palladin gene. RESULTS: By using promoter prediction programs, we successfully identified the transcription start sites for the Palld isoforms and revealed the presence of a variety of transcriptional regulatory elements including TATA box, GATA, MyoD, myogenin, MEF, Nkx2-5, and Tcf3 upstream promoter regions. The transcriptome profiling approach confirmed the active role of predicted transcription factors in the mouse genome. This study complements the missing piece in the characterization of palladin gene and certainly contributes to understanding the complexity and enrollment of palladin regulatory factors in gene transcription.

Genetic Diversity and Phylogeography of the Important Medical Herb, Cultivated Huang-Lian Populations, and the Wild Relatives Coptis Species in China.

Huang-lian (Coptis plants in China) are essential medicinal plants in China, C. chinensis var. chinensis and C. deltoidea have been domesticated and cultivated for 700 years. In this study, the genetic diversity patterns and biogeographical information of cultivated Huang-lian and their wild relatives Coptis species were assessed using three plastids DNA regions. A total of 186 individuals from twenty-seven populations representing two species of cultivated Huang-lian and four species of wild relatives were collected and analyzed. Twenty-four haplotypes of six species were identified when three plastid spacers were combined. Historical biogeography inference revealed multiple dispersal events in the groups of cultivated Huang-lian and C. omeiensis. This evidence can infer that large initial population size and interbreeding with co-existing wild relatives in expanding new planting areas might be the main reason for maintaining the high genetic diversity of cultivated Huang-lian. Nevertheless, the multimodal curve of mismatch analysis and positive or negative differed among species and populations by neutrality tests indicated some groups of cultivated Huang-lian experienced genetic bottlenecks. Phylogeny analysis (NJ, MP, BI) showed that cultivated Huang-lian and C. omeiensis were clustered into a monophyletic group while C. chinensis var. brevisepala was paraphyletic, having earlier divergence time from C. chinensis var. chinensis (7.6 Ma) than C. omeiensis. Parsimony network demonstrated that C. deltoidea had more shared haplotypes with C. omeiensis than C. chinensis var. chinensis, and other haplotypes of C. deltoidea and C. omeiensis had less mutation steps than that of C. chinensis var. chinensis and C. omeiensis. This evidence suggests that C. omeiensis has a closer relationship with cultivated Huang-lian and might be a potential wild relative to C. deltoidea. The results reported here provide the baseline data for preserving genetic resources of Huang-lian and also evaluating the genetic impacts of long-term cultivation on medicinal plants, which could be instructive to future cultivation projects of traditional Chinese medicinal plants.

Phylogeny and Historical Biogeography of Paphiopedilum Pfitzer (Orchidaceae) Based on Nuclear and Plastid DNA.

The phylogeny and biogeography of the genus Paphiopedilum were evaluated by using phylogenetic trees derived from analysis of nuclear ribosomal internal transcribed spacer (ITS) sequences, the plastid trnL intron, the trnL-F spacer, and the atpB-rbcL spacer. This genus was divided into three subgenera: Parvisepalum, Brachypetalum, and Paphiopedilum. Each of them is monophyletic with high bootstrap supports according to the highly resolved phylogenetic tree reconstructed by combined sequences. There are five sections within the subgenus Paphiopedilum, including Coryopedilum, Pardalopetalum, Cochlopetalum, Paphiopedilum, and Barbata. The subgenus Parvisepalum is phylogenetic basal, which suggesting that Parvisepalum is comprising more ancestral characters than other subgenera. The evolutionary trend of genus Paphiopedilum was deduced based on the maximum likelihood (ML) tree and Bayesian Evolutionary Analysis Sampling Trees (BEAST). Reconstruct Ancestral State in Phylogenies (RASP) analyses based on the combined sequence data. The biogeographic analysis indicates that Paphiopedilum species were firstly derived in Southern China and Southeast Asia, subsequently dispersed into the Southeast Asian archipelagoes. The subgenera Paphiopedilum was likely derived after these historical dispersals and vicariance events. Our research reveals the relevance of the differentiation of Paphiopedilum in Southeast Asia and geological history. Moreover, the biogeographic analysis explains that the significant evolutionary hotspots of these orchids in the Sundaland and Wallacea might be attributed to repeated migration and isolation events between the south-eastern Asia mainland and the Sunda Super Islands.

Collecting wild Miscanthus germplasm in Asia for crop improvement and conservation in Europe whilst adhering to the guidelines of the United Nations' Convention on Biological Diversity.

BACKGROUND AND AIMS: Germplasm with diverse, agronomically relevant traits forms the foundation of a successful plant breeding programme. Since 1993, the United Nations has been advocating the implementation of the Convention on Biological Diversity (CBD) and the subsequent 2002 Bonn Guidelines as international best practice on germplasm collection and use. In 2006, a European team made an expedition to Asia to collect wild germplasm of Miscanthus, a C4 perennial rhizomatous grass, for breeding an environmentally adaptable, resilient and high-yielding bioenergy crop. We outline general aspects of germplasm collection, conservation, breeding and biomass production evaluation while following the CBD's guidelines, respecting biodiversity and conservation needs, and the ethical use of genetic resources. METHODS: Effective protocols, quarantine, methods for collecting seed and rhizomes, and a genebank for conservation were established. Versatile informatics and database architecture were used to assist in selection, flowering synchronization, crossing, evaluation, phenotyping and data integration. Approaches were formulated to comply with the CBD guidelines. KEY RESULTS: A total of 303 accessions of M. sinensis, M. sacchariflorus and M. floridulus were collected from 158 geographically and environmentally diverse locations. These species were shown to accumulate different amounts of aerial biomass due to combinations of stem count, height and thickness. Progeny from one interspecies cross accumulated more biomass in early trials and has shown double the yield performance in years 3-4 compared with the existing commercial cultivar M. × giganteus. An example of an F1 hybrid has already demonstrated the long-term potential of exploiting this collection for a breeding programme. CONCLUSIONS: By conforming to the CBD principles, the authors' international collaboration provides a practical example of implementing the CBD. The collection widened the genetic diversity of Miscanthus available to allow for breeding of novel hybrids that exhibit more diverse traits to increase yield and resilience for growth on marginal land and in climate-challenged environments.

Nuclear and chloroplast DNA phylogeography suggests an Early Miocene southward expansion of Lithocarpus (Fagaceae) on the Asian continent and islands.

BACKGROUND: Most genera of Fagaceae are thought to have originated in the temperate regions except for the genus Lithocarpus, the stone oaks. Lithocarpus is distributed in subtropical and tropical Asia, and its ancestral population is hypothesized to be distributed in tropical regions in Borneo and Indochina. Borneo and the nearby islands (the Greater Sunda Islands) were connected to the Malay Peninsula and Indochina prior to the Pliocene epoch and formed the former Sundaland continent. The Southeast Asian Lithocarpus, is thought to have dispersed between continental Asia and the present Sundaland. The drastic climate changes during the Pliocene and Pleistocene epochs which caused periodic sea-level changes is often used to explain the cause of its diversity. The aim of this study was to establish phylogenetic relationships by analyzing nuclear (nrDNA) and chloroplast (cpDNA) DNA in order to describe and analyze the origin, causes of diversification and historical biogeography of Lithocarpus. RESULTS: Phylogeny reconstructed through the multiple-species coalescent method with nrDNA and cpDNA revealed that the continental-Asian taxa were clustered at the basal lineages. The derived lineages of tropical Lithocarpus, with the inference of a subtropical ancestral state, imply a southward migration in the Early Miocene period with subsequent in situ diversification in the Greater Sunda Islands. The gradual decrease in temperature since the Middle Miocene period is proposed as a cause of the increase in the net diversification rate. CONCLUSIONS: The historical ancestral origin of Lithocarpus has been suggested to be mainland Asia. Southward migration in the Early Miocene period with subsequent in situ diversification could explain the current diversity of stone oaks in Southeast Asia. This study also considered the multiple origins of stone oaks currently indigenous to the subtropical islands offshore and near mainland China. Our results provide phylogenetic evidence for a subtropical origin of Asian stone oaks and reveal the process of diversification and how it fits into the timeline of major geologic and climatic events rather than local, episodic, rate-shifting events.

Characterization of 39 microsatellite markers from Nuphar shimadai (Nymphaeaceae) and cross-amplification in two related taxa.

PREMISE OF THE STUDY: Microsatellite loci were developed for Nuphar shimadai (Nymphaeaceae) to evaluate the population genetic dynamics for conservation purposes. The species is an endemic aquatic species in Taiwan that is endangered by anthropogenic activities. METHODS AND RESULTS: A magnetic bead enrichment protocol was used to identify 72 potential microsatellite loci and develop 39 microsatellite markers from N. shimadai. The number of alleles per locus ranged from one to 10 per locus, with levels of observed heterozygosity ranging from 0 to 1.0 within populations. As a result of inbreeding within isolated populations, 65% of loci significantly deviated from Hardy-Weinberg equilibrium within populations. CONCLUSIONS: These novel markers should be valuable tools to evaluate the genetic diversity within the endangered aquatic taxon N. shimadai for conservation and reintroduction purposes in Taiwan.

Phylogeny of the gymnosperm genus Cycas L. (Cycadaceae) as inferred from plastid and nuclear loci based on a large-scale sampling: Evolutionary relationships and taxonomical implications.

The gymnosperm genus Cycas is the sole member of Cycadaceae, and is the largest genus of extant cycads. There are about 115 accepted Cycas species mainly distributed in the paleotropics. Based on morphology, the genus has been divided into six sections and eight subsections, but this taxonomy has not yet been tested in a molecular phylogenetic framework. Although the monophyly of Cycas is broadly accepted, the intrageneric relationships inferred from previous molecular phylogenetic analyses are unclear due to insufficient sampling or uninformative DNA sequence data. In this study, we reconstructed a phylogeny of Cycas using four chloroplast intergenic spacers and seven low-copy nuclear genes and sampling 90% of extant Cycas species. The maximum likelihood and Bayesian inference phylogenies suggest: (1) matrices of either concatenated cpDNA markers or of concatenated nDNA lack sufficient informative sites to resolve the phylogeny alone, however, the phylogeny from the combined cpDNA-nDNA dataset suggests the genus can be roughly divided into 13 clades and six sections that are in agreement with the current classification of the genus; (2) although with partial support, a clade combining sections Panzhihuaenses + Asiorientales is resolved as the earliest diverging branch; (3) section Stangerioides is not monophyletic because the species resolve as a grade; (4) section Indosinenses is not monophyletic as it includes Cycas macrocarpa and C. pranburiensis from section Cycas; (5) section Cycas is the most derived group and its subgroups correspond with geography.

Adaptive Genetic Divergence Despite Significant Isolation-by-Distance in Populations of Taiwan Cow-Tail Fir (Keteleeria davidiana var. formosana).

Double digest restriction site-associated DNA sequencing (ddRADseq) is a tool for delivering genome-wide single nucleotide polymorphism (SNP) markers for non-model organisms useful in resolving fine-scale population structure and detecting signatures of selection. This study performs population genetic analysis, based on ddRADseq data, of a coniferous species, Keteleeria davidiana var. formosana, disjunctly distributed in northern and southern Taiwan, for investigation of population adaptive divergence in response to environmental heterogeneity. A total of 13,914 SNPs were detected and used to assess genetic diversity, FST outlier detection, population genetic structure, and individual assignments of five populations (62 individuals) of K. davidiana var. formosana. Principal component analysis (PCA), individual assignments, and the neighbor-joining tree were successful in differentiating individuals between northern and southern populations of K. davidiana var. formosana, but apparent gene flow between the southern DW30 population and northern populations was also revealed. Fifteen of 23 highly differentiated SNPs identified were found to be strongly associated with environmental variables, suggesting isolation-by-environment (IBE). However, multiple matrix regression with randomization analysis revealed strong IBE as well as significant isolation-by-distance. Environmental impacts on divergence were found between populations of the North and South regions and also between the two southern neighboring populations. BLASTN annotation of the sequences flanking outlier SNPs gave significant hits for three of 23 markers that might have biological relevance to mitochondrial homeostasis involved in the survival of locally adapted lineages. Species delimitation between K. davidiana var. formosana and its ancestor, K. davidiana, was also examined (72 individuals). This study has produced highly informative population genomic data for the understanding of population attributes, such as diversity, connectivity, and adaptive divergence associated with large- and small-scale environmental heterogeneity in K. davidiana var. formosana.