Out of chaos: Phylogenomics of Asian Sonerileae.

Sonerileae is a diverse Melastomataceae lineage comprising ca. 1000 species in 44 genera, with >70% of genera and species distributed in Asia. Asian Sonerileae are taxonomically intractable with obscure generic circumscriptions. The backbone phylogeny of this group remains poorly resolved, possibly due to complexity caused by rapid species radiation in early and middle Miocene, which hampers further systematic study. Here, we used genome resequencing data to reconstruct the phylogeny of Asian Sonerileae. Three parallel datasets, viz. single-copy ortholog (SCO), genomic SNPs, and whole plastome, were assembled from genome resequencing data of 205 species for this purpose. Based on these genome-scale data, we provided the first well resolved phylogeny of Asian Sonerileae, with 34 major clades identified and 74% of the interclade relationships consistently resolved by both SCO and genomic data. Meanwhile, widespread phylogenetic discordance was detected among SCO gene trees as well as species trees reconstructed using different tree estimation methods (concatenation/site-based coalescent method/summary method) or different datasets (SCO/genomic/plastome). We explored sources of discordance using multiple approaches and found that the observed discordance in Asian Sonerileae was mainly caused by a combination of biased distribution of missing data, random noise from uninformative genes, incomplete lineage sorting, and hybridization/introgression. Exploration of these sources can enable us to generate hypotheses for future testing, which is the first step towards understanding the evolution of Asian Sonerileae. We also detected high levels of homoplasy for some characters traditionally used in taxonomy, which explains current chaotic generic delimitations. The backbone phylogeny of Asian Sonerileae revealed in this study offers a solid basis for future taxonomic revision at the generic level.

Three new species of Bredia (Sonerileae, Melastomataceae) from the Sino-Vietnamese border area.

Brediabullata, B.enchengensis, and B.nitida (Sonerileae, Melastomataceae), three species occurring in Sino-Vietnamese limestone karst regions, are described as new. Molecular phylogenetic analyses and morphological divergence indicate that these species are well separated from their close relatives in Bredia, justifying their recognition as distinct species. Brediabullata is unique in its interveinal areas prominently bullate each with an apical seta, a character otherwise never recorded in the genus. Bredianitida resembles B.malipoensis in habit, leaf shape, and inflorescence morphology, but differs in the glabrescent and nitid adaxial leaf surface (vs. densely pubescent and subvelvety), ovate-elliptic or elliptic calyx lobes (vs. triangular to semiorbicular), and white petals (vs. purplish-red). Brediaenchengensis is closest to B.longiradiosa, but easily recognized by its prostrate habit (vs. erect), the yellowish-green, membranous and fragile leaves (vs. green or dark green, papery), and white anthers (vs. pink to purplish). These new discoveries show that further botanical exploration is warranted in the remote Sino-Vietnamese bordering region.

Phylogenomic analyses based on genome-skimming data reveal cyto-nuclear discordance in the evolutionary history of Cotoneaster (Rosaceae).

As a consequence of hybridization, polyploidization, and apomixis, the genus Cotoneaster (Rosaceae) represents one of the most complicated and controversial lineages in Rosaceae, with ca. 370 species which have been classified into two subgenera and several sections, and is notorious for its taxonomic difficulty. The infrageneric relationships and taxonomy of Cotoneaster have remained poorly understood. Previous studies have focused mainly on natural hybridization involving only several species, and phylogeny based on very limited markers. In the present study, the sequences of complete chloroplast genomes and 204 low-copy nuclear genes of 72 accessions, representing 69 species as ingroups, were used to conduct the most comprehensive phylogenetic analysis so far for Cotoneaster. Based on the sequences of complete chloroplast genomes and many nuclear genes, our analyses yield two robust phylogenetic trees respectively. Chloroplast genome and nuclear data confidently resolved relationships of this genus into two major clades which largely supported current classification based on morphological evidence. However, conflicts between the chloroplast genome and low-copy nuclear phylogenies were observed in both the species level and clade level. Cyto-nuclear discordance in the phylogeny could be caused by frequent hybridization events and incomplete sorting lineage (ILS). In addition, our divergence-time analysis revealed an evolutionary radiation of the genus from late Miocene to date.

A new species of Bredia (Sonerileae, Melastomataceae) from Sichuan, China.

Bredia hispida (Sonerileae, Melastomataceae), a species occurring in southeastern Sichuan, China, is newly described based on morphological and molecular data. The generic placement of B. hispida is well supported by phylogenetic analysis and morphological characters, including basally cordate, hairy leaf blade, cymose inflorescence, basally gibbous anthers and enlarged ovary crown enclosing an inverted frustum-shaped depression. Both molecular and morphological divergence showed that B. hispida is well separated from its close relatives, justifying its recognition as a distinct species. The new species resembles B. repens, B. changii and B. guidongensis in the prostrate habit and isomorphic stamens but differs markedly in the unequal opposed leaves, the 2-4 mm long, stout bristles on the adaxial surface of leaf blade and acuminate leaf apex. Bredia hispida co-occurs with B. esquirolii in the wild. No morphologically putative hybrids between them were observed despite their overlap in flowering season. The isolating mechanism remains unclear, pending further investigation.

Recircumscription of Bredia and resurrection of Tashiroea (Sonerileae, Melastomataceae) with description of a new species T.villosa.

Bredia (Melastomataceae) is an Asian genus that extends from central and southern mainland China to Taiwan and the Ryukyu islands. Molecular phylogenetic analyses reveal that the type of Bredia is nested in a clade of 20 species, while Tashiroea, a genus previously synonymized in Bredia, falls in another distantly related clade of 10 species. Our morphological survey shows that the two clades can be distinguished by several diagnostic features including leaf indumentum, texture, leaf surface sculpture under SEM, presence/absence of yellowish uniseriate trichomes, and capsule morphology. Based on molecular and morphological evidence, Bredia is recircumscribed and Tashiroea is resurrected. Description and a list of species are provided for the two genera with the description of a new species, T.villosa.

A new species of Fordiophyton (Sonerileae, Melastomataceae) from Yunnan, China.

Fordiophytonjinpingense (Melastomataceae; Sonerileae), a species occurring in south-eastern Yunnan, China, is described as new, based on morphological and molecular data. Phylogenetic analyses, based on nrITS sequence data, showed that, except F.breviscapum, all species sampled in Fordiophyton formed a strongly supported clade in which two geographical lineages were recovered. The generic placement of F.jinpingense is well supported by phylogenetic analyses and a character combination of 4-merous flowers, distinctly dimorphic stamens and the connectives basally not calcarate. Molecular divergence and morphological evidence indicate that F.jinpingense is well separated from other members of the genus, thus justifying its recognition as a distinct species. Fordiophytonjinpingense is phylogenetically closest to F.repens, but differs markedly from the latter in stem morphology (short, obtusely 4-sided vs. long, 4-angular), habit (erect vs. creeping), leaf size (6-16.5 × 4.5-13 cm vs. 4-7.5 × 4-6.5 cm) and flower number per inflorescence (5-13 vs. 3-6).

The specific status of Melastoma kudoi (Melastomataceae, Melastomeae).

BACKGROUND: Melastoma has undergone rapid species radiation during the last one million years, and circumscription of some species in the genus has remained controversial. Melastoma kudoi, an erect species narrowly endemic to central Taiwan was previously treated as a synonym of M. intermedium, a semicreeping hybrid between the erect species M. candidum and the creeping M. dodecandrum, making its identity questionable. We addressed this question based on molecular and morphological data. RESULTS: Phylogenetic analyses based on nrITS sequence data revealed that M. kudoi is most closely related to M. dodecandrum. Further analyses of six nuclear genes (cam, chi, gapC, gbss, tpi and vr) and two chloroplast markers (trnL-trnF and psbA) showed that M. kudoi is well diverged from its close relatives. Morphologically, it is also easily distinguished from related species by its erect habit, center-positioned stigma, and spreading, basally enlarged hairs on the hypanthium. CONCLUSIONS: Both molecular phylogenetic and morphological data suggest that M. kudoi is well separated from M. intermedium, M. dodecandrum, and O. scaberrima, and should be treated as a distinct species. Taxonomic treatment and detailed description of M. kudoi are provided.

The Antibiosis Action and Rice-Induced Resistance, Mediated by a Lipopeptide from Bacillus amyloliquefaciens B014, in Controlling Rice Disease Caused by Xanthomonas oryzae pv. oryzae.

In the present study, a lipopeptide (named AXLP14) antagonistic to Xanthomonas oryzae pv. oryzae (Xoo) was obtained from the culture supernatant of Bacillus amyloliquefaciens B014. Matrix-assisted laser desorption/ionization time-of-flight mass spectrometry analysis demonstrated that AXLP14 consisted of surfactin homologs. The minimum inhibition concentration and minimum bactericidal concentration of AXLP14 against Xoo were determined to be 1.25 and 2.50 mg/ml, respectively. At a concentration of 0.613 mg/ml, AXLP14 strongly inhibited the formation of Xoo biofilm. AXLP14 also inhibited the motility of Xoo in a concentration-dependent manner. Applying AXLP14 to rice seedlings significantly reduced the incidence and severity of disease caused by Xoo. In Xoo-infected rice seedlings, AXLP14 strongly and continuously up-regulated the expression of both OsNPR1 and OsWRKY45. In addition, AXLP14 effectively inhibited the Xoo-induced up-regulation of the expression of the abscisic acid biosynthesis gene OsNECD3 and the abscisic acid signalingresponsive gene OsLip9, indicating that AXLP14 may protect rice against Xoo-induced disease by enhancing salicylic acid defense and interfering with the abscisic acid response to virulence.

De Novo Transcriptome Assembly in Firmiana danxiaensis, a Tree Species Endemic to the Danxia Landform.

Many Firmiana species are locally endemic, providing an interesting system for studying adaptation and speciation. Among these species, F. danxiaensis is a tree species endemic to Mount Danxia in Guangdong, China, which is an area known for presenting the Danxia landform. How F. danxiaensis could have adapted to the stressful environment of rocky cliffs covered with barren soils in the Danxia landform is still unknown. In this study, we performed de novo assembly of the transcriptome of F. danxiaensis, obtaining 47,221 unigenes with an N50 value of 987 bp. Homology analysis showed that 32,318 of the unigenes presented hits in the NCBI non-redundant database, and 31,857 exhibited significant matches with the protein database of Theobroma cacao. Gene Ontology (GO) annotation showed that hundreds of unigenes participated in responses to various stresses or nutritional starvation, which may help us to understand the adaptation of F. danxiaensis to Danxia landform. Additionally, we found 263 genes related to responses to Cd, partially explaining the high accumulation of Cd observed in Firmiana species. The EuKaryotic Orthologous Groups (KOG) and Kyoto Encyclopedia of Genes and Genomes (KEGG) annotations revealed many genes playing roles in the biosynthesis of secondary metabolites and environmental adaptation, which may also contribute to the survivor and success of Firmiana species in extreme environments. Based on the obtained transcriptome, we further identified a Firmiana-specific whole-genome duplication event that occurred approximately 20 Mya, which may have provided raw materials for the diversification of Firmiana species.

Natural hybridization and asymmetric introgression at the distribution margin of two Buddleja species with a large overlap.

BACKGROUND: Natural hybridization in plants is universal and plays an important role in evolution. Based on morphology it has been presumed that hybridization occurred in the genus Buddleja, though genetic studies confirming this assumption have not been conducted to date. The two species B. crispa and B. officinalis overlap in their distributions over a wide range in South-West China, and we aimed to provide genetic evidence for ongoing hybridization in this study. RESULTS: We investigated the occurrence of hybrids between the two species at the southern-most edge of the distribution of B. crispa using five nuclear loci and pollination experiments. The genetic data suggest substantial differentiation between the two species as species-specific alleles are separated by at least 7-28 mutations. The natural hybrids found were nearly all F1s (21 of 23), but backcrosses were detected, and some individuals, morphologically indistinguishable from the parental species, showed introgression. Pollen viability test shows that the percentage of viable pollen grains was 50 ± 4% for B. crispa, and 81 ± 2% for B. officinalis. This difference is highly significant (t = 7.382, p < 0.0001). Hand cross-pollination experiments showed that B. crispa is not successful as pollen-parent, but B. officinalis is able to pollinate B. crispa to produce viable hybrid seed. Inter-specific seed-set is low (8 seeds per fruit, as opposed to about 65 for intra-specific pollinations), suggesting post-zygotic reproductive barriers. In addition, one of the reference populations also suggests a history of introgression at other localities. CONCLUSIONS: The occurrence of morphologically intermediate individuals between B. crispa and B. officinalis at Xishan Mountain is unequivocally linked to hybridization and almost all examined individuals of the putative hybrids were likely F1s. Despite pollination experiments indicating higher chances for introgression into B. officinalis (hybrids only produced viable seed when crossed with B. officinalis), observed introgression was asymmetrical into B. crispa. This could be due to seeds produced by hybrids not contributing to seedlings, or other factors favoring the establishment of backcrosses towards B. crispa. However, further research will be needed to confirm these observations, as the small number of plants used for the pollination experiments could have introduced an artifact, for example if used individuals were more or less compatible than the species average, and also the small number of loci used could convey a picture of introgression that is not representative for the whole genome.

[Diversity and tissue distribution of fungal endophytes in Alpinia officinarum: an important south-China medicinal plant].

In the present study, terminal-restriction fragment length polymorphism (T-RFLP) technique was applied to assess the diversity and tissue distribution of the fungal endophyte communities of Alpinia officinarum collected from Longtang town in Xuwen county, Guangdong province, China, at which the pharmacological effect of the medicine plant is traditional considered to be the significantly higher than that in any other growth areas in China. A total of 28 distinct Terminal-Restriction Fragment (T-RFs) were detected with HhaI Mono-digestion targeted amplified fungal nuclear ribosomal internal transcribed spacer region sequences (rDNA ITS) from the root, rhizome, stem, and leaf internal tissues of A. officinarum plant, indicating that at least 28 distinct fungal species were able to colonize the internal tissue of the host plant. The rDNA ITS-T-RFLP profiles obtained from different tissues of the host plant were obvious distinct. And the numbers of total T-RFs, and the dominant T-RFs detected from various tissues were significantly different. Based on the obtained T-RFLP profiles, Shannon's diversity index and the Shannon's evenness index were calculated, which were significantly different among tissues (P < 0.05). Furthermore, two types of active chemicals, total volatile oils by water vapor distillation method and galangin by methanol extraction-HPLC method, were examined in the each tissue of the tested plant. Both of tested components were detected in all of the four tissues of the medicine plant with varying contents. And the highest was in rhizome tissue. Correlation analysis revealed there were significant negative correlations between both of the tested active components contents and calculated Shannon's diversity index, as well as the Shannon's evenness index of the fungal endophyte communities of the host plant (P = 0, Pearson correlation coefficient ≤ -0.962), and significant positive correlations between both of the tested active components contents and 325 bp dominant T-RF linkage to Pestalotiopsis (P = 0, Pearson correlation coefficient ≥ 0.975). In conclusion, A. officinarum is colonized by diverse fungal endophytes communities. The diversity of the fungal endophytes was found in the A. officinarum varied with differences of the tissue types of the host plants and was closely correlated with the accumulation of main active components, total volatile oils and galangin contents in the host plant tissue.

Development of 11 microsatellite loci for an endangered herb, Paraisometrum mileense (Gesneriaceae), in southwest China.

PREMISE OF THE STUDY: Microsatellite markers were developed to investigate the genetic diversity and population structure of the endangered species Paraisometrum mileense, as well as its evolutionary history. • METHODS AND RESULTS: Using the Fast Isolation by AFLP of Sequences COntaining repeats (FIASCO) protocol, 11 polymorphic primer sets were obtained in P. mileense. The number of alleles per locus ranged from one to nine with an average 3.8, and the observed and expected heterozygosities ranged from 0 to 1 and 0 to 0.89, respectively, for the available three populations. • CONCLUSIONS: These markers may be useful for further investigation of the conservation genetics of P. mileense.

Development and characterization of nine microsatellites for an endangered tree, Pinus wangii (Pinaceae).

PREMISE OF THE STUDY: Pinus wangii is an endemic and endangered species in southwestern China, and microsatellite primers were developed to characterize its genetic diversity and population structure. • METHODS AND RESULTS: Using the Fast Isolation by AFLP of Sequences COntaining repeats (FIASCO) protocol, nine sets of microsatellite primers were developed in P. wangii. One population with 26 individuals of P. wangii, as well as 11 individuals each for two congeneric species, P. taiwanensis and P. squamata, were used to test their polymorphism and transferability. The number of alleles per locus ranged from one to seven with an average of 3.7, and the observed heterozygosity and expected heterozygosity ranged from 0 to 0.91 and 0 to 0.75, respectively. • CONCLUSIONS: We developed nine sets of polymorphic microsatellite loci that are suitable for investigating genetic diversity and population structure of P. wangii, and these markers may be useful for other Pinus species.

Development of microsatellite loci for the invasive weed Wedelia trilobata (Asteraceae).

PREMISE OF THE STUDY: Microsatellite markers were developed to help elucidate the population genetics of the invasive species Wedelia trilobata. • METHODS AND RESULTS: Using the Fast Isolation by AFLP of Sequences COntaining (FIASCO) repeats protocol, 23 sets of primers for amplifying microsatellite loci were identified in W. trilobata, 10 of which showed polymorphism (two to five alleles per locus) in samples of two populations of W. trilobata, one from China and one from Peru. Six of these loci were successfully amplified from samples of the native congener W. chinensis, with expected sizes. • CONCLUSIONS: These markers may be useful for further investigation of population genetics of Wedelia trilobata and other congener species.

[Effects of divergent habitat on genetic structure of population of Excoecaria agallocha, a mangrove associate].

Genetic diversity and population genetic structure of Excoecaria agallocha, a typical mangrove associate species,were surveyed at divergent habitats (intertidal and inland). In general, intertidal populations had higher genetic diversity than inland populations. Genetic differentiation among intertidal populations (G(ST) = 0.191) were smaller than that among inland populations (G(ST) = 0.218), suggesting that gene flow via seed among intertidal populations were stronger. In an analysis of molecular variance (AMOVA), we found that 15.13% of the genetic variance could be explained by the differentiation between habitats, as compared to only 11.63% to geographical effects among five sits 181 -759 km distant from each other. This implies that markedly selection regimes result in habitat adaptation. Isolation-by-distance, Southwest Monsoon Current,China Coastal Current and genetic drift played important role in genetic differentiation of China population of Excoecaria agalocha.