Integration of large and diverse angiosperm DNA fragments into Asian Gnetum mitogenomes.

BACKGROUND: Horizontal gene transfer (HGT) events have rarely been reported in gymnosperms. Gnetum is a gymnosperm genus comprising 25‒35 species sympatric with angiosperms in West African, South American, and Southeast Asian rainforests. Only a single acquisition of an angiosperm mitochondrial intron has been documented to date in Asian Gnetum mitogenomes. We wanted to develop a more comprehensive understanding of frequency and fragment length distribution of such events as well as their evolutionary history in this genus. RESULTS: We sequenced and assembled mitogenomes from five Asian Gnetum species. These genomes vary remarkably in size and foreign DNA content. We identified 15 mitochondrion-derived and five plastid-derived (MTPT) foreign genes. Our phylogenetic analyses strongly indicate that these foreign genes were transferred from diverse eudicots-mostly from the Rubiaceae genus Coptosapelta and ten genera of Malpighiales. This indicates that Asian Gnetum has experienced multiple independent HGT events. Patterns of sequence evolution strongly suggest DNA-mediated transfer between mitochondria as the primary mechanism giving rise to these HGT events. Most Asian Gnetum species are lianas and often entwined with sympatric angiosperms. We therefore propose that close apposition of Gnetum and angiosperm stems presents opportunities for interspecific cell-to-cell contact through friction and wounding, leading to HGT. CONCLUSIONS: Our study reveals that multiple HGT events have resulted in massive amounts of angiosperm mitochondrial DNA integrated into Asian Gnetum mitogenomes. Gnetum and its neighboring angiosperms are often entwined with each other, possibly accounting for frequent HGT between these two phylogenetically remote lineages.

Hedyotislongiramulis (Rubiaceae), a new species from south China.

Hedyotislongiramulissp. nov. (Rubiaceae) is described from Guangdong Province, China. It is similar to H.caudatifolia but differs in having puberulent, more or less tetragonal and decussately sulcate juvenile stems, waxy leaf surface, short inflorescence peduncles, high length ratio of corolla lobe to tube, and subglobose capsules. The phylogenetic analysis reveals that H.longiramulis is sister to H.pubirachis. Dimorphism concerning pollen size was observed in the heterostylous flowers. The complete chloroplast genome of the new species comprises a typical quadripartite structure of 153,616 bp in length, with two inverted repeats of 25,457 bp, a large single-copy of 85,050 bp and a small single-copy of 17,652 bp. It contains 112 unique genes, including 79 protein-coding genes, 29 tRNA genes, and four rRNA genes, the GC content of the chloroplast genome is 32.4%. The new species is provisionally evaluated as "Least Concern" because it is common and well-protected in two Provincial Nature Reserves.

Phylogenomic Analysis Reconstructed the Order Matoniales from Paleopolyploidy Veil.

Phylogenetic conflicts limit our understanding of the evolution of terrestrial life under multiple whole genome duplication events, and the phylogeny of early terrestrial plants remains full of controversy. Although much incongruence has been solved with so-called robust topology based on single or lower copy genes, the evolutionary mechanisms behind phylogenetic conflicts such as polyploidization remain poorly understood. Here, through decreasing the effects of polyploidization and increasing the samples of species, which represent all four orders and eight families that comprise early leptosporangiate ferns, we have reconstructed a robust phylogenetic tree and network with 1125 1-to-1 orthologs based on both coalescent and concatenation methods. Our data consistently suggest that Matoniales, as a monophyletic lineage including Matoniaceae and Dipteridaceae, should be redefined as an ordinal rank. Furthermore, we have identified and located at least 11 whole-genome duplication events within the evolutionary history of four leptosporangiates lineages, and associated polyploidization with higher speciation rates and mass extinction events. We hypothesize that paleopolyploidization may have enabled leptosporangiate ferns to survive during mass extinction events at the end Permian period and then flourish throughout the Mesozoic era, which is supported by extensive fossil records. Our results highlight how ancient polyploidy can result in rapid species radiation, thus causing phylogenetic conflicts yet allowing plants to survive and thrive during mass extinction events.

Convergent molecular evolution of phosphoenolpyruvate carboxylase gene family in C4 and crassulacean acid metabolism plants.

Phosphoenolpyruvate carboxylase (PEPC), as the key enzyme in initial carbon fixation of C4and crassulacean acid mechanism (CAM) pathways, was thought to undergo convergent adaptive changes resulting in the convergent evolution of C4 and CAM photosynthesis in vascular plants. However, the integral evolutionary history and convergence of PEPC in plants remain poorly understood. In the present study, we identified the members of PEPC gene family across green plants with seventeen genomic datasets, found ten conserved motifs and modeled three-dimensional protein structures of 90 plant-type PEPC genes. After reconstructing PEPC gene family tree and reconciled with species tree, we found PEPC genes underwent 71 gene duplication events and 16 gene loss events, which might result from whole-genome duplication events in plants. Based on the phylogenetic tree of the PEPC gene family, we detected four convergent evolution sites of PEPC in C4 species but none in CAM species. The PEPC gene family was ubiquitous and highly conservative in green plants. After originating from gene duplication of ancestral C3-PEPC, C4-PEPC isoforms underwent convergent molecular substitution that might facilitate the convergent evolution of C4 photosynthesis in Angiosperms. However, there was no evidence for convergent molecular evolution of PEPC genes between CAM plants. Our findings help to understand the origin and convergent evolution of C4 and CAM plants and shed light on the adaptation of plants in dry, hot environments.

Dryopteriswulingshanensis (Dryopteridaceae), a new species from Hunan, China.

Dryopteriswulingshanensis, a new species growing on limestone in the Wulingshan Mountains, Hunan, China, is described and illustrated. This species is most similar to D.jishouensis and D.gymnophylla on general morphological traits, such as the form of scales, rhizome and sori, but differs by the number of vascular bundles at the base of the petiole, length to width ratio of lamina, stalk length of basal pinnae, division of the lamina, apex form of the pinnule and habitat. Moreover, molecular phylogenetic analysis using the chloroplast rbcL gene suggested that D.wulingshanensis, as the sister group of D.jishouensis, is a monophyletic clade. According to its restricted geographic range, small populations and few individuals, D.wulingshanensis should be considered endangered, according to the IUCN Red List criteria.

Morphology and molecules support the new monotypic genus Parainvolucrella (Rubiaceae) from Asia.

Parainvolucrella R.J. Wang, a new monotypic genus for P.scabra (Wall. ex Kurz) M.D.Yuan & R.J.Wang, new combination, is segregated from the Hedyotis-Oldenlandia complex, based on morphological and molecular evidence. Phylogenetically, the new genus is sister to Scleromitrion, from which it differs by a combination of morphological characters: herbaceous habit, terminal inflorescence with subtended leaves, heterostylous flowers, indehiscent fruits and pollen with double microreticulate tectum. A key to the genera of the Hedyotis-Oldenlandia complex in China is provided for further identification.

Chromosomal-level assembly of the Leptodermis oblonga (Rubiaceae) genome and its phylogenetic implications.

Rubiaceae is the fourth largest and a taxonomically complex family of angiosperms. Many species in this family harbor low reproductive isolation and frequently exhibit inconsistent phenotypic characteristics. Therefore, taxonomic classification and their phylogenetic relationships in the Rubiaceae family is challenging, especially in the genus Leptodermis. Considering the low taxonomic confusion and wide distribution, Leptodermis oblonga is selected as a representative Leptodermis for genome sequencing. The assemblies resulted in 497 Mbp nuclear and 155,100 bp chloroplast genomes, respectively. Using the nuclear genome as a reference, SNPs were called from 37 Leptodermis species or varieties. The phylogenetic tree based on SNPs exhibited high resolution for species delimitation of the complex and well-resolved phylogenetic relationships in the genus. Moreover, 28,987 genes were predicted in the nuclear genome and used for comparative genomics study. As the first chromosomal-level genome of the subfamily Rubioideae in Rubiaceae, it will provide fruitfully evolutionary understanding in the family.

Morphology and molecules support the new monotypic genus Fenghwaia (Rhamnaceae) from south China.

Fenghwaia, a new monotypic genus, along with the new species Fenghwaia gardeniicarpa, is described from Guangdong Province, China. The combined features of inferior ovary, cylindrical drupaceous fruits and orbicular and dorsiventrally-compressed seeds with an elongate and pronounced basal appendage make the new genus significantly different from other genera of the family. In addition, its pollen morphology also showed great similarity to other species of this stenopalynous family. The molecular phylogenetic analysis, based on nuclear ribosomal internal transcribed spacer (ITS) and plastid trnL-F intron spacer (trnL-F) DNA sequence data from the new genus and the other 375 species representing 58 genera of Rhamnaceae, indicates that Fenghwaia is nested within the 'rhamnoid' group and sister to the tribe Rhamneae and then both sister to the tribe Maesopsideae. A taxonomic classification key to the 'rhamnoid' group is provided, based on morphological characters. A global conservation assessment is also performed and classifies Fenghwaia gardeniicarpa as Near Threatened (NT).

Nutritional and Functional Properties of Wild Food-Medicine Plants From the Coastal Region of South China.

Food-medicine plants play an important role in providing nutrition and treating chronic diseases, especially in many minority communities and developing regions. The coastal region of South China has abundant resources of medicinal plants. A long history of cross-cultural medicinal practices among different minority groups has facilitated the development of a remarkable dietary culture by using food-medicine plants. However, integrative ethnobotanical research on both nutritional and functional properties of the food-medicine plants in this region is still limited. In this study, 27 commonly used wild food-medicine plants were recorded and analyzed from the coastal region of South China. Most of them are good sources for calcium (47.83-1099.89 mg/100 g fresh weight), dietary fiber (3.00-31.87 mg/100 g fresh weight), iron (1.17-24.73 mg/100 g fresh weight), and vitamin C (0.44-68.32 mg/100 g fresh weight). Solanum americanum has the highest average nutritive value and is also considered to be good sources for proteins (7.90 g/100 g fresh weight). Medicinal properties of the studied species can be classified into 8 categories: treatment of the damp-heat syndrome, digestive diseases, urologic diseases, arthropathy, respiratory diseases, gynecological diseases, snake or insect bites, and uses as a tonic. Treating the damp-heat syndrome or expelling warm pathogenic factors is the most commonly used ethnomedicinal practice in the study area. The present study highlights that the local ethnomedicinal practices are deeply influenced by local natural conditions and customs. Food-medicine plants with superior key nutrients have been used regularly in the diet as medicinal food to alleviate common endemic diseases.

Development of microsatellite markers for a monotypic and globally endangered species, Glyptostrobus pensilis (Cupressaceae).

PREMISE OF THE STUDY: Microsatellite markers were developed to facilitate studies of genetic diversity and structure in Glyptostrobus pensilis, a critically endangered and monotypic conifer species. METHODS AND RESULTS: Using restriction site-associated DNA sequencing (RAD-Seq), we developed 10 polymorphic and 27 monomorphic microsatellite markers. Polymorphism was characterized using 333 individuals from nine populations. The number of alleles per locus ranged from one to 14 at the population level. The levels of observed and unbiased expected heterozygosities varied from 0.058 to 0.844 and 0.219 to 0.583, respectively. Nine of these 10 polymorphic markers were successfully cross-amplified in Taxodium distichum, the species most closely related to G. pensilis. CONCLUSIONS: These microsatellite markers can be used to reveal the genetic diversity in existing populations of G. pensilis, enabling its conservation and restoration.

Endangered but genetically stable-Erythrophleum fordii within Feng Shui woodlands in suburbanized villages.

Feng Shui woodlands are naturally or artificially formed green areas in southern China. They are precious for maintaining ecosystem balance in modern semiurban environments. However, they are generally small and geographically isolated from each other, and the status of genetic diversity of the plant species within them has been almost neglected. Therefore, we studied the genetic diversity of the endangered Erythrophleum fordii in eight Feng Shui woodlands (a total of 1,061 individuals) in Guangzhou, a large city in southern China, using microsatellites. For comparison, one population with 33 individuals sampled in a nature reserve was also studied. Although our results indicate that significant demographic declines occurred historically in E. fordii, such declines have not resulted in consistent reductions in genetic variation over generations in Feng Shui populations in the recent past, and the levels of genetic variation in these populations were higher than or comparable to the genetic variation of the population in the nature reserve. In addition, our parentage and paternity analyses indicated widespread and potential long-distance pollen flow within one Feng Shui woodland, indicating the presence of an unbroken pollination network, which would at least partially alleviate the genetic erosion due to habitat fragmentation and the unequal gene contributions of E. fordii parents to their progenies when favorable recruitment habitats are absent under most of the parent trees. Overall, our results suggest that E. fordii in Feng Shui woodlands may not be driven to extinction in the near future. Nevertheless, uncontrolled fast urban development with a lack of awareness of Feng Shui woodlands will cause the local extinction of E. fordii, which has already happened in some Feng Shui woodlands.

Spiradiclis pengshuiensis (Ophiorrhizeae, Rubioideae), a new species from Chongqing, China.

Spiradiclis pengshuiensis Bo Pan & R. J. Wang (Rubiaceae) is described as a new species from Chongqing in SW China. It is morphologically compared with Spiradiclis pauciflora L. Wu & Q. R. Liu because of their similarities in habit, pubescent surface, small leaf laminas and subglobose capsules. Its conservation status is evaluated as "VU" according to the IUCN categories and criteria.

Phylogeny of the Asian Hedyotis-Oldenlandia complex (Spermacoceae, Rubiaceae): evidence for high levels of polyphyly and the parallel evolution of diplophragmous capsules.

Generic delimitation in the Hedyotis-Oldenlandia complex has a long taxonomically confused history because of the controversy of lumping or splitting these two taxa. Previous morphological and phylogenetic studies with a paucity of Asian taxa suggested that Hedyotis should include only Asian species characterized by diplophragmous capsules. In order to test the reliability of this conclusion, assess the phylogenetic value of capsular characters, and evaluate generic circumscriptions in this complex, a phylogenetic study based on expanded inclusion of 63 Asian species was performed using two nuclear regions and eight plastid regions with parsimony and likelihood analyses. The results show that the Hedyotis-Oldenlandia complex is a highly polyphyletic group. Hedyotis should only include most Asian species with erect, robust herbs or shrubs and diplophragmous capsules. Oldenlandia s. str. consists primarily of African species, including the type O. corymbosa, that are characterized by small herbs, paniculate or corymbose inflorescences, inserted styles and stamens, and loculicidally dehiscent capsules. Dimetia, Scleromitrion and Thecagonum are proposed to be resurrected to accommodate three newly resolved clades. Morphological character optimizations indicate that the diplophragmous capsule evolved independently twice within this complex. Plant habit, stipule shape, and capsular dehiscent pattern are of great value in generic circumscriptions.

Comparative chloroplast genomes of pinaceae: insights into the mechanism of diversified genomic organizations.

Pinaceae, the largest family of conifers, has diversified organizations of chloroplast genomes (cpDNAs) with the two typical inverted repeats (IRs) highly reduced. To unravel the mechanism of this genomic diversification, we examined the cpDNA organizations from 53 species of the ten Pinaceous genera, including those of Larix decidua (122,474 bp), Picea morrisonicola (124,168 bp), and Pseudotsuga wilsoniana (122,513 bp), which were firstly elucidated. The results uncovered four distinct cpDNA forms (A-C and P) that are due to rearrangements of two ∼20 and ∼21 kb specific fragments. The C form was documented for the first time and the A form might be the most ancestral one. In addition, only the individuals of Ps. macrocarpa and Ps. wilsoniana were detected to have isomeric cpDNA forms. Three types (types 1-3) of Pinaceae-specific repeats situated nearby the rearranged fragments were found to be syntenic. We hypothesize that type 1 (949 ± 343 bp) and type 3 (608 ± 73 bp) repeats are substrates for homologous recombination (HR), whereas type 2 repeats are likely inactive for HR because of their relatively short sizes (151 ± 30 bp). Conversions among the four distinct forms may be achieved by HR and mediated by type 1 or 3 repeats, thus resulting in increased diversity of cpDNA organizations. We propose that in the Pinaceae cpDNAs, the reduced IRs have lost HR activity, then decreasing the diversity of cpDNA organizations, but the specific repeats that the evolution endowed Pinaceae complement the reduced IRs and increase the diversity of cpDNA organizations.

Dynamics and evolution of the inverted repeat-large single copy junctions in the chloroplast genomes of monocots.

BACKGROUND: Various expansions or contractions of inverted repeats (IRs) in chloroplast genomes led to fluxes in the IR-LSC (large single copy) junctions. Previous studies revealed that some monocot IRs contain a trnH-rps19 gene cluster, and it has been speculated that this may be an evidence of a duplication event prior to the divergence of monocot lineages. Therefore, we compared the organizations of genes flanking two IR-LSC junctions in 123 angiosperm representatives to uncover the evolutionary dynamics of IR-LSC junctions in basal angiosperms and monocots. RESULTS: The organizations of genes flanking IR-LSC junctions in angiosperms can be classified into three types. Generally each IR of monocots contains a trnH-rps19 gene cluster near the IR-LSC junctions, which differs from those in non-monocot angiosperms. Moreover, IRs expanded more progressively in monocots than in non-monocot angiosperms. IR-LSC junctions commonly occurred at polyA tract or A-rich regions in angiosperms. Our RT-PCR assays indicate that in monocot IRA the trnH-rps19 gene cluster is regulated by two opposing promoters, S10A and psbA. CONCLUSION: Two hypotheses are proposed to account for the evolution of IR expansions in monocots. Based on our observations, the inclusion of a trnH-rps19 cluster in majority of monocot IRs could be reasonably explained by the hypothesis that a DSB event first occurred at IRB and led to the expansion of IRs to trnH, followed by a successive DSB event within IRA and lead to the expansion of IRs to rps19 or to rpl22 so far. This implies that the duplication of trnH-rps19 gene cluster was prior to the diversification of extant monocot lineages. The duplicated trnH genes in the IRB of most monocots and non-monocot angiosperms have distinct fates, which are likely regulated by different expression levels of S10A and S10B promoters. Further study is needed to unravel the evolutionary significance of IR expansion in more recently diverged monocots.