Phylogeny and Historical Biogeography of the East Asian Clematis Group, Sect. Tubulosae, Inferred from Phylogenomic Data.

The evolutionary history of Clematis section Tubulosae, an East Asian endemic lineage, has not been comprehensively studied. In this study, we reconstruct the phylogeny of this section with a complete sampling using a phylogenomic approach. The genome skimming method was applied to obtain the complete plastome sequence, the nuclear ribosomal DNA (nrDNA), and the nuclear SNPs data for phylogenetic reconstruction. Using a Bayesian molecular clock approach and ancestral range reconstruction, we reconstruct biogeographical history and discuss the biotic and abiotic factors that may have shaped the distribution patterns of the section. Both nuclear datasets better resolved the phylogeny of the sect. Tubulosae than the plastome sequence. Sect. Tubulosae was resolved as a monophyletic group sister to a clade mainly containing species from the sect. Clematis and sect. Aspidanthera. Within sect. Tubulosae, two major clades were resolved by both nuclear datasets. Two continental taxa, C. heracleifolia and C. tubulosa var. ichangensis, formed one clade. One continental taxon, C. tubulosa, and all the other species from Taiwan island, the Korean peninsula, and the Japanese archipelago formed the other clade. Molecular dating results showed that sect. Tubulosae diverged from its sister clade in the Pliocene, and all the current species diversified during the Pleistocene. Our biogeographical reconstruction suggested that sect. Tubulosae evolved and began species diversification, most likely in mainland China, then dispersed to the Korean peninsula, and then expanded its range through the Japanese archipelago to Taiwan island. Island species diversity may arise through allopatric speciation by vicariance events following the range fragmentation triggered by the climatic oscillation and sea level change during the Pleistocene epoch. Our results highlight the importance of climatic oscillation during the Pleistocene to the spatial-temporal diversification patterns of the sect. Tubulosae.

Genome-partitioning strategy, plastid and nuclear phylogenomic discordance, and its evolutionary implications of Clematis (Ranunculaceae).

Clematis is one of the largest genera of Ranunculaceae with many phylogenetic problems left to be resolved. Clematis species have considerable genome size of more than 7 Gbp, and there was no whole-genome reference sequence published in this genus. This raises difficulties in acquiring nuclear genome data for its phylogenetic analysis. Previous studies based on Sanger sequencing data, plastid genome data, and nrDNA sequences did not well resolve the phylogeny of Clematis. In this study, we used genome skimming and transcriptome data to assemble the plastid genome sequences, nuclear single nucleotide polymorphisms (SNPs) datasets, and single-copy nuclear orthologous genes (SCOGs) to reconstruct the phylogenetic backbone of Clematis, and test effectiveness of these genome partitioning methods. We also further analyzed the discordance among nuclear gene trees and between plastid and nuclear phylogenies. The results showed that the SCOGs datasets, assembled from transcriptome method, well resolved the phylogenetic backbone of Clematis. The nuclear SNPs datasets from genome skimming method can also produce similar results with the SCOGs data. In contrast to the plastid phylogeny, the phylogeny resolved by nuclear genome data is more robust and better corresponds to morphological characters. Our results suggested that rapid species radiation may have generated high level of incomplete lineage sorting, which was the major cause of nuclear gene discordance. Our simulation also showed that there may have been frequent interspecific hybridization events, which led to some of the cyto-nuclear discordances in Clematis. This study not only provides the first robust phylogenetic backbone of Clematis based on nuclear genome data, but also provides suggestions of genome partitioning strategies for the phylogenomic study of other plant taxa.

A phylotranscriptome study using silica gel-dried leaf tissues produces an updated robust phylogeny of Ranunculaceae.

The utility of transcriptome data in plant phylogenetics has gained popularity in recent years. However, because RNA degrades much more easily than DNA, the logistics of obtaining fresh tissues has become a major limiting factor for widely applying this method. Here, we used Ranunculaceae to test whether silica-dried plant tissues could be used for RNA extraction and subsequent phylogenomic studies. We sequenced 27 transcriptomes, 21 from silica gel-dried (SD-samples) and six from liquid nitrogen-preserved (LN-samples) leaf tissues, and downloaded 27 additional transcriptomes from GenBank. Our results showed that although the LN-samples produced slightly better reads than the SD-samples, there were no significant differences in RNA quality and quantity, assembled contig lengths and numbers, and BUSCO comparisons between two treatments. Using these data, we conducted phylogenomic analyses, including concatenated- and coalescent-based phylogenetic reconstruction, molecular dating, coalescent simulation, phylogenetic network estimation, and whole genome duplication (WGD) inference. The resulting phylogeny was consistent with previous studies with higher resolution and statistical support. The 11 core Ranunculaceae tribes grouped into two chromosome type clades (T- and R-types), with high support. Discordance among gene trees is likely due to hybridization and introgression, ancient genetic polymorphism and incomplete lineage sorting. Our results strongly support one ancient hybridization event within the R-type clade and three WGD events in Ranunculales. Evolution of the three Ranunculaceae chromosome types is likely not directly related to WGD events. By clearly resolving the Ranunculaceae phylogeny, we demonstrated that SD-samples can be used for RNA-seq and phylotranscriptomic studies of angiosperms.

Corrigendum: Comparative Analysis of Complete Chloroplast Genomes of 13 Species in Epilobium, Circaea, and Chamaenerion and Insights into Phylogenetic Relationships of Onagraceae.

[This corrects the article DOI: 10.3389/fgene.2021.730495.].

Comparative Analysis of Complete Chloroplast Genomes of 13 Species in Epilobium, Circaea, and Chamaenerion and Insights Into Phylogenetic Relationships of Onagraceae.

The evening primrose family, Onagraceae, is a well defined family of the order Myrtales, comprising 22 genera widely distributed from boreal to tropical areas. In this study, we report and characterize the complete chloroplast genome sequences of 13 species in Circaea, Chamaenerion, and Epilobium using a next-generation sequencing method. We also retrieved chloroplast sequences from two other Onagraceae genera to characterize the chloroplast genome of the family. The complete chloroplast genomes of Onagraceae encoded an identical set of 112 genes (with exclusion of duplication), including 78 protein-coding genes, 30 transfer RNAs, and four ribosomal RNAs. The chloroplast genomes are basically conserved in gene arrangement across the family. However, a large segment of inversion was detected in the large single copy region of all the samples of Oenothera subsect. Oenothera. Two kinds of inverted repeat (IR) region expansion were found in Oenothera, Chamaenerion, and Epilobium samples. We also compared chloroplast genomes across the Onagraceae samples in some features, including nucleotide content, codon usage, RNA editing sites, and simple sequence repeats (SSRs). Phylogeny was inferred by the chloroplast genome data using maximum-likelihood (ML) and Bayesian inference methods. The generic relationship of Onagraceae was well resolved by the complete chloroplast genome sequences, showing potential value in inferring phylogeny within the family. Phylogenetic relationship in Oenothera was better resolved than other densely sampled genera, such as Circaea and Epilobium. Chloroplast genomes of Oenothera subsect. Oenothera, which are biparental inheritated, share a syndrome of characteristics that deviate from primitive pattern of the family, including slightly expanded inverted repeat region, intron loss in clpP, and presence of the inversion.

Natural Hybrid Origin of the Controversial "Species" Clematis × pinnata (Ranunculaceae) Based on Multidisciplinary Evidence.

Interspecific hybridization is common and has often been viewed as a driving force of plant diversity. However, it raises taxonomic problems and thus impacts biodiversity estimation and biological conservation. Although previous molecular phylogenetic studies suggested that interspecific hybridization may be rather common in Clematis, and artificial hybridization has been widely applied to produce new Clematis cultivars for nearly two centuries, the issue of natural hybridization of Clematis has never been addressed in detail. In this study, we tested the hybrid origin of a mesophytic and cold-adapted vine species, Clematis pinnata, which is a rare and taxonomically controversial taxon endemic to northern China. Using field investigations, flow cytometry (FCM), phylogenomic analysis, morphological statistics, and niche modeling, we tested hybrid origin and species status of C. pinnata. The FCM results showed that all the tested species were homoploid (2n = 16). Phylonet and HyDe analyses based on transcriptome data showed the hybrid origins of C. × pinnata from either C. brevicaudata × C. heracleifolia or C. brevicaudata × C. tubulosa. The plastome phylogeny depicted that C. × pinnata in different sampling sites originated by different hybridization events. Morphological analysis showed intermediacy of C. × pinnata between its putative parental species in many qualitative and quantitative characters. Niche modeling results suggested that C. × pinnata had not been adapted to a novel ecological niche independent of its putative parents. These findings demonstrated that plants of C. × pinnata did not formed a self-evolved clade and should not be treated as a species. The present study also suggests that interspecific hybridization is a common mechanism in Clematis to generate diversity and variation, and it may play an important role in the evolution and diversification of this genus. Our study implies that morphological diversity caused by natural hybridization may overstate the real species diversity in Clematis.

An updated phylogenetic and biogeographic analysis based on genome skimming data reveals convergent evolution of shrubby habit in Clematis in the Pliocene and Pleistocene.

Convergent evolution, often viewed as the inevitable outcome of natural selection, has received special attention since the time of Darwin. Clematis is well known for its climbing habit, but it has some shrubby species, known as sect. Fruticella s.l. The shrubby Clematis species are distributed in the dry habitats of Central Asia and adjacent areas showing possible convergent evolution. In this study, we assembled the complete plastome and nuclear ribosomal DNA (nrDNA) sequences of 56 Clematis species, representing most sections and covering most of the shrubby species, to reconstruct their evolutionary histories. Using both maximum likelihood and Bayesian methods, the plastome and nrDNA datasets generated similar, but not identical, phylogenetic relationships, which are better resolved than in previous studies. Then, molecular dating, historical range reconstruction, and character optimization analyses were conducted based on this updated phylogenetic framework. All the morphological characters widely used for taxonomy were shown to have evolved multiple times. Molecular dating inferred that Clematis diverged from its sister in the mid Miocene, and all six major clades of Clematis originated during the late Miocene, with a species radiation during the Pliocene to Pleistocene. The results clearly showed that the shrubby habit evolved independently in four lineages of Clematis in Asia. We also revealed that the shrubby lineages have emerged since the very beginning of Pliocene. Asian monsoon variation in the Pliocene and glacial period fluctuation in the Pleistocene may be the driving forces for the origin and diversification of the shrubby Clematis in Central Asia and adjacent dry areas.

The complete chloroplast genome of a distinctive meadow-rue, Thalictrum foeniculaceum (Ranunculaceae).

Thalictrum foeniculaceum is a morphologically distinctive species in the genus with very reduced filiform compound leaves and big pinkish flowers occurring in dry slopes of northern China. Herein, we report the first complete chloroplast genome sequence of T. foeniculaceum. The chloroplast genome sequence was 155,923 bp in length, with large and small single-copy regions (LSC with 85,323 bp and SSC with 17,628 bp in length) separated by two inverted repeat regions (IRs with 26,486 bp). The total GC content was 38.3%. The complete plastome sequence contained 111 genes, including 77 protein-coding, 30 tRNA, and four rRNA genes. The phylogenetic analysis of Thalictrum based on the complete chloroplast genomes available online was also presented in this study.

Predictive modelling of the distribution of Clematis sect. Fruticella s. str. under climate change reveals a range expansion during the Last Glacial Maximum.

BACKGROUND: The knowledge of distributional dynamics of living organisms is a prerequisite for protecting biodiversity and for the sustainable use of biotic resources. Clematis sect. Fruticella s. str. is a small group of shrubby, yellow-flowered species distributed mainly in arid and semi-arid areas of China. Plants in this section are both horticulturally and ecologically important. METHODS: Using past, present, and future environmental variables and data with Maximum Entropy (Maxent) modeling, we evaluated the importance of the environmental variables on the section's estimated distributions, thus simulating its distributional dynamics over time. The contractions and expansions of suitable habitat between the past and future scenarios and the present were then compared. RESULTS AND DISCUSSION: The models revealed that the areas with high and moderate suitability currently encompass about 725,110 km2. The distribution centroid location varies between points in Ningxia and Inner Mongolia during the different scenarios. Elevation, Mean UV-B of Lowest Month, Precipitation of Coldest Quarter, and Mean Temperature of Driest Quarter were major factors determining the section's distribution. Our modeling indicated that Clematis sect. Fruticella underwent a significant range contraction during the last interglacial period, and then expanded during the last glacial maximum (LGM) to amounts like those of the present. Cold, dry, and relatively stable climate, as well as steppe or desert steppe environments may have facilitated range expansion of this cold-adapted, drought-resistant plant taxon during the LGM. Predicted future scenarios show little change in the amounts of suitable habitat for Clematis sect. Fruticella. This study aids understanding of the distributional dynamics of Clematis sect. Fruticella, and the results will help the conservation and sustainable use of these important woody plants in Chinese arid and semiarid areas.

Structural variation of the complete chloroplast genome and plastid phylogenomics of the genus Asteropyrum (Ranunculaceae).

Two complete chloroplast genome sequences of Asteropyrum, as well as those of 25 other species from Ranunculaceae, were assembled using both Illumina and Sanger sequencing methods to address the structural variation of the cp genome and the controversial systematic position of the genus. Synteny and plastome structure were compared across the family. The cp genomes of the only two subspecies of Asteropyrum were found to be differentiated with marked sequence variation and different inverted repeat-single copy (IR-SC) borders. The plastomes of both subspecies contains 112 genes. However, the IR region of subspecies peltatum carries 27 genes, whereas that of subspecies cavaleriei has only 25 genes. Gene inversions, transpositions, and IR expansion-contraction were very commonly detected in Ranunculaceae. The plastome of Asteropyrum has the longest IR regions in the family, but has no gene inversions or transpositions. Non-coding regions of the cp genome were not ideal markers for inferring the generic relationships of the family, but they may be applied to interpret species relationship within the genus. Plastid phylogenomic analysis using complete cp genome with Bayesian method and partitioned modeling obtained a fully resolved phylogenetic framework for Ranunculaceae. Asteropyrum was detected to be sister to Caltha, and diverged early from subfamily Ranunculoideae.

Clematismae (Ranunculaceae), a new species of C.sect.Meclatis from Xinjiang, China.

Clematismae Z.Z.Yang & L.Xie, a new species of Ranunculaceae from Xinjiang, China, is described and illustrated. The new species is morphologically similar to C.orientalis and C.glauca but can be distinguished for being a less hairy plant (hairy in C.orientalis), often 2-ternate leaves (1-2-pinnate for C.orientalis and C.glauca), lanceolate to linear-lanceolate leaflets (elliptic or ovate in C.glauca), larger flowers (smaller flower in C.orientalis) and narrowly lanceolate sepals with acute to slightly attenuate apex (narrowly oblong sepals in C.orientalis and ovate to broadly lanceolate sepals in C.glauca). The new species is endemic to the southern slope of North Tianshan Mountain in Central Xinjiang. The conservation status of the species is also discussed.

Comparative Analysis of Complete Chloroplast Genomes of Anemoclema, Anemone, Pulsatilla, and Hepatica Revealing Structural Variations Among Genera in Tribe Anemoneae (Ranunculaceae).

Structural rearrangements of Anemone species' chloroplast genome has been reported based on genetic mapping of restriction sites but has never been confirmed by genomic studies. We used a next-generation sequencing method to characterize the complete chloroplast genomes of five species in the tribe Anemoneae. Plastid genomes were assembled using de novo assembling methods combined with conventional Sanger sequencing to fill the gaps. The gene order of the chloroplast genomes of tribe Anemoneae was compared with that of other Ranunculaceae species. Multiple inversions and transpositions were detected in tribe Anemoneae. Anemoclema, Anemone, Hepatica, and Pulsatilla shared the same gene order, which contained three inversions in the large single copy region (LSC) compared to other Ranunculaceae genera. Archiclematis, Clematis, and Naravelia shared the same gene order containing two inversions and one transposition in LSC. A roughly 4.4 kb expansion region in inverted repeat (IR) regions was detected in tribe Anemoneae, suggesting that this expansion event may be a synapomorphy for this group. Plastome phylogenomic analyses using parsimony and a Bayesian method with implementation of partitioned models generated a well resolved phylogeny of Ranunculaceae. These results suggest that evaluation of chloroplast genomes may result in improved resolution of family phylogenies. Samples of Anemone, Hepatica, and Pulsatilla were tested to form paraphyletic grades within tribe Anemoneae. Anemoclema was a sister clade to Clematis. Structual variation of the plastid genome within tribe Anemoneae provided strong phylogenetic information for Ranunculaceae.