Lactobacillus plantarum LPYC225 mixture partially modulates the vaginal bacterial community of Gardnerella vaginalis-infected bacterial vaginosis in mice.

Bacterial vaginosis (BV) is defined as dysbiosis of the vaginal microbiome associated with the depletion of Lactobacilli and excessive growth of commensal or pathogenic bacteria. This study investigated the effects of lactic acid bacteria (LAB) mixture (LM; InoRexyne™) on the vaginal bacterial community of Gardnerella vaginalis (G. vaginalis)-infected BV mice. Single LAB and LM exhibited antibacterial and anti-inflammatory effects by inhibiting G. vaginalis growth and pro-inflammatory markers in RAW 264.7 cells. Administering LM did not significantly alter the vaginal architecture or fecal short-chain fatty acids but did significantly inhibit the vaginal interleukin-6 levels in the high LM group compared to the GV group. LM administration decreased the relative abundances of Enterobacter, Escherichia coli, and Bacteroides vulgatus in vaginal flushing fluids compared to the GV group. LM partially alleviated BV by inhibiting G. vaginalis growth and modulating the vaginal bacterial community, providing new insights into its modulatory effects on the vaginal microbiome in BV. Supplementary Information: The online version contains supplementary material available at 10.1007/s10068-024-01641-w.

The Possible Earliest Allopolyploidization in Tracheophytes Revealed by Phylotranscriptomics and Morphology of Selaginellaceae.

Selaginellaceae, originated in the Carboniferous and survived the Permian-Triassic mass extinction, is the largest family of lycophyte, which is sister to other tracheophytes. It stands out from tracheophytes by exhibiting extraordinary habitat diversity and lacking polyploidization. The organelle genome-based phylogenies confirmed the monophyly of Selaginella, with six or seven subgenera grouped into two superclades, but the phylogenetic positions of the enigmatic Selaginella sanguinolenta clade remained problematic. Here, we conducted a phylogenomic study on Selaginellaceae utilizing large-scale nuclear gene data from RNA-seq to elucidate the phylogeny and explore the causes of the phylogenetic incongruence of the S. sanguinolenta clade. Our phylogenetic analyses resolved three different positions of the S. sanguinolenta clade, which were supported by the sorted three nuclear gene sets, respectively. The results from the gene flow test, species network inference, and plastome-based phylogeny congruently suggested a probable hybrid origin of the S. sanguinolenta clade involving each common ancestor of the two superclades in Selaginellaceae. The hybrid hypothesis is corroborated by the evidence from rhizophore morphology and spore micromorphology. The chromosome observation and Ks distributions further suggested hybridization accompanied by polyploidization. Divergence time estimation based on independent datasets from nuclear gene sets and plastid genome data congruently inferred that allopolyploidization occurred in the Early Triassic. To our best knowledge, the allopolyploidization in the Mesozoic reported here represents the earliest record of tracheophytes. Our study revealed a unique triad of phylogenetic positions for a hybrid-originated group with comprehensive evidence and proposed a hypothesis for retaining both parental alleles through gene conversion.

Correction to: Enzyme­derived deer velvet extract activate the immune response in cyclophosphamide­induced immunosuppressive mice.

[This corrects the article DOI: 10.1007/s10068-023-01275-4.].

Evolution of the Araliaceae family involved rapid diversification of the Asian Palmate group and Hydrocotyle specific mutational pressure.

The Araliaceae contain many valuable species in medicinal and industrial aspects. We performed intensive phylogenomics using the plastid genome (plastome) and 45S nuclear ribosomal DNA sequences. A total of 66 plastome sequences were used, 13 of which were newly assembled in this study, 12 from new sequences, and one from existing data. While Araliaceae plastomes showed conserved genome structure, phylogenetic reconstructions based on four different plastome datasets revealed phylogenetic discordance within the Asian Palmate group. The divergence time estimation revealed that splits in two Araliaceae subfamilies and the clades exhibiting phylogenetic discordances in the Asian Palmate group occurred at two climatic optima, suggesting that global warming events triggered species divergence, particularly the rapid diversification of the Asian Palmate group during the Middle Miocene. Nucleotide substitution analyses indicated that the Hydrocotyloideae plastomes have undergone accelerated AT-biased mutations (C-to-T transitions) compared with the Aralioideae plastomes, and the acceleration may occur in their mitochondrial and nuclear genomes as well. This implies that members of the genus Hydrocotyle, the only aquatic plants in the Araliaceae, have experienced a distinct evolutionary history from the other species. We also discussed the intercontinental disjunction in the genus Panax and proposed a hypothesis to complement the previously proposed hypothesis. Our results provide the evolutionary trajectory of Araliaceae and advance our current understanding of the evolution of Araliaceae species.

Authentication of Allium ulleungense, A. microdictyon and A. ochotense based on super-barcoding of plastid genome and 45S nrDNA.

Allium ulleungense (AU) and A. microdictyon (AM) are valuable medicinal and edible vegetables, referred to as mountain garlic in Korea. The identification of AU, AM and a neighboring species A. ochotense (AO) is difficult because of their morphological similarities. We collected samples from three species and 46 cultivated collections to understand the genetic diversity of these valuable Allium species. Among them, we sequenced six collections, including three species and three cultivating collections to obtain data from the plastid genome (plastome) and nuclear 45S ribosomal DNA (nrDNA) for super-barcoding. The AM and AO showed around 60 single nucleotide polymorphisms (SNPs) and 39 Insertion/Deletion (InDels) in the plastome but no variations in the nrDNA sequences. Conversely, the AU and AM showed more than 170 SNPs and 80 InDels in the plastomes, and 20 SNPs and 1 InDel were found in the 45S nrDNA sequences. Among the three cultivating collections, one TB collection was determined to be the AU type in both plastome and nrDNA sequences. However, the other two collections, JB and SA, showed the AM type plastome but were heterozygous in the 45S nrDNA sequences, indicating both AU and AM types (putative AM x AU hybrid). Ten molecular markers were developed based on sequence variations to identify these three species and assess their genetic diversity. A total of 49 collections were genotyped using the ten developed markers and classified into five groups: 14 AU, 22 AM, 1 AO, 3 putative AM x AU hybrids, and 9 putative AU x AM hybrid collections. Super-barcoding with plastomes and nrDNAs revealed the genetic diversity of the three Allium species and putative hybrids between species. The newly developed markers will facilitate species and hybrid identification, thereby benefiting marker-assisted molecular breeding of Allium species.

Extensive characterization of 28 complete chloroplast genomes of Hydrangea species: A perspective view of their organization and phylogenetic and evolutionary relationships.

The tribe Hydrangeeae displays a unique, distinctive disjunct distribution encompassing East Asia, North America and Hawaii. Despite its complex trait variations and polyphyletic nature, comprehensive phylogenomic and biogeographical studies on this tribe have been lacking. To address this gap, we sequenced and characterized 28 plastomes of Hydrangeeae. Our study highlights the highly conserved nature of Hydrangeaceae chloroplast (cp) genomes in terms of gene content and arrangement. Notably, synapomorphic characteristics of tandem repeats in the conserved domain of accD were observed in the Macrophyllae, Chinenses, and Dichroa sections within the Hydrangeeae tribe. Additionally, we found lower expression of accD in these sections using structure prediction and quantitative real-time PCR analysis. Phylogenomic analyses revealed the subdivision of the Hydrangeeae tribe into two clades with robust support values. Consistent with polyphyletic relationships, sect. Broussaisia was identified as the basal group in the tribe Hydrangeeae. Our study also provides insights into the phylogenetic relationships of Hydrangea petiolaris in the Jeju and Ulleung Island populations, suggesting the need for further studies with more samples and molecular data. Divergence time estimation and biogeographical analyses suggested that the common ancestors of the tribe Hydrangeeae likely originated from North America and East Asia during the Paleocene period via the Bering Land Bridge, potentially facilitating migration within the tribe between these regions. In conclusion, this study enhances our understanding of the evolutionary history and biogeography of the tribe Hydrangeeae, shedding light on the dispersal patterns and origins of this intriguing plant group with its unique disjunct distribution.

Comparative Plastid Genome and Phylogenomic Analyses of Potamogeton Species.

Potamogetonaceae are aquatic plants divided into six genera. The largest genus in the family is Potamogeton, which is morphologically diverse with many hybrids and polyploids. Potamogetonaceae plastomes were conserved in genome size (155,863 bp-156,669 bp), gene contents (113 genes in total, comprising 79 protein-coding genes and 30 tRNA and 4 rRNA genes), and GC content (36.5%). However, we detected a duplication of the trnH gene in the IR region of the Potamogeton crispus and P. maakianus plastomes. A comparative analysis of Alismatales indicated that the plastomes of Potamogetonaceae, Cymodaceae, and Ruppiaceae have experienced a 6-kb inversion of the rbcL-trnV region and the ndh complex has been lost in the Najas flexilis plastome. Five divergent hotspots (rps16-trnQ, atpF intron, rpoB-trnC, trnC-psbM, and ndhF-rpl32) were identified among the Potamogeton plastomes, which will be useful for species identification. Phylogenetic analyses showed that the family Potamogetonaceae is a well-defined with 100% bootstrap support and divided into two different clades, Potamogeton and Stuckenia. Compared to the nucleotide substitution rates among Alismatales, we found neutral selection in all plastid genes of Potamogeton species. Our results reveal the complete plastome sequences of Potamogeton species, and will be helpful for taxonomic identification, the elucidation of phylogenetic relationships, and the plastome structural analysis of aquatic plants.

Enzyme-derived deer velvet extract activate the immune response in cyclophosphamide-induced immunosuppressive mice.

Deer velvet (DV) is an oriental traditional medicine used to treat various diseases. The present study examined the effect of flavourzyme-derived DV extract (YC-1101) on macrophages and an immunosuppressed mouse model. YC-1101 induced activation of macrophages as measured by nitric oxide production, cell proliferation, and cytokine release via concentration-dependent phosphorylation of c-Jun N-terminal kinase, extracellular signal-regulated kinase, and AKT, and nuclear translocation of p65 in macrophages. In addition, oral YC-1101 administration significantly increased splenocyte proliferation and natural killer cell activity in the immunosuppressed mouse model. Moreover, the levels of immune-related cytokines such as tumor necrotic factor-α, interferon-γ, and interleukin-2 were significantly increased by YC-1101 treatment comparable to the control group. Thus, these results suggest that YC-1101 is an efficient natural ingredient that has an immune-enhancing effect, and it might be a potential functional food for improving immunity.

The complete plastome of Cynanchum rostellatum (Apocynaceae), an indigenous plant in Korea.

The climbing plant Cynanchum rostellatum (Turcz.) Liede & Khanum is widely distributed throughout Korea and Northeast Asia as a member of the Apocynaceae family. Although this plant has a high value in medicinal and industrial purposes, genetic research on this plant is insufficient. This study announces the complete plastid genome (plastome) sequence of C. rostellatum with 663× mean coverage, which was assembled using 763 Mbp short-read data generated by the Illumina HiSeq X platform. The C. rostellatum plastome was 158,018 bp in length and displayed the typical quadripartite structure composed of the large single-copy (LSC) region (89,058 bp), the small single-copy (SSC) region (18,718 bp), and a pair of inverted repeat (IR) regions (25,116 bp). A total of 129 genes have been annotated, including 84 protein-coding genes, 37 transfer RNA genes, and eight ribosomal RNA genes. Phylogenetic analysis indicated the genus Cynanchum including 12 Cynanchum plastome sequences, was monophyletic and was located within the sub-family Asclepiadoideae. Two C. rostellatum plastomes, including the plastome assembled in this study, formed a subclade and were sister to the C. thesioides plastome, whereas the other C. rostellatum, which was previously reported one, was located within the clade of C. wilfordii and C. bungei.

Health-related quality of life using WHODAS 2.0 and associated factors 1 year after stroke in Korea: a multi-centre and cross-sectional study.

BACKGROUND: Little is known about the self-perceived level of disability of stroke survivors in the community. We aimed to characterise Health-related quality of life (HRQoL) 1 year after stroke and investigate how sociodemographic and stroke-related factors and medical adherence explain the self-perceived level of disability in a Korean stroke population. METHODS: This was a multicentre cross-sectional study. A total of 382 ischaemic stroke survivors at 1 year after onset from 11 university hospitals underwent a one-session assessment, including socioeconomic variables, the modified Rankin Scale (mRS), various neurological sequelae, the Morisky, Green and Levin-Medication Adherence Questionnaire (MGL), and the World Health Organization Disability Assessment Schedule 2.0 (WHODAS 2.0) 36-items. The relationship between disability and different variables was analysed using ordinal logistic regression. RESULTS: The prevalence of disability based on global WHODAS 2.0 was 62.6% (mild, 41.6%; moderate, 16.0%; severe, 5.0%). The prevalence of severe disability was higher in participation in society (16.8%) and getting around (11.8%) than in other domains. Low MGL- motivation was the only factor determining a significant association between all six domains of disability after adjustment. Different predictors for specific domains were age, mRS, dysarthria, trouble seeing, cognition problems, and MGL-motivation for understanding and communicating; age, recurrent stroke, mRS, hemiplegia, facial palsy, general weakness, and MGL-motivation for getting around; age, education, mRS, hemiplegia, and MGL-motivation for self-care; education, recurrent stroke, hemiplegia, dysarthria, and MGL-motivation for getting along with people; age, education, income, mRS, hemiplegia, dysarthria, MGL-knowledge, and MGL-motivation for life activities; living without a spouse, mRS, hemiplegia, dysarthria, trouble seeing, cognition problems, general weakness, and MGL-motivation for participation in society. CONCLUSIONS: Self-perceived disability according to the WHODAS 2.0 at 1 year after stroke was highly prevalent. Each disability domain showed a different prevalence and associated factors. Interventions promoting medical adherence to motivation seemed to help achieve high HRQoL in all domains.

The complete plastid genome sequence of Chloranthus fortunei (A. Gray) Solms-Laub. in Chloranthaceae.

Chloranthus fortunei (A. Gray) Solms-Laub. is a perennial herb in a basal angiosperm family Chloranthaceae. Here, we reported the complete plastid genome of C. fortunei using Illumina short-read data. The total genome size was 157,063 bp in length, containing 79 protein-coding genes, 30 tRNA genes, and four rRNA genes. The gene content and order were consistent with previously reported Chloranthus plastid genomes. The overall GC content of the C. fortunei plastid genome was 39.0%. In the phylogenetic result, genus Chloranthus was monophyletic and divided into two subclades: C. japonicus+C. angustifolius+C. fortunei, and C. henryi+C. spicatus+C. erectus. Our phylogenetic result was consistent with previous phylogenetic studies, and was supported by a previously proposed infrageneric classification of the genus Chloranthus.

Analysis of the complete plastomes and nuclear ribosomal DNAs from Euonymus hamiltonianus and its relatives sheds light on their diversity and evolution.

Euonymus hamiltonianus and its relatives (Celastraceae family) are used for ornamental and medicinal purposes. However, species identification in Euonymus is difficult due to their morphological diversity. Using plastid genome (plastome) data, we attempt to reveal phylogenetic relationship among Euonymus species and develop useful markers for molecular identification. We assembled the plastome and nuclear ribosomal DNA (nrDNA) sequences from five Euonymus lines collected from South Korea: three Euonymus hamiltonianus accessions, E. europaeus, and E. japonicus. We conducted an in-depth comparative analysis using ten plastomes, including other publicly available plastome data for this genus. The genome structures, gene contents, and gene orders were similar in all Euonymus plastomes in this study. Analysis of nucleotide diversity revealed six divergence hotspots in their plastomes. We identified 339 single nucleotide polymorphisms and 293 insertion or deletions among the four E. hamiltonianus plastomes, pointing to abundant diversity even within the same species. Among 77 commonly shared genes, 9 and 33 were identified as conserved genes in the genus Euonymus and E. hamiltonianus, respectively. Phylogenetic analysis based on plastome and nrDNA sequences revealed the overall consensus and relationships between plastomes and nrDNAs. Finally, we developed six barcoding markers and successfully applied them to 31 E. hamiltonianus lines collected from South Korea. Our findings provide the molecular basis for the classification and molecular taxonomic criteria for the genus Euonymus (at least in Korea), which should aid in more objective classification within this genus. Moreover, the newly developed markers will be useful for understanding the species delimitation of E. hamiltonianus and closely related species.

The evolution of extremely diverged plastomes in Selaginellaceae (lycophyte) is driven by repeat patterns and the underlying DNA maintenance machinery.

Two factors are proposed to account for the unusual features of organellar genomes: the disruptions of organelle-targeted DNA replication, repair, and recombination (DNA-RRR) systems in the nuclear genome and repetitive elements in organellar genomes. Little is known about how these factors affect organellar genome evolution. The deep-branching vascular plant family Selaginellaceae is known to have a deficient DNA-RRR system and convergently evolved organellar genomes. However, we found that the plastid genome (plastome) of Selaginella sinensis has extremely accelerated substitution rates, a low GC content, pervasive repeat elements, a dynamic network structure, and it lacks direct or inverted repeats. Unexpectedly, its organelle DNA-RRR system is short of a plastid-targeted Recombinase A1 (RecA1) and a mitochondrion-targeted RecA3, in line with other explored Selaginella species. The plastome contains a large collection of short- and medium-sized repeats. Given the absence of RecA1 surveillance, we propose that these repeats trigger illegitimate recombination, accelerated mutation rates, and structural instability. The correlations between repeat quantity and architectural complexity in the Selaginella plastomes support these conclusions. We, therefore, hypothesize that the interplay of the deficient DNA-RRR system and the high repeat content has led to the extraordinary divergence of the S. sinensis plastome. Our study not only sheds new light on the mechanism of plastome divergence by emphasizing the power of cytonuclear integration, but it also reconciles the longstanding contradiction on the effects of DNA-RRR system disruption on genome structure evolution.

The complete plastid genome sequence of Peucedanum hakuunense Nakai (Apiaceae), an endemic and rare species in Korea.

Peucedanum hakuunense Nakai is one of the rare species in the Korean Peninsula. This study characterized the complete plastid genome (plastome) sequence of P. hakuunense by de novo assembly with next-generation sequencing data. The complete plastome of P. hakuunense is 147,426 bp in length with a typical quadripartite structure comprising a large single-copy region of 91,915 bp, a small single-copy region of 17,425 bp, and two inverted repeat regions of 19,043 bp in length. The plastome of P. hakuunense is composed of 85 protein-coding genes, 36 tRNA genes, and 8 rRNA genes. The phylogenetic analysis revealed that two Peucedanum species formed an independent subclade, sister to the subclade of Angelica species within the tribe Selineae.

Distinctive evolutionary pattern of organelle genomes linked to the nuclear genome in Selaginellaceae.

Plastids and mitochondria are endosymbiotic organelles that store genetic information. The genomes of these organelles generally exhibit contrasting patterns regarding genome architecture and genetic content. However, they have similar genetic features in Selaginellaceae, and little is known about what causes parallel evolution. Here, we document the multipartite plastid genomes (plastomes) and the highly divergent mitochondrial genomes (mitogenomes) from spikemoss obtained by combining short- and long-reads. The 188-kb multipartite plastome has three ribosomal operon copies in the master genomic conformation, creating the alternative subgenomic conformation composed of 110- and 78-kb subgenomes. The long-read data indicated that the two different genomic conformations were present in almost equal proportions in the plastomes of Selaginella nipponica. The mitogenome of S. nipponica was assembled into 27 contigs with a total size of 110 kb. All contigs contained directly arranged repeats at both ends, which introduced multiple conformations. Our results showed that plastomes and mitogenomes share high tRNA losses, GC-biased nucleotides, elevated substitution rates and complicated organization. The exploration of nuclear-encoded organelle DNA replication, recombination and repair proteins indicated that, several single-targeted proteins, particularly plastid-targeted recombinase A1, have been lost in Selaginellaceae; conversely, the dual-targeted proteins remain intact. According to the reported function of recombinase A1, we propose that the plastomes of spikemoss often fail to pair homologous sequences during recombination, and the dual-targeted proteins play a key role in the convergent genetic features of plastomes and mitogenomes. Our results provide a distinctive evolutionary pattern of the organelle genomes in Selaginellaceae and evidence of their convergent evolution.

Hospital Volume Threshold Associated with Higher Survival after Endovascular Recanalization Therapy for Acute Ischemic Stroke.

BACKGROUND AND PURPOSE: Endovascular recanalization therapy (ERT) is becoming increasingly important in the management of acute ischemic stroke (AIS). However, the hospital volume threshold for optimal ERT remains unknown. We investigated the relationship between hospital volume of ERT and risk-adjusted patient outcomes. METHODS: From the National Health Insurance claims data in Korea, 11,745 patients with AIS who underwent ERT from July 2011 to June 2016 in 111 hospitals were selected. We measured the hospital's ERT volume and patient outcomes, including the 30-day mortality, readmission, and postprocedural intracranial hemorrhage (ICH) rates. For each outcome measure, we constructed risk-adjusted prediction models incorporating demographic variables, the modified Charlson comorbidity index, and the stroke severity index (SSI), and validated them. Risk-adjusted outcomes of AIS cases were compared across hospital quartiles to confirm the volume-outcome relationship (VOR) in ERT. Spline regression was performed to determine the volume threshold. RESULTS: The mean AIS volume was 14.8 cases per hospital/year and the unadjusted means of mortality, readmission, and ICH rates were 11.6%, 4.6%, and 8.6%, respectively. The VOR was observed in the risk-adjusted 30-day mortality rate across all quartile groups, and in the ICH rate between the first and fourth quartiles (P<0.05). The volume threshold was 24 cases per year. CONCLUSIONS: There was an association between hospital volume and outcomes, and the volume threshold in ERT was identified. Policies should be developed to ensure the implementation of the AIS volume threshold for hospitals performing ERT.

Intracranial Venous Reflux Caused by Occlusion of the Brachiocephalic Vein Mimicking Dural Arteriovenous Fistula.

BACKGROUND: High signal intensity of cerebral venous sinuses on magnetic resonance angiography (MRA) indicates high flow of shunt from arterial blood, suggesting dural arteriovenous fistula (dAVF). Herein, we describe the cases of 2 patients with intracranial venous reflux caused by brachiocephalic vein occlusion (BVO) mimicking dAVF on MRA. CASE DESCRIPTION: Case 1 was a 71-year-old woman who had received arteriovenous graft (AVG) surgery for hemodialysis and presented with headache. MRA depicted high signal intensities in the left jugular vein, inferior petrosal sinus, and ophthalmic vein suggesting intracranial dAVF. However, cerebral angiography revealed intracranial venous reflux through the jugular vein caused by BVO. After balloon angioplasty with stenting for BVO, normal venous drainage was restored and her symptoms resolved. Case 2 was a 62-year-old woman who had received AVG for hemodialysis and was hospitalized due to seizure. MRA revealed high signal intensities of the left transverse-sigmoid sinus and jugular vein, and it was suspected that they were associated with BVO. She did not receive immediate intervention because of her septic condition. The patient ultimately died of fatal intracranial hemorrhage and subarachnoid hemorrhage. CONCLUSIONS: Central venous occlusion may occur in patients with end-stage renal disease who are receiving AVG for hemodialysis. Intracranial venous reflux resulting from BVO can induce various neurologic disorders including intracranial venous hypertension or hemorrhage. Prompt recognition and implementation of endovascular treatment can be beneficial in cases of symptomatic central venous occlusion.

Effects of Carotid Calcification on Restenosis After Carotid Artery Stenting: A Follow-Up Study with Computed Tomography Angiography.

BACKGROUND: Carotid Doppler sonography has been used routinely in the screening for restenosis after carotid artery stenting (CAS) by applying the criteria of flow velocity; however, computed tomography angiography (CTA) enables us to detect restenosis directly and has some advantages that help to overcome the limitations of Carotid Doppler sonography. We aimed to investigate the incidence of restenosis with CTA and to identify factors related to restenosis after CAS. METHODS: We conducted a retrospective study of 225 patients (235 arteries, mean age of 71.9 ± 7.7 years) who had undergone CTA of the neck for the evaluation of restenosis after CAS between July 2002 and June 2012. We assessed the cumulative incidence of restenosis (>50%) and the predictors of restenosis after CAS. RESULTS: During 1064.4 patient-years of follow-up (mean 4.5 years; range 1-9.7 years), the incidence of restenosis was 10.2% (24/235). The cumulative rate of freedom from restenosis was 94.1%, 88.8%, and 86.7% at 1, 3, and 5 years, respectively. The statistically significant factors associated with restenosis after CAS were residual stenosis (adjusted odds ratio 1.07, 95% confidence interval 1.03-1.11, P < 0.01) and high-grade (>50%) calcification (adjusted odds ratio 6.04, 95% confidence interval 1.29-28.36, P = 0.02) on multivariate analysis. CONCLUSIONS: This study revealed that carotid arteries with high-grade calcification were prone to develop restenosis after CAS. Regular follow-up should be performed for the monitoring of restenosis after CAS in patients with highly calcified carotid plaques.

Plastid Phylogenomics Resolve Deep Relationships among Eupolypod II Ferns with Rapid Radiation and Rate Heterogeneity.

The eupolypods II ferns represent a classic case of evolutionary radiation and, simultaneously, exhibit high substitution rate heterogeneity. These factors have been proposed to contribute to the contentious resolutions among clades within this fern group in multilocus phylogenetic studies. We investigated the deep phylogenetic relationships of eupolypod II ferns by sampling all major families and using 40 plastid genomes, or plastomes, of which 33 were newly sequenced with next-generation sequencing technology. We performed model-based analyses to evaluate the diversity of molecular evolutionary rates for these ferns. Our plastome data, with more than 26,000 informative characters, yielded good resolution for deep relationships within eupolypods II and unambiguously clarified the position of Rhachidosoraceae and the monophyly of Athyriaceae. Results of rate heterogeneity analysis revealed approximately 33 significant rate shifts in eupolypod II ferns, with the most heterogeneous rates (both accelerations and decelerations) occurring in two phylogenetically difficult lineages, that is, the Rhachidosoraceae-Aspleniaceae and Athyriaceae clades. These observations support the hypothesis that rate heterogeneity has previously constrained the deep phylogenetic resolution in eupolypods II. According to the plastome data, we propose that 14 chloroplast markers are particularly phylogenetically informative for eupolypods II both at the familial and generic levels. Our study demonstrates the power of a character-rich plastome data set and high-throughput sequencing for resolving the recalcitrant lineages, which have undergone rapid evolutionary radiation and dramatic changes in substitution rates.