Comparative Analysis of Six Complete Plastomes of Tripterospermum spp.

The Tripterospermum, comprising 34 species, is a genus of Gentianaceae. Members of Tripterospermum are mostly perennial, entwined herbs with high medicinal value and rich in iridoids, xanthones, flavonoids, and triterpenes. However, our inadequate understanding of the differences in the plastid genome sequences of Tripterospermum species has severely hindered the study of their evolution and phylogeny. Therefore, we first analyzed the 86 Gentianae plastid genomes to explore the phylogenetic relationships within the Gentianae subfamily where Tripterospermum is located. Then, we analyzed six plastid genomes of Tripterospermum, including two newly sequenced plastid genomes and four previously published plastid genomes, to explore the plastid genomes' evolution and phylogenetic relationships in the genus Tripterospermum. The Tripterospermum plastomes have a quadripartite structure and are between 150,929 and 151,350 bp in size. The plastomes of Tripterospermum encoding 134 genes were detected, including 86 protein-coding genes (CDS), 37 transfer RNA (tRNA) genes, eight ribosomal RNA (rRNA) genes, and three pseudogenes (infA, rps19, and ycf1). The result of the comparison shows that the Tripterospermum plastomes are very conserved, with the total plastome GC content ranging from 37.70% to 37.79%. In repeat sequence analysis, the number of single nucleotide repeats (A/T) varies among the six Tripterospermum species, and the identified main long repeat types are forward and palindromic repeats. The degree of conservation is higher at the SC/IR boundary. The regions with the highest divergence in the CDS and the intergenic region (IGS) are psaI and rrn4.5-rrn5, respectively. The average pi of the CDS and the IGS are only 0.071% and 0.232%, respectively, indicating that the Tripterospermum plastomes are highly conserved. Phylogenetic analysis indicated that Gentianinae is divided into two clades, with Tripterospermum as a sister to Sinogeniana. Phylogenetic trees based on CDS and CDS + IGS combined matrices have strong support in Tripterospermum. These findings contribute to the elucidation of the plastid genome evolution of Tripterospermum and provide a foundation for further exploration and resource utilization within this genus.

The molecular population structure of Swertia perennis (Gentianaceae) in Central Europe.

Phylogeographic analysis of Swertia perennis, a typical European subalpine springtime species, revealed the existence of at least five major phylogenetic lineages. A large phylogeographic separation exists among these geographical regions, which confirms the existence of glacial refugia in the Pyrenees, but also in the Eastern and Central Alps. The results obtained from the analyzes indicate the existence of three major differences between the populations of the Alps and the Pyrenees, but also between the populations of the Alps and those of other geographical regions (Carpathians, southern Czech Republic, Sudetes and northern Poland). The studied populations from the Black Forest and from north-eastern and southern Poland are a relic of the former wider distribution of these (periglacial) genetic groups within Swertia perennis. Our results also confirm the existence of biogeographical links between the Carpathians and the Hercynian Range and the Alps. Certainly, there was an exchange of genes between populations located in the eastern Alps, the Carpathians and the Hercynian ranges (Czech Republic, Jeseníky, Sudetes, Ore Mountains). This confirms previous results of comparative studies on the genetic diversity of populations of other vascular plant species.

Phylogeography and ecological niche modeling implicate multiple microrefugia of Swertia tetraptera during quaternary glaciations.

BACKGROUND: Climate fluctuations during the Pleistocene and mountain uplift are vital driving forces affecting geographic distribution. Here, we ask how an annual plant responded to the Pleistocene glacial cycles. METHODS: In this study, we analyzed the population demographic history of the annual herb Swertia tetraptera Maxim (Gentianaceae) endemic to Qinghai-Tibetan Plateau (QTP). A total of 301 individuals from 35 populations of S. tetraptera were analyzed based on two maternally inherited chloroplast fragments (trnL-trnF and trnS-trnG). Phylogeographic analysis was combined with species distribution modeling to detect the genetic variations in S. tetraptera. RESULTS: The genetic diversity of S. tetraptera was high, likely due to its wide natural range, high proportion of endemic haplotypes and evolutionary history. Fifty-four haplotypes were identified in S. tetraptera. Only a few haplotypes were widespread (Hap_4, Hap_1, Hap_3), which were dispersed throughout the present geographical range of S. tetraptera, while many haplotypes were confined to single populations. The cpDNA dataset showed that phylogeographic structuring was lacking across the distribution range of S. tetraptera. Analyses of molecular variance showed that most genetic variation was found within populations (70.51%). In addition, the relationships of the haplotypes were almost completely unresolved by phylogenetic reconstruction. Both mismatch distribution analysis and neutrality tests showed a recent expansion across the distribution range of S. tetraptera. The MaxEnt analysis showed that S. tetraptera had a narrow distribution range during the Last Glacial Maximum (LGM) and a wide distribution range during the current time, with predictions into the future showing the distribution range of S. tetraptera expanding. CONCLUSION: Our study implies that the current geographic and genetic distribution of S. tetraptera is likely to have been shaped by Quaternary periods. Multiple microrefugia of S. tetraptera existed during Quaternary glaciations. Rapid intraspecific diversification and hybridization and/or introgression may have played a vital role in shaping the current distribution patterns of S. tetraptera. The distribution range of S. tetraptera appeared to have experienced contraction during the LGM; in the future, when the global climate becomes warmer with rising carbon dioxide levels, the distribution of S. tetraptera will expand.

Impact of mycoheterotrophy on the growth of Gentiana zollingeri (Gentianaceae), as suggested by size variation, morphology, and 13C abundance of flowering shoots.

Gentiana zollingeri is an annual photosynthetic plant that employs a mycoheterotrophic growth strategy during its underground seedling stage (initial mycoheterotrophy). Notably, the morphological characteristics of its flowering shoots, such as shoot size, leaf size, and leaf color, are highly variable, and it was hypothesized that these variations may be linked to nutritional mode. The morphological characteristics of G. zollingeri individuals were thus investigated alongside environmental factors, 13C abundance, and diversity of colonizing arbuscular mycorrhizal (AM) fungi. The majority of G. zollingeri flowering individuals were found to exhibit a high affinity for the specific AM fungi that exclusively colonize roots of the mycoheterotrophic seedlings, while other phylogenetically diverse AM fungi could also be detected. The leaves to shoot dry weight ratio (leaf ratio) was negatively correlated with the canopy openness in the habitat, suggesting that leaf development is impeded in sunny conditions. Furthermore, the shoot weight of G. zollingeri was positively correlated with leaf 13C abundance. Given that 13C enrichment can provide indirect evidence of mycoheterotrophy in AM plants, the results suggest that the utilization of carbon obtained through mycoheterotrophy, at least during the underground seedling stage, is crucial for G. zollingeri.

Association of mycoheterotrophic Gentianaceae with specific Glomus lineages.

Some plant species took an alternative evolutionary pathway in which they lost their photosynthetic capacity to depend exclusively on carbon supplied by arbuscular mycorrhizal fungi (AMF) in an association called mycoheterotrophy. Among them is Voyriella parviflora, a species of the family Gentianaceae, which is found in tropical regions such as the Amazon basin. Here, we assessed the identity of AMF symbionts associated with this species. DNA was isolated from eight Gentianaceae specimens and from litter and surrounding roots of photosynthetic plants. The atp1 gene was amplified by Sanger sequencing to determine the taxonomic affiliation of the mycoheterotrophic plants. A 280 bp region of the 18S rRNA gene of AMF was amplified with primers NS31/AML2 by high-throughput sequencing. The mycoheterotrophic specimens were assigned to V. parviflora with a bootstrap support of 72%. Glomus was the most abundant AMF genus, both in the mycoheterotrophic plants and in the litter and roots of photosynthetic plants. In addition, a few Glomus genotypes were abundantly enriched in the mycoheterotrophic plants, with only a few specimens colonized by Gigaspora, Acaulospora, and Scutellospora in a low proportion. These genotypes formed a cluster within a larger clade, suggesting that V. parviflora shows a preferential association with a narrow Glomus lineage which is not phylogenetically close to a previously identified V. parviflora's associated lineage. Furthermore, detecting fungi from other families suggests that V. parviflora is colonized by other genera, although with low frequency. These findings provide new insights into the association between AMF and mycoheterotrophic species and highlight the importance of considering trap culture-independent approaches in understanding this symbiosis.

Acylated iridoid glucoside and xanthones from Canscora lucidissima: Their structures and ferroptosis inhibitory activity.

Phytochemical study on the whole plants of a Gentianaceous medicinal plant, Canscora lucidissima, gave one new acylated iridoid glucoside, canscorin A (1), and two new xanthone glycosides (2 and 3) together with 17 known compounds including five xanthones, eight xanthone glycosides, two benzophenone glucosides, caffeic acid, and loganic acid. Canscorin A (1) was assigned as a loganic acid derivative having a hydroxyterephthalic acid moiety by spectroscopic analysis together with chemical evidence, while 2 and 3 were elucidated to be a rutinosylxanthone and a glucosylxanthone, respectively. The absolute configurations of the sugar moieties of 2 and 3 were determined by HPLC analysis. The isolated compounds were evaluated for their inhibitory activities against erastin-induced ferroptosis on human hepatoma Hep3B cells and LPS-stimulated IL-1ß production from murine microglial cells.

Comparative chloroplast genomics of 34 species in subtribe Swertiinae (Gentianaceae) with implications for its phylogeny.

BACKGROUND: Subtribe Swertiinae, a medicinally significant and highly speciose Subtribe of family Gentianaceae. Despite previous extensive studies based on both morphology and molecular data, intergeneric and infrageneric relationships within subtribe Swertiinae remain controversial. METHODS: Here, we employed four newly generated Swertia chloroplast genomes with thirty other published genomes to elucidate their genomic characteristics. RESULTS: The 34 chloroplast genomes were small and ranged in size from 149,036 to 154,365 bp, each comprising two inverted repeat regions (size range 25,069-26,126 bp) that separated large single-copy (80,432-84,153 bp) and small single-copy (17,887-18,47 bp) regions, and all the chloroplast genomes showed similar gene orders, contents, and structures. These chloroplast genomes contained 129-134 genes each, including 84-89 protein-coding genes, 37 tRNAs, and 8 rRNAs. The chloroplast genomes of subtribe Swertiinae appeared to have lost some genes, such as rpl33, rpl2 and ycf15 genes. Comparative analyses revealed that two mutation hotspot regions (accD-psaI and ycf1) could serve as effective molecular markers for further phylogenetic analyses and species identification in subtribe Swertiinae. Positive selection analyses showed that two genes (ccsA and psbB) had high Ka/Ks ratios, indicating that chloroplast genes may have undergone positive selection in their evolutionary history. Phylogenetic analysis showed that the 34 subtribe Swertiinae species formed a monophyletic clade, with Veratrilla, Gentianopsis and Pterygocalyx located at the base of the phylogenetic tree. Some genera of this subtribe, however, were not monophyletic, including Swertia, Gentianopsis, Lomatogonium, Halenia, Veratrilla and Gentianopsis. In addition, our molecular phylogeny was consistent with taxonomic classification of subtribe Swertiinae in the Roate group and Tubular group. The results of molecular dating showed that the divergence between subtrib Gentianinae and subtrib Swertiinae was estimated to occur in 33.68 Ma. Roate group and Tubular group in subtribe Swertiinae approximately diverged in 25.17 Ma. CONCLUSION: Overall, our study highlighted the taxonomic utility of chloroplast genomes in subtribe Swertiinae, and the genetic markers identified here will facilitate future studies on the evolution, conservation, population genetics, and phylogeography of subtribe Swertiinae species.

Phylotranscriptomics of Swertiinae (Gentianaceae) reveals that key floral traits are not phylogenetically correlated.

Establishing how lineages with similar traits are phylogenetically related remains critical for understanding the origin of biodiversity on Earth. Floral traits in plants are widely used to explore phylogenetic relationships and to delineate taxonomic groups. The subtribe Swertiinae (Gentianaceae) comprises more than 350 species with high floral diversity ranging from rotate to tubular corollas and possessing diverse nectaries. Here we performed phylogenetic analysis of 60 species from all 15 genera of the subtribe Swertiinae sensu Ho and Liu, representing the range of floral diversity, using data from the nuclear and plastid genomes. Extensive topological conflicts were present between the nuclear and plastome trees. Three of the 15 genera represented by multiple species are polyphyletic in both trees. Key floral traits including corolla type, absence or presence of lobe scales, nectary type, nectary position, and stigma type are randomly distributed in the nuclear and plastome trees without phylogenetic correlation. We also revealed the likely ancient hybrid origin of one large clade comprising 10 genera with diverse floral traits. These results highlight the complex evolutionary history of this subtribe. The phylogenies constructed here provide a basic framework for further exploring the ecological and genetic mechanisms underlying both species diversification and floral diversity.

Comparative analysis of mitochondrial genomes of two alpine medicinal plants of Gentiana (Gentianaceae).

Gentiana crassicaulis and G. straminea are alpine plants of Gentiana with important medicinal value and complex genetic backgrounds. In this study, the mitochondrial genomes (mtDNAs) of these two species were sequenced. The mtDNAs of G. crassicaulis and G. straminea are 368,808 and 410,086 bp long, respectively, 52 and 49 unique genes are annotated in the two species, and the gene arrangement varies widely. Compared to G. crassicaulis, G. straminea loses three effective genes, namely atp6, trnG-GCC and trnV-GAC. As a pseudogene, the atp6 gene of G. straminea is incomplete, which is rare in higher plants. We detected 1696 and 1858 pairs of long repeats and 213 SSRs and 250 SSs in the mtDNAs of G. crassicaulis and G. straminea, respectively. There are 392 SNPs and 18 InDels between the two genomes, and syntenic sequence and structural variation analysis show low collinearity between the two genomes. Chloroplast DNA transferring to mtDNA is observed in both species, and 46,511 and 55,043 bp transferred segments containing three tRNA genes are identified, respectively. Comparative analysis of mtDNAs of G. crassicaulis, G. straminea and four species of Gentianales determined 18 core genes, and there is no specific gene in G. crassicaulis and G. straminea. The phylogenetic tree based on mtDNAs places Gentianaceae in a branch of Gentianales. This study is the first to analyze the mtDNAs of Gentianaceae, which could provide information for analysis of the structure of mtDNAs of higher plants and phylogenetic research of Gentianaceae and Gentianales.

Unraveling the secretion mechanism of the curious nectaries in Gentianaceae.

Unusual nectaries were anatomically described as being usual traits for Gentianaceae. They are small, avascularized, and formed by 3 to 5 rosette cells with labyrinthine walls around one central cell. Such as nectaries have been reported for stems, petals, and sepals of different species of the family, however, there is no information on the mechanisms involved with the synthesis and release of secretion. Thus, this work aimed to unravel the mechanism of secretion and exudation of nectar for these curious nectaries using Calolisianthus speciosus as a model. Samples were processed according to standard methods for light and electron microscopy. Leaf and sepal nectaries were described, as were those of the apex of petals where ants were observed patrolling a darkened region. The enzymatic method was used for the detection of sugars, proteins, and amino acids in leaf and sepal exudates. The nectaries of petals of C. speciosus are similar to those of its leaves, sepals, and stem, although their activities are asynchronous. Polysaccharides were detected on the labyrinthine walls of rosette cells and protein in the opposite region of the cytoplasm. Labyrinthine walls increase the contact surface between rosette cells and the central cell, allowing for the transfer of secretion. After accumulation, the secretion is released to the subcuticular space of the central cell through disruption of the cuticle. The secretion and exudation of nectar were elucidated and involve distinct organelles.

Isolation and cytotoxic activities of undescribed iridoid and xanthone glycosides from Centaurium erythraea Rafn. (Gentianaceae).

Centaurium erythraea Rafn. (Gentianaceae) is used in internal traditional therapy as an anthelmintic, hypotensive, antipyretic, and antidiabetic. It is used externally for the treatment of wounds. Ursolic acid, maslinic acid, secologanin, secologanin dimethyl acetal, centauroside A, erythraeaxanthone I, erythraeaxanthone II, and demethyleustomin were isolated from aerial parts of Centaurium erythraea and were identified using spectroscopic methods, including NMR and mass spectrometry. The cytotoxic potency of undescribed compounds was evaluated by the XTT assay against human breast cancer MCF-7, MDA-MB-453 and mouse fibroblast 3T3-L1 cell lines. Erythraeaxanthone II was found to have the most potent cytotoxic activity.

Secondary metabolites from the Mongolian medicine Lomatogonium carinthiacum.

Systematic phytochemical investigation on the Mongolian medicinal herb Lomatogonium carinthiacum led to the isolation of 12 monoterpenoids including three new secoiridoids (1, 2 and 4) and one new iridoid glycoside (13), one new monoterpenoid alkaloid (3), and three new sesquiterpenoids (14-16). Comprehensive spectroscopic analysis (including 1D and 2D NMR, and HRESIMS) and quantum chemistry computations (including ECD and NMR calculations) were applied to elucidate their structures. Weak immunosuppressive activities were observed for the new isolates via inhibiting T cell proliferation and cytokine IFN-γ secretion in vitro.

Chromosome-scale genome assembly of Eustoma grandiflorum, the first complete genome sequence in the genus Eustoma.

Eustoma grandiflorum (Raf.) Shinn. is an annual herbaceous plant native to the southern United States, Mexico, and the Greater Antilles. It has a large flower with a variety of colors and is an important flower crop. In this study, we established a chromosome-scale de novo assembly of E. grandiflorum genome sequences by integrating four genomic and genetic approaches: (1) Pacific Biosciences (PacBio) Sequel deep sequencing, (2) error correction of the assembly by Illumina short reads, (3) scaffolding by chromatin conformation capture sequencing (Hi-C), and (4) genetic linkage maps derived from an F2 mapping population. Thirty-six pseudomolecules and 64 unplaced scaffolds were created, with a total length of 1,324.8 Mb. A total of 36,619 genes were predicted on the genome as high-confidence genes. A comparison of genome structure between E. grandiflorum and C. canephora or O. pumila suggested whole-genome duplication after the divergence between the families Gentianaceae and Rubiaceae. Phylogenetic analysis with single-copy genes suggested that the divergence time between Gentianaceae and Rubiaceae was 74.94 MYA. Genetic diversity analysis was performed for nine commercial E. grandiflorum varieties bred in Japan, from which 254,205 variants were identified. This first report on the construction of a reference genome sequence in the genus Eustoma is expected to contribute to genetic and genomic studies in this genus and in the family Gentianaceae.

Effect of novel therapeutic medicine swertiamarin from Enicostema axillare in zebrafish infected with Salmonella typhi.

Herbal treatments have been practiced by humans over centuries and therefore possess time-proven safety. However, it is crucial to evaluate the toxic effects of herbal medicine to confirm their safety, particularly when developing therapeutic drugs. Use of laboratory animals such as mice, rat, and rabbits was considered as gold standard in herbal toxicity assessments. However, in the last few decades, the ethical consideration of using higher vertebrates for toxicity testing has become more controversial. As a possible alternative model involving lower vertebrates such as zebra fish were introduced. Hence in the present study, swertiamain compound isolated from E. axillare was assessed for it antimicrobial activity in zebra fish larvae againt S. typhi. The cumulative mortality rate and bacterial localization in zebra fish larvae were studied. Biochemical markers assays were performed to find the preventive role of the compound during the typhoid infection. The results showed that zebra fish can be successfully used as a model to study typhoid infection and the anti-bacterial compound swertiamarin used in this study clears the bacterial load and pathogenic symptoms to a great extent.

Complete chloroplast genomes of two medicinal Swertia species: the comparative evolutionary analysis of Swertia genus in the Gentianaceae family.

MAIN CONCLUSION: The complete chloroplast genome of Swertia kouitchensis has been sequenced and assembled, compared with that of S. bimaculata to determine the evolutionary relationships among species of the Swertia in the Gentianaceae family. Swertia kouitchensis and S. bimaculata are from the Gentianaceae family. The complete chloroplast genome of S. kouitchensis was newly assembled, annotated, and analyzed by Illumina Hiseq 2500 platform. The chloroplast genomes of the two species encoded a total of 133, 134 genes, which included 88-89 protein-coding genes, 37 transfer RNA (tRNA) genes, and 8 ribosomal RNA genes. One intron was contained in each of the eight protein-coding genes and eight tRNA-coding genes, whereas two introns were found in two genes (ycf3 and clpP). The most abundant codon of the two species was for isoleucine, and the least abundant codon was for cysteine. The number of microsatellite repeat sequences was twenty-eight and thirty-two identified in the chloroplast genomes of S. kouitchensis and S. bimaculata, respectively. A total of 1127 repeat sequences were identified in all the 23 Swertia chloroplast genomes, and they fell into four categories. Furthermore, five divergence hotspot regions can be applied to discriminate these 23 Swertia species through genomes comparison. One pair of genus-specific DNA barcodes primer has been accurately identified. Therefore, the diverse regions cloned by a specific primer may become an effective and powerful molecular marker for the identification of Swertia genus. Moreover, four genes (ccsA, ndhK, rpoC1, and rps12) were positive selective pressure. The phylogenetic tree showed that the 23 Swertia species were clustered into a large clade including four evident subbranches, whereas the two species of S. kouitchensis and S. bimaculata were separately clustered into the diverse but correlated species group.

Complete chloroplast genome of Gentianopsis barbata and comparative analysis with related species from Gentianaceae.

Gentianopsis barbata is an essential medicinal plant in China with high ornamental and medicinal values. Unfortunately, the study of the chloroplast genome of this plant still has a gap. This study sequenced and characterized the complete chloroplast genome of G. barbata. The complete chloroplast genome of G. barbata is a typical circular structure of 151 123 bp. It consists of a large single-copy region (82 690 bp) and a small single-copy region (17 887 bp) separated by a pair of inverted repeats (25 273 bp), which covers 78 protein-coding genes, 30 tRNAs, and 4 rRNAs. The long repeat sequence analysis showed that the P-type (palindromic) sequences were the major long repeat sequences. Thirty-seven simple sequence repeats were identified, most of which were single nucleotides. The Bayesian inference tree, maximum likelihood tree, and neighbor-joining tree suggested that G. barbata is grouped with Gentianopsis grandis and Gentianopsis paludosa. The divergence time analysis showed that G. barbata diverged at 1.243 Mya. Comparative analysis of chloroplast genomes can reveal interspecific diversity, and regions with high variation can be used to develop molecular markers applicable to various research areas. Our results provide a new insight into plastome evolution and a valuable resource for further studies on G. barbata.

Plastome structure, phylogenomics and evolution of plastid genes in Swertia (Gentianaceae) in the Qing-Tibetan Plateau.

BACKGROUND: The genus Swertia is of great medicinal importance and one of the most taxonomically challenging taxa within Gentianaceae, largely due to the morphological similarities of species within this genus and with its closely related genera. Previous molecular studies confirmed its polyphyly but suffered from low phylogenetic resolutions because only limited sequence loci were used. Thus, we conducted the structural, gene evolutionary, and phylogenetic analyses of 11 newly obtained plastomes of Swertia. Our result greatly improved the phylogenetic resolutions in Swertia, shed new light on the plastome evolution and phylogenetic relationships of this genus. RESULTS: The 11 Swertia plastomes together with the published seven species proved highly similar in overall size, structure, gene order, and content, but revealed some structural variations caused by the expansion and contraction of the IRb region into the LSC region, due to the heterogeneous length of the ψycf1. The gene rps16 was found to be in a state flux with pseudogenes or completely lost. Similar situation was also documented in other genera of Gentianaceae. This might imply loss of the gene in the common ancestor of Gentianaceae. The distribution plot of ENC vs. GC3 showed all these plastomes arranging very close in the Wright line with an expected ENC value (49-52%), suggesting the codon usage of Swertia was mainly constrained by a GC mutation bias. Most of the genes remained under the purifying selection, however, the cemA was identified under positive selection, possibly reflecting an adaptive response to low CO2 atmospheric conditions during the Late Miocene. Our phylogenomic analyses, based on 74 protein-coding genes (CDS), supported the polyphyly of Swertia with its close allies in the subtribe Swertiinae, presumably due to recent rapid radiation. The topology inferred from our phylogenetic analyses partly supported the current taxonomic treatment. Finally, several highly variable loci were identified, which can be used in future phylogenetic studies and accurate identification of medicinal genuineness of Swertia. CONCLUSIONS: Our study confirmed the polyphyly of Swertia and demonstrated the power of plastome phylogenomics in improvement of phylogenetic resolution, thus contributing to a better understanding of the evolutionary history of Swertia.

The chromosome-level genome assembly of Gentiana dahurica (Gentianaceae) provides insights into gentiopicroside biosynthesis.

Gentiana dahurica Fisch. is a perennial herb of the family Gentianaceae. This species is used as a traditional Tibetan medicine because of its rich gentiopicroside constituents. Here, we generate a high-quality, chromosome-level genome of G. dahurica with a total length of 1,416.54 Mb. Comparative genomic analyses showed that G. dahurica shared one whole-genome duplication (WGD) event with Gelsemium sempervirens of the family Gelsemiaceaei and had one additional species-specific WGD after the ancient whole-genome triplication with other eudicots. Further transcriptome analyses identified numerous enzyme coding genes and the transcription factors related to gentiopicroside biosynthesis. A set of candidate cytochrome P450 genes were identified for being involved in biosynthetic shifts from swertiamarin to gentiopicroside. Both gene expressions and the contents measured by high-performance liquid chromatography indicated that the gentiopicrosides were mainly synthesized in the rhizomes with the highest contents. In addition, we found that two above-mentioned WGDs, contributed greatly to the identified candidate genes involving in gentiopicroside biosynthesis. The first reference genome of Gentianaceae we generated here will definitely accelerate evolutionary, ecological, and pharmaceutical studies of this family.

Effects of HIIT associated with Coutoubea spicata supplementation on tissue and oxidative damage biomarkers in Wistar rats / Efectos del HIIT asociado a la suplementación de Coutoubea spicata en los biomarcadores del tejido y daño oxidativo en ratas Wistar

SUMMARY: This study aimed to evaluate the effects of six weeks of HIIT on tissue and oxidative damage markers in rats supplemented with Coutoubea spicata fraction. Thirty-two male Wistar rats were divided into 4 groups: Baseline (GB); supplemented with 100 mg/kg of Coutoubea spicata fraction (GSCS); exercised for 6 weeks with the HIIT protocol (GH); supplemented with 100 mg/kg of Coutoubea spicata fraction + HIIT for 6 weeks (GHCS). Exercised animals performed the HIIT protocol (2 x 2). Tissue damage CK, LDH, ALT and AST markers in plasma were analyzed, as well as oxidative stress MDA and SH biomarkers in plasma and in cardiac, hepatic and muscle tissues. The results showed that CK, LDH, AST and ALT enzymes showed increase in GH when compared to GB (p<0.0001). However, CK, AST and ALT markers reduced their concentrations in GHCS when compared to GH (p<0.0001), indicating that Coutoubea spicata supplementation attenuated the damage in muscle and liver tissues induced by HIIT. Plasma, liver and muscle MDA showed increase in GH after HIIT sessions; however, when compared to GHCS, it showed reduced levels (p<0.0001). SH was elevated in the GH group when compared to GB in plasma and liver tissues (p<0.0001); in contrast, reduction in GHCS when compared to GH was observed in plasma, liver and cardiac tissues, demonstrating the redox effect of HIIT on some tissues. Thus, our findings showed that Coutoubea spicata has antioxidant activity, reducing oxidative damage markers and consequently tissue damage in healthy Wistar rats after HIIT protocol, but it also demonstrated redox balance after analyzing oxidative stress markers.

RESUMEN: Este estudio tuvo como objetivo evaluar los efectos de HIIT en los marcadores de daño tisular y oxidativo en ratas suplementadas con Coutoubea spicata durante seis semanas. Treinta y dos ratas Wistar macho se dividieron en 4 grupos: línea de base (GB); suplementados con 100 mg/kg de fracción de Coutoubea spicata (GSCS); ejercitados durante 6 semanas con el protocolo HIIT (GH); suplementado con 100 mg/kg de fracción de Coutoubea spicata + HIIT durante 6 semanas (GHCS). Los animales ejercitados realizaron el protocolo HIIT (2x2). Se analizaron los marcadores de daño tisular CK, LDH, ALT y AST en plasma, así como los biomarcadores de estrés oxidativo MDA y SH en plasma y en tejidos cardiaco, hepático y muscular. Los resultados indicaron que las enzimas CK, LDH, AST y ALT mostraron aumento en GH en comparación con GB (p<0,0001). Sin embargo, los marcadores CK, AST y ALT redujeron sus concentraciones en GHCS en comparación con GH (p<0,0001), lo que indica que la suplementación con Coutoubea spicata atenuó el daño en los tejidos musculares y hepáticos inducido por HIIT. La MDA de plasma, hígado y músculo mostró un aumento en la GH después de las sesiones de HIIT; sin embargo, en comparación con GHCS, mostró niveles reducidos (p<0,0001). Se observó SH elevado en el grupo de GH en comparación con GB en plasma y tejidos hepáticos (p<0,0001); en contraste, se observó una reducción en GHCS en comparación con GH en plasma, hígado y tejidos cardíacos, lo que demuestra el efecto redox de HIIT en algunos tejidos. Por lo tanto, nuestros hallazgos mostraron que Coutoubea spicata tiene actividad antioxidante, con reducción de los marcadores de daño oxidativo y, en consecuencia, el daño tisular en ratas Wistar sanas después del protocolo HIIT, pero además demostró el equilibrio redox después de analizar los marcadores de estrés oxidativo.

Changes in female function and autonomous selfing across floral lifespan interact to drive variation in the cost of selfing.

PREMISE: Morphological and developmental changes as flowers age can impact patterns of mating. At the same time, direct or indirect costs of floral longevity can alter their fitness outcomes. This influence has been less appreciated, particularly with respect to the timing of selfing. We investigated changes in stigma events, autonomous selfing, outcross seed set capacity, and autofertility-a measure representing the potential for reproductive assurance-across floral lifespan in the mixed-mating biennial Sabatia angularis. METHODS: We examined stigma morphology and receptivity, autonomous self-pollen deposition, and seed number and size under autonomous self-pollination and hand outcross-pollination for flowers of different ages, from 1 d of female phase until 14 d. We compared autonomous seed production to maximal outcross seed production at each flower age to calculate an index of autofertility. RESULTS: The stigmatic lobes begin to untwist 1 d post anthesis. They progressively open, sextend, coil, and increase in receptivity, peaking or saturating at 8-11 d, depending on the measure. Autonomous seed production can occur early, but on average remains low until 6 d, when it doubles. In contrast, outcross seed number and size start out high, then decline precipitously. Consequently, autofertility increases steeply across floral lifespan. CONCLUSIONS: Changes in stigma morphology and receptivity, timing of autonomous self-pollen deposition, and floral senescence can interact to influence the relative benefit of autonomous selfing across floral lifespan. Our work highlights the interplay between evolution of floral longevity and the mating system, with implications for the maintenance of mixed mating in S. angularis.