Profile of a flower: How rates of morphological evolution drive floral diversification in Ericales and angiosperms.

PREMISE: Recent studies of floral disparity in the asterid order Ericales have shown that flowers vary strongly among families and that disparity is unequally distributed between the three flower modules (perianth, androecium, gynoecium). However, it remains unknown whether these patterns are driven by heterogeneous rates of morphological evolution or other factors. METHODS: Here, we compiled a data set of 33 floral characters scored for 414 species of Ericales sampled from 346 genera and all 22 families. We conducted ancestral state reconstructions using an equal-rates Markov model for each character. We estimated rates of morphological evolution for Ericales and for a separate angiosperm-wide data set of 19 characters and 792 species, creating "rate profiles" for Ericales, angiosperms, and major angiosperm subclades. We compared morphological rates among flower modules within each data set separately and between data sets, and we compared rates among angiosperm subclades using the angiosperm data set. RESULTS: The androecium exhibits the highest evolutionary rates across most characters, whereas most perianth and gynoecium characters evolve more slowly in both Ericales and angiosperms. Both high and low rates of morphological evolution can result in high floral disparity in Ericales. Analyses of an angiosperm-wide floral data set reveal that this pattern appears to be conserved across most major angiosperm clades. CONCLUSIONS: Elevated rates of morphological evolution in the androecium of Ericales may explain the higher disparity reported for this floral module. Comparing rates of morphological evolution through rate profiles proves to be a powerful tool in understanding floral evolution.

Genomic-based microsatellite development for Ternstroemia (Pentaphylacaceae) and transferability to other Ericales.

BACKGROUND: The genus Ternstroemia is associated with the vulnerable tropical montane cloud forest in Mexico and with other relevant vegetation types worldwide. It contains threatened and pharmacologically important species and has taxonomic issues regarding its species limits. This study describes 38 microsatellite markers generated using a genomic-based approach. METHODS AND RESULTS: We tested 23 of these markers in a natural population of Ternstroemia lineata. These markers are highly polymorphic (all loci polymorphic with 3-14 alleles per locus and expected heterozygosity between 0.202 and 0.908), most of them (19 out of 23) are in Hardy-Weinberg Equilibrium and free of null alleles (18 out of 23). Also we found no evidence of linkage among them. Finally, we tested the transferability to six other American species of Ternstroemia, two other Pentaphylacaceae species, and four species from different families within the order Ericales. CONCLUSIONS: These molecular resources are promising tools to investigate genetic diversity loss and as barcodes for ethnopharmacological applications and species delimitation in the family Pentaphylacaceae and some Ericales, among other applications.

Comparative analyses of three complete Primula mitogenomes with insights into mitogenome size variation in Ericales.

BACKGROUND: Although knowledge of the sizes, contents, and forms of plant mitochondrial genomes (mitogenomes) is increasing, little is known about the mechanisms underlying their structural diversity. Evolutionary information on the mitogenomes of Primula, an important ornamental taxon, is more limited than the information on their nuclear and plastid counterparts, which has hindered the comprehensive understanding of Primula mitogenomic diversity and evolution. The present study reported and compared three Primula mitogenomes and discussed the size expansion of mitogenomes in Ericales. RESULTS: Mitogenome master circles were sequenced and successfully assembled for three Primula taxa and were compared with publicly available Ericales mitogenomes. The three mitogenomes contained similar gene contents and varied primarily in their structures. The Primula mitogenomes possessed relatively high nucleotide diversity among all examined plant lineages. In addition, high nucleotide diversity was found among Primula species between the Mediterranean and Himalaya-Hengduan Mountains. Most predicted RNA editing sites appeared in the second amino acid codon, increasing the hydrophobic character of the protein. An early stop in atp6 caused by RNA editing was conserved across all examined Ericales species. The interfamilial relationships within Ericales and interspecific relationships within Primula could be well resolved based on mitochondrial data. Transfer of the two longest mitochondrial plastid sequences (MTPTs) occurred before the divergence of Primula and its close relatives, and multiple independent transfers could also occur in a single MTPT sequence. Foreign sequence [MTPTs and mitochondrial nuclear DNA sequences (NUMTs)] uptake and repeats were to some extent associated with changes in Ericales mitogenome size, although none of these relationships were significant overall. CONCLUSIONS: The present study revealed relatively conserved gene contents, gene clusters, RNA editing, and MTPTs but considerable structural variation in Primula mitogenomes. Relatively high nucleotide diversity was found in the Primula mitogenomes. In addition, mitogenomic genes, collinear gene clusters, and locally collinear blocks (LCBs) all showed phylogenetic signals. The evolutionary history of MTPTs in Primula was complicated, even in a single MTPT sequence. Various reasons for the size variation observed in Ericales mitogenomes were found.

Phylotranscriptomics of Theaceae: generic-level relationships, reticulation and whole-genome duplication.

BACKGROUND AND AIMS: Theaceae, with three tribes, nine genera and more than 200 species, are of great economic and ecological importance. Recent phylogenetic analyses based on plastomic data resolved the relationships among the three tribes and the intergeneric relationships within two of those tribes. However, generic-level relationships within the largest tribe, Theeae, were not fully resolved. The role of putative whole-genome duplication (WGD) events in the family and possible hybridization events among genera within Theeae also remain to be tested further. METHODS: Transcriptomes or low-depth whole-genome sequencing of 57 species of Theaceae, as well as additional plastome sequence data, were generated. Using a dataset of low-copy nuclear genes, we reconstructed phylogenetic relationships using concatenated, species tree and phylogenetic network approaches. We further conducted molecular dating analyses and inferred possible WGD events by examining the distribution of the number of synonymous substitutions per synonymous site (Ks) for paralogues in each species. For plastid protein-coding sequences , phylogenies were reconstructed for comparison with the results obtained from analysis of the nuclear dataset. RESULTS: Based on the 610 low-copy nuclear genes (858 606 bp in length) investigated, Stewartieae was resolved as sister to the other two tribes. Within Theeae, the Apterosperma-Laplacea clade grouped with Pyrenaria, leaving Camellia and Polyspora as sister. The estimated ages within Theaceae were largely consistent with previous studies based mainly on plastome data. Two reticulation events within Camellia and one between the common ancestor of Gordonia and Schima were found. All members of the tea family shared two WGD events, an older At-γ and a recent Ad-ß; both events were also shared with the outgroups (Diapensiaceae, Pentaphylacaceae, Styracaceae and Symplocaceae). CONCLUSIONS: Our analyses using low-copy nuclear genes improved understanding of phylogenetic relationships at the tribal and generic levels previously proposed based on plastome data, but the phylogenetic position of the Apterosperma-Laplacea clade needs more attention. There is no evidence for extensive intergeneric hybridization within Theeae or for a Theaceae-specific WGD event. Land bridges (e.g. the Bering land bridge) during the Late Oligocene may have permitted the intercontinental plant movements that facilitated the putative ancient introgression between the common ancestor of Gordonia and Schima.

Genetic diversity and correlation between morphological traits of pequi fruits (Caryocar brasiliense Camb.) with and without thorns at the endocarp.

Morphological characteristics of pequi fruits (Caryocar brasiliense) allows us to obtain information for the conservation, breeding, and detect genetic variability. Thus, the aims of this research were to characterize the genetic diversity of Caryocar brasiliense that produce fruits with and without thorn at the endocarp; determine the important characters in the study of genetic diversity, and to estimate phenotypic and genotypic correlations. 80 fruits with thorns and 90 fruits without thorns were evaluated. Data were used for analysis: Euclidian distance as a measure of dissimilarity and the Tocher method for delimitation of groups; Principal Component Analysis, and genotypic and phenotypic correlations. Genetic dissimilarity of C. brasiliense ranged from 0.066 (B6-D6) to 0.908 (A14-B3). Tocher method allowed dividing the matrices into three groups, the dendrogram into five groups, and the principal components allowed the distribution in 8 groups. We observed 15 significant correlations for characters of fruit with thorns and 31 correlations for characters of fruit without thorns. The weight of the fruit (with thorns) and the pyrene yield (without thorns) are relevant correlations for production. The groupings generated allowed the classification of the matrices into distinct sets. Allowed the identification of potential producers plants that could be used in breeding programs.

Global patterns and a latitudinal gradient of flower disparity: perspectives from the angiosperm order Ericales.

Morphological diversity (disparity) is an essential but often neglected aspect of biodiversity. Hence, it seems timely and promising to re-emphasize morphology in modern evolutionary studies. Disparity is a good proxy for the diversity of functions and interactions with the environment of a group of taxa. In addition, geographical and ecological patterns of disparity are crucial to understand organismal evolution and to guide biodiversity conservation efforts. Here, we analyse floral disparity across latitudinal intervals, growth forms, climate types, types of habitats, and regions for a large and representative sample of the angiosperm order Ericales. We find a latitudinal gradient of floral disparity and a decoupling of disparity from species richness. Other factors investigated are intercorrelated, and we find the highest disparity for tropical trees growing in African and South American forests. Explanations for the latitudinal gradient of floral disparity may involve the release of abiotic constraints and the increase of biotic interactions towards tropical latitudes, allowing tropical lineages to explore a broader area of the floral morphospace. Our study confirms the relevance of biodiversity parameters other than species richness and is consistent with the importance of species interactions in the tropics, in particular with respect to angiosperm flowers and their pollinators.

Efficacy of Souroubea-Platanus Dietary Supplement Containing Triterpenes in Beagle Dogs Using a Thunderstorm Noise-Induced Model of Fear and Anxiety.

A novel botanical dietary supplement, formulated as a chewable tablet containing a defined mixture of Souroubea spp. vine and Platanus spp. Bark, was tested as a canine anxiolytic for thunderstorm noise-induced stress (noise aversion). The tablet contained five highly stable triterpenes and delivered 10 mg of the active ingredient betulinic acid (BA) for an intended 1 mg/kg dose in a 10 kg dog. BA in tablets was stable for 30 months in storage at 23 °C. Efficacy of the tablets in reducing anxiety in dogs was assessed in a blinded, placebo-controlled study by recording changes in blood cortisol levels and measures of behavioral activity in response to recorded intermittent thunder. Sixty beagles were assigned into groups receiving: placebo, 0.5×, 1×, 2×, and 4× dose, or the positive control (diazepam), for five days. Reduction in anxiety measures was partially dose-dependent and the 1× dose was effective in reducing inactivity time (p = 0.0111) or increased activity time (p = 0.0299) compared with placebo, indicating a decrease in anxiety response. Cortisol measures also showed a dose-dependent reduction in cortisol in dogs treated with the test tablet.

Effect of Extract and Synthesized Derivatives of Isolated Compound from Symplocos chinensis f. Pilosa Ohwi on Neuropathic Pain in Mice.

Neuropathic pain is described as the "most terrible of all tortures that a nerve wound may inflict." The aim of the present study was to demonstrate the antinociceptive effect of Symplocos chinensis f. pilosa Ohwi water extract (SCW) and synthesized derivatives of the isolated compound. The antinociceptive effect was tested using the acetic acid-induced writhing and 5% formalin tests. Antinociceptive effects on neuropathic pain were evaluated using the von Frey test with chronic constriction injury (CCI) and surgical nerve injury (SNI) models and tail-flick test with a vincristine-induced pain model. An Ames test was also conducted. 5-hydroxymethylfurfural (5-HMF) was isolated and derivatives were synthesized with various acid groups. Among the plant water extracts, SCW showed significantly effective activity. Additionally, SCW presented antinociceptive effects in the neuropathic pain models. The SCW water fraction resulted in fewer writhes than the other fractions, and isolated 5-HMF was identified as an effective compound. Because 5-HMF revealed a positive response in the Ames test, derivatives were synthesized. Among the synthesized derivations, 5-succinoxymethylfurfural (5-SMF) showed the best effect in the neuropathic pain model. Our data suggest that SCW and the synthesized compound, 5-SMF, possess effective antinociceptive activity against neuropathic pain.

Activity Guided Isolation of Phenolic Compositions from Anneslea fragrans Wall. and Their Cytoprotective Effect against Hydrogen Peroxide Induced Oxidative Stress in HepG2 Cells.

Anneslea fragrans Wall., commonly known as "Pangpo Tea", is traditionally used as a folk medicine and healthy tea for the treatment of liver and intestine diseases. The aim of this study was to purify the antioxidative and cytoprotective polyphenols from A. fragrans leaves. After fractionation with polar and nonpolar organic solvents, the fractions of aqueous ethanol extract were evaluated for their total phenolic (TPC) and flavonoid contents (TFC) and antioxidant activities (DPPH, ABTS, and FRAP assays). The n-butanol fraction (BF) showed the highest TPC and TFC with the strongest antioxidant activity. The bio-guided chromatography of BF led to the purification of six flavonoids (1-6) and one benzoquinolethanoid (7). The structures of these compounds were determined by NMR and MS techniques. Compound 6 had the strongest antioxidant capacity, which was followed by 5 and 2. The protective effect of the isolated compounds on hydrogen peroxide (H2O2)-induced oxidative stress in HepG2 cells revealed that the compounds 5 and 6 exhibited better protective effects by inhibiting ROS productions, having no significant difference with vitamin C (p > 0.05), whereas 6 showed the best anti-apoptosis activity. The results suggest that A. fragrans could serve as a valuable antioxidant phytochemical source for developing functional food and health nutraceutical products.

A herbal treatment approach for the management of clinical, hormonal and ultrasound parameters in reproductive age group women with polycystic ovarian syndrome: A randomized clinical trial.

Around fifteen percent women of reproductive age have been effected by Polycystic Ovarian Syndrome (PCOS); a complicated disorder; and apparently there is no standard therapy available. Considering this lack, we design present work; for the assessment of a herbal medicine (Femitex-SP4) in managing PCOs. During 2016-17; this study was carried out at Abbasi Shaheed hospital, Karachi, Pakistan. A total of 150 patients aged between 18-44 years were included as per Rotterdam criteria. Patients received 500 mg of powdered herbs in capsule form twice daily. The primary outcomes were regular menstruation and ovulation plus change in fasting blood sugar levels. Changes in free testosterone levels and ovarian morphology was secondary outcome measures. Continuous outcomes before and after treatment were compared by Student's t-test (one tailed, independent). P = 0.05 was considered as significant. Women menstrual cycle was considerably improved. Fasting blood sugar levels did not change (p=0.103392). Progesterone levels were same at the starting point and after treatment (P=0.318322). With complete recovery in 6 patients; a notable change was found in ovarian size. Free testosterone levels were also dropped significantly (p<0.00001). Our main success was drastic improvement in normalizing menstrual cycle during therapy. Herbal treatment is proven to be clinically effective in most of the patients; particularly PCOs patients with menstrual irregularities. Hence, Femitex-SP4 can be taken as a better treatment for PCOs.

The N-terminal and third transmembrane domain of PsCor413im1 are essential for targeting to chloroplast envelope membrane.

Cold-regulated (COR) genes, located downstream of the C-repeat binding factors (CBFs) in cold signaling pathways, play a central role in plant response to cold stress. In our previous studies, a Cor413 chloroplast envelope membrane protein, PsCor413im1, was identified from the cold-tolerant plant Phlox subulata. Its overexpression enhanced cold tolerance and altered AtCor15 expression in Arabidopsis. In the present study, the function of PsCor413im1 was further investigated. Transmission electron microscope observation showed that the chloroplast envelope membrane of cold-treated transgenic Arabidopsis seedlings was more stable than that of cold-treated wild-type seedlings. Subcellular localization of green fluorescent protein as a marker revealed that the N-terminal and putative third transmembrane domain (TMD) of PsCor413im1 were essential for its targeting of the chloroplast envelope membrane. Furthermore, overexpression of PsCor413im1 fragments containing N-terminal and third TMD also altered the expression of AtCor15 genes in Arabidopsis. Overall, our results suggest that PsCor413im1 may stabilize the chloroplast envelope membrane under cold stress, and its N-terminal and third TMD are important for its targeting capability and function.

A consensus phylogenomic approach highlights paleopolyploid and rapid radiation in the history of Ericales.

PREMISE: Large genomic data sets offer the promise of resolving historically recalcitrant species relationships. However, different methodologies can yield conflicting results, especially when clades have experienced ancient, rapid diversification. Here, we analyzed the ancient radiation of Ericales and explored sources of uncertainty related to species tree inference, conflicting gene tree signal, and the inferred placement of gene and genome duplications. METHODS: We used a hierarchical clustering approach, with tree-based homology and orthology detection, to generate six filtered phylogenomic matrices consisting of data from 97 transcriptomes and genomes. Support for species relationships was inferred from multiple lines of evidence including shared gene duplications, gene tree conflict, gene-wise edge-based analyses, concatenation, and coalescent-based methods, and is summarized in a consensus framework. RESULTS: Our consensus approach supported a topology largely concordant with previous studies, but suggests that the data are not capable of resolving several ancient relationships because of lack of informative characters, sensitivity to methodology, and extensive gene tree conflict correlated with paleopolyploidy. We found evidence of a whole-genome duplication before the radiation of all or most ericalean families, and demonstrate that tree topology and heterogeneous evolutionary rates affect the inferred placement of genome duplications. CONCLUSIONS: We provide several hypotheses regarding the history of Ericales, and confidently resolve most nodes, but demonstrate that a series of ancient divergences are unresolvable with these data. Whether paleopolyploidy is a major source of the observed phylogenetic conflict warrants further investigation.

Biological Potential of Fruit and Leaves of Strawberry Tree (Arbutus unedo L.) from Croatia.

The strawberry tree fruit and leaf are a rich source of minerals, easily digestible sugars, dietary fibers, vitamins (especially vitamin C) and many bioactive compounds of significant functional value. Due to their favorable chemical composition, fruits have recently become increasingly popular in consumption. The aim of this study was to determine the physical-chemical composition, content of bioactive compounds, and also the antioxidant capacity of the fruit and leaves of wild strawberry tree populations among the Adriatic coast in Croatia, as well as to investigate the influence of location on the content of specific bioactive compounds. According to the obtained results, both fruit and leaves are pronouncedly high in vitamin C content, the average value for fruits amounted to 224.21 mg/100 g FW, while that for leaves amounted to 138.08 mg/100 g FW. Additionally, significantly high values of total polyphenolic compounds were recorded both in fruits (average value of 637.94 mg GAE/100 g FW) and especially in leaves (average value of 2157.01 mg GAE/100 g FW). Several pigments from different categories were determined in the fruit depending on fruit maturity, including: total anthocyanins, ß-carotene and lycopene; while in leaves chlorophylls and carotenoids. Given the high content of different bioactive compounds high values of antioxidant capacity were determined (the average value for fruits was 2269.96 µmol TE/kg and for leaves, 2237.16 µmol TE/kg). Location strongly influenced the physical-chemical composition and also the content of specialized metabolites; populations collected from southern areas (central and south Dalmatia) of the Adriatic coast tended to have higher amounts of vitamin C, total phenols, total anthocyanins and ß-carotene.

Caryocar brasiliense Cambess. Pulp Oil Supplementation Reduces Total Cholesterol, LDL-c, and Non-HDL-c in Animals.

The fruit of Caryocar brasiliense Cambess. is a source of oil with active compounds that are protective to the organism. In our work, we analyzed the physicochemical characteristics and evaluated the effects of supplementation with C. brasiliense oil in an animal model. We characterized the oil by indices of quality and identity, optical techniques of absorption spectroscopy in the UV-Vis region and fluorescence, and thermogravimetry/derived thermogravimetry (TG/DTG). For the animal experiment, we utilized mice (Mus musculus) supplemented with lipidic source in different dosages. The results demonstrated that C. brasiliense oil is an alternative source for human consumption and presents excellent oxidative stability. Primarily, it exhibited oleic MFA (53.56%) and palmitic SFA (37.78%). The oil level of tocopherols and tocotrienols was superior to the carotenoids. The supplementation with C. brasiliense oil reduced the levels of total cholesterol, LDL-c, and non-HDL-c. Regarding visceral fats and adiposity index, the treatment synergically supplemented with olive oil and C. brasiliense oil (OO + CO) obtained the best result. Therefore, C. brasiliense oil is a high quality product for consumption. Its supplementation promotes beneficial effects mainly on the lipidic profile.

Stem succulence controls flower and fruit production but not stem growth in the desert shrub ocotillo (Fouquieria splendens).

PREMISE OF THE STUDY: The C3 desert shrub ocotillo (Fouquieria splendens) completely lacks xeromorphic leaves but is uncommonly both stem succulent and repetitively drought deciduous (documented to have produced many foliation-defoliation cycles during a growing season). Both adaptations conserve water in this xerophyte, but are the roles of succulence and deciduousness merely redundant? The observation that year-to-year reproductive effort was relatively consistent while vegetative growth was not offered a critical clue that, coupled with long-term precipitation data, helped answer this question. METHODS: At two sites in the Chihuahuan Desert in southern New Mexico, United States, 22 ocotillos were studied annually for more than two decades to explore the relationships among reproductive effort, vegetative stem growth, and patterns of precipitation. KEY RESULTS: Vegetative stem growth occurred in mid- to late summer (July-September), the season of maximum precipitation in the Chihuahuan Desert, and was significantly related to summer precipitation received in the year of growth. Reproductive effort occurred in early to late spring (April-June), which with winter account for minimum precipitation during the year, but was significantly related to summer precipitation received in the previous year, suggesting the importance of stem succulence and stored water. CONCLUSIONS: While highly variable summer precipitation was responsible for enormous fluctuations in annual ocotillo stem growth, stem succulence insulated reproductive effort from such immense variability. Stem-stored water allowed the production of flowers and fruits to proceed relatively consistently during the driest years and during the driest time of year in the Chihuahuan Desert.

Chemical Constituents of Anneslea fragrans and Their Antiausterity Activity against the PANC-1 Human Pancreatic Cancer Cell Line.

An ethanolic extract of Anneslea fragrans leaves showed potent preferential cytotoxicity against PANC-1 human pancreatic cancer cells under a nutrient-deprived condition, with a PC50 value of 9.6 µg/mL. Phytochemical investigation of this active extract led to the isolation of two new secondary metabolites, fragranones A (1) and B (2), along with 15 previously reported compounds. The structure elucidation of the new compounds was achieved by HRFABMS, acid hydrolysis, NMR, and ECD spectroscopic analysis. Fragranone A (1) is the first example of a rare natural product bearing an acetonide glucose moiety. Fragranone B (2) is representative of a rare class of natural products with a threonolactone unit linked to a chalcone through an ether linkage. The isolated compounds exhibited antiausterity activity against PANC-1 cells under nutrient-deprived conditions, and betulin (14) was found to be the most potent compound tested, with a PC50 value of 8.4 µM. In addition, fragranone A (1) was found to suppress PANC-1 cancer cell migration in real time.

Pequi enriched diets protect Drosophila melanogaster against paraquat-induced locomotor deficits and oxidative stress.

The species Caryocar coriaceum Wittm (C. coriaceum), is popularly employed in northeast of Brazil for culinary purposes and in folk medicine. The oil from its fruit, deignated Pequi, is commonly used to treat inflammatory problems, and its leaves to treat viral infections. However, comprehensive knowledge regarding the pharmacological properties attributed to these plant parts is still scarce. Thus, this study aimed to explore the in vivo antioxidant potential of aqueous extract of the leaves (AEL) and Pequi pulp oil (PPO) on the pro-oxidative effects induced by paraquat (PQ) using Drosophila melanogaster (D. melanogaster) as a model. These flies were fed with either standard or AEL and PPO supplemented diets prior to (pre-treatment for 7 days) or concomitantly (co-treatment for 5 days) with PQ. D. melanogaster administered PQ exhibited locomotor deficits and a higher rate of mortality. PQ induced significant changes in the antioxidant/oxidant status of D. melanogaster, including significant (1) increase in levels of reactive oxygen species (ROS) and lipid peroxidation; (2) elevation in the activity of antioxidant enzymes catalase (CAT) and glutathione-S-transferase (GST) and marked up-regulation in mRNA expression of stress-related genes for CAT, superoxide dismutase (SOD), thioredoxin reductase and Keap-1. Aside for mortality rates, AEL and PPO treatments reduced PQ-induced oxidative stress and motor impairments. No apparent evidence of toxicity was observed in D. melanogaster fed with AEL and PPO alone. Our findings provide evidence that AEL and PPO may confer protection against oxidant conditions by stimulating antioxidant responses.

Development and Evaluation of Laboratory Bioassays to Study Powdery Mildew Pathogens of Phlox In Vitro.

The genus Phlox consists of approximately 65 species that include some of the most prevalent ornamental plants in the temperate zone. These popular ornamentals are extremely susceptible to powdery mildew (PM) caused by the biotrophic fungi Golovinomyces magnicellulatus and Podosphaera sp. In this study, we used Phlox paniculata and P. glaberrima to develop a set of laboratory tools to study these pathogens in vitro, including a detached leaf and a micropropagated plantlet bioassay. We assessed pathogen growth under different experimental conditions, which included the use of four different media variations (1/2 MS medium amended with benzimidazole and tetracycline), three ages of pathogen culture (14, 18, and 22 days), three phenological stages of the host tissue (1st, 3rd, and 5th node leaves), placement of inoculum on both leaf surfaces (abaxial and adaxial), and three different inoculation techniques (single spore transfer, colony tapping, colony brushing). Detached P. paniculata leaves were successfully maintained on benzimidazole-amended 1/2 MS medium for up to 3 weeks. For both pathogens, the use of 18-day-old cultures resulted in a higher number of larger, higher sporulating colonies compared with 1-4 and 22-day-old cultures. The adaxial side of 3rd node leaves supported statistically significant more fungal growth compared with the adaxial side of 1st and 5th node leaves. Both pathogens also successfully infected micropropagated plantlets of P. glaberrima. These newly developed tools should facilitate in vitro studies on PM of Phlox and possibly be applicable to other ornamental species attacked by the same fungi.

A novel cold-regulated gene from Phlox subulata, PsCor413im1, enhances low temperature tolerance in Arabidopsis.

Low temperature stress adversely affects plant growth, development, and crop productivity. Analysis of the function of genes in the response of plants to low temperature stress is essential for understanding the mechanism of chilling and freezing tolerance. In this study, PsCor413im1, a novel cold-regulated gene isolated from Phlox subulata, was transferred to Arabidopsis to investigate its function under low temperature stress. Real-time quantitative PCR analysis revealed that PsCor413im1 expression was induced by cold and abscisic acid. Subcellular localization revealed that PsCor413im1-GFP fusion protein was localized to the periphery of the chloroplast, consistent with the localization of chloroplast inner membrane protein AtCor413im1, indicating that PsCor413im1 is a chloroplast membrane protein. Furthermore, the N-terminal of PsCor413im1 was determined to be necessary for its localization. Compared to the wild-type plants, transgenic plants showed higher germination and survival rates under cold and freezing stress. Moreover, the expression of AtCor15 in transgenic plants was higher than that in the wild-type plants under cold stress. Taken together, our results suggest that the overexpression of PsCor413im1 enhances low temperature tolerance in Arabidopsis.

Recent radiation and dispersal of an ancient lineage: The case of Fouquieria (Fouquiericeae, Ericales) in North American deserts.

Arid biomes are particularly prominent in the Neotropics providing some of its most emblematic landscapes and a substantial part of its species diversity. To understand some of the evolutionary processes underlying the speciation of lineages in the Mexican Deserts, the diversification of Fouquieria is investigated, which includes eleven species, all endemic to the warm deserts and dry subtropical regions of North America. Using a phylogeny from plastid DNA sequences with samples of individuals from populations of all the species recognized in Fouquieria, we estimate divergence times, test for temporal diversification heterogeneity, test for geographical structure, and conduct ancestral area reconstruction. Fouquieria is an ancient lineage that diverged from Polemoniaceae ca. 75.54 Ma. A Mio-Pliocene diversification of Fouquieria with vicariance, associated with Neogene orogenesis underlying the early development of regional deserts is strongly supported. Test for temporal diversification heterogeneity indicates that during its evolutionary history, Fouquieria had a drastic diversification rate shift at ca.12.72 Ma, agreeing with hypotheses that some of the lineages in North American deserts diversified as early as the late Miocene to Pliocene, and not during the Pleistocene. Long-term diversification dynamics analyses suggest that extinction also played a significant role in Fouquieria's evolution, with a very high rate at the onset of the process. From the late Miocene onwards, Fouquieria underwent substantial diversification change, involving high speciation decreasing to the present and negligible extinction, which is congruent with its scant fossil record during this period. Geographic phylogenetic structure and the pattern of most sister species inhabiting different desert nucleus support that isolation by distance could be the main driver of speciation.