Cytokinins act synergistically with heat acclimation to enhance rice thermotolerance affecting hormonal dynamics, gene expression and volatile emission.

Heat stress is a frequent environmental constraint. Phytohormones can significantly affect plant thermotolerance. This study compares the effects of exogenous cytokinin meta-topolin-9-(tetrahydropyran-2-yl)purine (mT9THP) on rice (Oryza sativa) under control conditions, after acclimation by moderate temperature (A; 37 °C, 2h), heat stress (HS; 45 °C, 6h) and their combination (AHS). mT9THP is a stable cytokinin derivative that releases active meta-topolin gradually, preventing the rapid deactivation reported after exogenous cytokinin application. Under control conditions, mT9THP negatively affected jasmonic acid in leaves and abscisic and salicylic acids in crowns (meristematic tissue crucial for tillering). Exogenous cytokinin stimulated the emission of volatile organic compounds (VOC), especially 2,3-butanediol. Acclimation upregulated trans-zeatin, expression of stress- and hormone-related genes, and VOC emission. The combination of acclimation and mT9THP promoted the expression of stress markers and antioxidant enzymes and moderately increased VOC emission, including 2-ethylhexyl salicylate or furanones. AHS and HS responses shared some common features, namely, increase of ethylene precursor aminocyclopropane-1-carboxylic acid (ACC), cis-zeatin and cytokinin methylthio derivatives, as well as the expression of heat shock proteins, alternative oxidases, and superoxide dismutases. AHS specifically induced jasmonic acid and auxin indole-3-acetic acid levels, diacylglycerolipids with fewer double bonds, and VOC emissions [e.g., acetamide, lipoxygenase (LOX)-derived volatiles]. Under direct HS, exogenous cytokinin mimicked some positive acclimation effects. The combination of mT9THP and AHS had the strongest thermo-protective effect, including a strong stimulation of VOC emissions (including LOX-derived ones). These results demonstrate for the first time the crucial contribution of volatiles to the beneficial effects of cytokinin and AHS on rice thermotolerance.

The nociceptive response during adult cardiac surgery measured by the qNOX index: A feasibility study.

BACKGROUND: Monitoring nociception during general anaesthesia remains a substantial challenge. The Conox monitor uses two EEG indices, the qCon and the qNox. The qNox refers to the probability that a nociceptive stimulation triggers a movement of the patient and the response probability of reaction to nociceptive stimulation. We decided to test the feasibility of monitoring the qNOX index during adult cardiac surgery and to investigate whether this index correlates with hemodynamic and hormonal signs of nociceptive stimulation. METHOD: We enrolled 19 patients undergoing elective cardiac surgery. These were randomised to 2 groups receiving different doses of sufentanil via target controlled infusion: group A (n=9) 0.25 ng/mL and group B (n=10) 0.75 ng/mL. All patients were maintained at the same depth of anaesthesia. We recorded the Conox monitor indices (qNOX, qCON, electromyographic), hemodynamic variables and plasmatic levels of cortisol and noradrenaline. RESULTS: There was significantly higher blood pressure (P=0.013) and plasmatic cortisol (P=0.003) in group A and a significant increase in plasmatic noradrenaline with increasing intensity of surgical stimulation in both groups: A (P=0.001), B (P=0.008). We found no significant corresponding changes in the qNOX index. There was a positive correlation between qNOX and hemodynamic signs of stimulation (P=0.012) and between the qNOX and EMG indices (P=0.013) after endotracheal intubation, but not later after EMG index dropped. CONCLUSION: Our results do not support the assumption that signs of nociceptive stimulation during adult cardiac surgery will be reflected by the qNOX index. This may be related to compounding of qNOX processing and use of muscle relaxants. Further investigation on this field is needed.

Seed Protection of Solanum lycopersicum with Pythium oligandrum against Alternaria brassicicola and Verticillium albo-atrum.

Pythium oligandrum, strain M1, is a soil oomycete successfully used as a biological control agent (BCA), protecting plants against fungal, yeast, and oomycete pathogens through mycoparasitism and elicitor-dependent plant priming. The not yet described Pythium strains, X42 and 00X48, have shown potential as BCAs given the high activity of their secreted proteases, endoglycosidases, and tryptamine. Here, Solanum lycopersicum L. cv. Micro-Tom seeds were coated with Pythium strains, and seedlings were exposed to fungal pathogens, either Alternaria brassicicola or Verticillium albo-atrum. The effects of both infection and seed-coating on plant metabolism were assessed by determining the activity and isoforms of antioxidant enzymes and endoglycosidases and the content of tryptamine, amino acids, and heat shock proteins. Dual culture competition testing and microscopy analysis confirmed mycoparasitism in all three Pythium strains. In turn, seed treatment significantly increased the total free amino acid content, changing their abundance in both non-infected and infected plants. In response to pathogens, plant Hsp70 and Hsp90 isoform levels also varied among Pythium strains, most likely as a strategy for priming the plant against infection. Overall, our results show in vitro mycoparasitism between Pythium strains and fungal pathogens and in planta involvement of heat shock proteins in priming.

Vasa nervorum of epicardial nerves and valves of small veins investigated in porcine heart by two types of vascular injections.

The morphology of the vascular supply of peripheral branches of cardiac nerves has not been systematically described until now. The aim of this study was to describe the architectonics of the vasa nervorum of epicardial nerves in porcine hearts by using two injection techniques. Twenty-three hearts from young healthy pigs were used. In 10 hearts India ink solution was injected into the origin of the anterior interventricular branch. In another 10 hearts India ink solution was injected retrogradely through the coronary sinus. The hearts were then analyzed using a magnifying glass and light microscopy. The arterial injection showed the entirety of the rich venous components of the vasa nervorum, which often consisted of paired veins accompanying the epicardial nerves. The thickness of the nerves ranged from 50 to 815 µm. The vasa nervorum drained into larger subepicardial veins. In seven of the hearts prepared with venous injections the vasa nervorum of epicardial nerves were visualized in the same detail as in the arterial preparations and India ink solution filled the right ventricle via the smallest cardiac veins. The histological analysis of these seven hearts showed complete dehiscence and functional insufficiency of small and larger veins valves. In the other three hearts prepared with venous injections the valves were competent, which prevented retrograde filling of larger and smaller veins. The results obtained expand the current knowledge on epicardial nerves vasa nervorum and provide anatomical evidence behind the mechanism of retrograde application of cardioplegic solutions in cardiac surgery.

Synthesis, Inhibitory Activity, and In Silico Modeling of Selective COX-1 Inhibitors with a Quinazoline Core.

Selective cyclooxygenase-1 (COX-1) inhibition has got into the spotlight with the discovery of COX-1 upregulation in various cancers and the cardioprotective role of COX-1 in control of thrombocyte aggregation. Yet, COX-1-selective inhibitors are poorly explored. Thus, three series of quinazoline derivatives were prepared and tested for their potential inhibitory activity toward COX-1 and COX-2. Of the prepared compounds, 11 exhibited interesting COX-1 selectivity, with 8 compounds being totally COX-1-selective. The IC50 value of the best quinazoline inhibitor was 64 nM. The structural features ensuring COX-1 selectivity were elucidated using in silico modeling.

European ferns as rich sources of antioxidants in the human diet.

Wild edible plants have become an attractive variation of the human diet, especially in East Asia, North America, and Oceania. However, their potential in nutrition is only rarely considered in Europe. This study aims to reveal the nutritional and antioxidant potential of mature fern leaves from 13 families grown in Europe. We found that most of the examined fern species displayed a high antioxidant capacity, exceeding 0.5 g Trolox equivalent per gram of extract dry weight in ORAC assay and reaching IC50 values lower than 30 µg·mL-1 in DPPH assay (with the value for Trolox 7 µg·mL-1). Most of the species also appeared to be a good source of carotenoids, especially of lutein (205 µg·g-1 DW on average) and ß-carotene (161 µg·g-1 DW on average) when compared to the reference leafy vegetables spinach and rocket. A cytotoxicity test using ovine hepatocytes showed a non-toxicity effect of fern leaf extracts.

Nutritional and Antioxidant Potential of Fiddleheads from European Ferns.

Ferns are part of the diet and traditional medicine in East Asia, North America, and Oceania, however, their importance has been forgotten in Europe. Here, the nutritional and antioxidant potential of young fern fronds (fiddleheads) of eight families were studied. Most of the tested fern species excelled in high antioxidant capacity when compared to the reference leafy vegetables spinach and rocket. On average, the total phenol content reached 220 mg·g-1 of extract dry weight for all fiddleheads, and 15 out of 24 tested species exceeded 1 g Trolox equivalent per gram of extract dry weight in Oxygen Radical Absorbance Capacity (ORAC) assay. On the other hand, fiddleheads contained a comparable amount of carotenoids and ascorbic acid with the reference vegetables. In the case of fatty acid composition, fiddleheads contained especially high amounts of essential omega-3 (n3) and omega-6 (n6) polyunsaturated fatty acids with a beneficial n6/n3 ratio. The n6/n3 ratio in all tested species was between 2 and 6.4, whereas the ratio in the reference vegetables was below 0.4. All in all, fiddleheads from European ferns are a rich source of valuable antioxidants and essential fatty acids with a desirable n-6/n-3 ratio and may thus form an alternative source of these compounds, especially for those people not consuming fish and fish products.

Novel Insights into the Effect of Pythium Strains on Rapeseed Metabolism.

Pythium oligandrum is a unique biological control agent. This soil oomycete not only acts as a mycoparasite, but also interacts with plant roots and stimulates plant defense response via specific elicitors. In addition, P. oligandrum can synthetize auxin precursors and stimulate plant growth. We analyzed the secretomes and biochemical properties of eleven Pythium isolates to find a novel and effective strain with advantageous features for plants. Our results showed that even closely related P. oligandrum isolates significantly differ in the content of compounds secreted into the medium, and that all strains secrete proteins, amino acids, tryptamine, phenolics, and hydrolytic enzymes capable of degrading cell walls (endo-ß-1,3-glucanase, chitinase, and cellulase), exoglycosidases (especially ß-glucosidase), proteases, and phosphatases. The most different strain was identified as a not yet described Pythium species. The changes in metabolism of Brassica napus plants grown from seeds coated with the tested Pythium spp. were characterized. Enhanced levels of jasmonates, ethylene precursor, and salicylic acid may indicate better resistance to a wide variety of pathogens. Glucosinolates, as defense compounds against insects and herbivores, were enhanced in young plants. Altogether, P. oligandrum strains varied in their life strategies, and either they could perform equally as plant growth promoters and mycoparasites or they had developed one of these strategies better.

Synthesis, inhibitory activity and in silico docking of dual COX/5-LOX inhibitors with quinone and resorcinol core.

Based on the significant anti-inflammatory activity of natural quinone primin (5a), series of 1,4-benzoquinones, hydroquinones, and related resorcinols were designed, synthesized, characterized and tested for their ability to inhibit the activity of cyclooxygenase (COX-1 and COX-2) and 5-lipoxygenase (5-LOX) enzymes. Structural modifications resulted in the identification of two compounds 5b (2-methoxy-6-undecyl-1,4-benzoquinone) and 6b (2-methoxy-6-undecyl-1,4-hydroquinone) as potent dual COX/5-LOX inhibitors. The IC50 values evaluated in vitro using enzymatic assay were for compound 5b IC50 = 1.07, 0.57, and 0.34 µM and for compound 6b IC50 = 1.07, 0.55, and 0.28 µM for COX-1, COX-2, and 5-LOX enzyme, respectively. In addition, compound 6d was identified as the most potent 5-LOX inhibitor (IC50 = 0.14 µM; reference inhibitor zileuton IC50 = 0.66 µM) from the tested compounds while its inhibitory potential against COX enzymes (IC50 = 2.65 and 2.71 µM for COX-1 and COX-2, respectively) was comparable with the reference inhibitor ibuprofen (IC50 = 4.50 and 2.46 µM, respectively). The most important structural modification leading to increased inhibitory activity towards both COXs and 5-LOX was the elongation of alkyl chain in position 6 from 5 to 11 carbons. Moreover, the monoacetylation in ortho position of bromo-hydroquinone 13 led to the discovery of potent (IC50 = 0.17 µM) 5-LOX inhibitor 17 (2-bromo-6-methoxy-1,4-benzoquinone) while bromination stabilized the hydroquinone form. Docking analysis revealed the interaction of compounds with Tyr355 and Arg120 in the catalytic site of COX enzymes, while the hydrophobic parts of the molecules filled the hydrophobic substrate channel leading up to Tyr385. In the allosteric catalytic site of 5-LOX, compounds bound to Tyr142 and formed aromatic interactions with Arg138. Taken together, we identified optimal alkyl chain length for dual COX/5-LOX inhibition and investigated other structural modifications influencing COX and 5-LOX inhibitory activity.

Ivermectin biotransformation and impact on transcriptome in Arabidopsis thaliana.

Veterinary drugs enter the environment in many ways and may affect non-target organisms, including plants. The present project was focused on the biotransformation of ivermectin (IVM), one of the mostly used anthelmintics, in the model plant Arabidopsis thaliana. Our results certified the ability of plants to uptake IVM by roots and translocate it to the aboveground parts. Using UHPLC-MS/MS, six metabolites in roots and only the parent drug in rosettes were found after 24- and 72-h incubation of A. thaliana with IVM. The metabolites were formed only via hydroxylation and demethylation, with no IVM conjugates detected. Although IVM did not induce changes in the activity of antioxidant enzymes in A. thaliana rosettes, the expression of genes was significantly affected. Surprisingly, a higher number of transcripts, 300 and 438, respectively, was dysregulated in the rosettes than in roots. The significantly affected genes play role in response to salt, osmotic and water deprivation stress, in response to pathogens and in ion homeostasis. We hypothesize that the above described changes in gene transcription in A. thaliana resulted from disrupted ionic homeostasis caused by certain ionophore properties of IVM. Our results underlined the negative impact of IVM presence in the environment.

Phytotoxicity of ZnO/kaolinite nanocomposite-is anchoring the right way to lower environmental risk?

The importance of studies on photoactive zinc oxide nanoparticles (ZnO NPs) increases with increasing environmental pollution. Since the ZnO NPs (and NPs in general) also pose an environmental risk, and since an understanding of the risk is still not sufficient, it is important to prevent their spread into the environment. Anchoring on phyllosilicate particles of micrometric size is considered to be a useful way to address this problem, however, so far mainly on the basis of leaching tests in pure water. In the present study, the phytotoxicity of kaolinite/ZnO NP (10, 30, and 50 wt.%) nanocomposites in concentrations 10, 100, and 1000 mg/dm3 tested on white mustard (Sinapis alba) seedlings was found to be higher (relative lengths of roots are ~ 1.4 times lower) compared with seedlings treated with pristine ZnO NPs. The amount of Zn accumulated from the nanocomposites in white mustard tissues was ~ 2 times higher than can be expected based on the ZnO content in the nanocomposites compared with the ZnO content (100 wt.%) in pristine ZnO NPs. For the false fox-sedge (Carex otrubae) plants, the amount of Zn accumulated in roots and leaves was ~ 2.25 times higher and ~ 2.85 times higher, respectively, compared with that of the pristine ZnO NPs (with respect to the ZnO content). Increased phytotoxicity of the nanocomposites and higher uptake of Zn by plants from the nanocomposites in comparison with pristine ZnO NPs suggest that the immobilization of ZnO NPs on the kaolinite does not reduce the environmental risk.

The effect of nanoparticles on the photosynthetic pigments in cadmium-zinc interactions.

Heavy metal contamination, one of the greatest global problems, not only endangers humans and animals but also negatively affects plants. New trends, the production and industrial applications of metals in nanoforms, lead to release of large amounts of nanoparticles into the environment. However, the influence of nanoparticles on living organisms is not well understood. Cadmium is a heavy metal not essential for plants, and to its phytotoxicity also contributes its chemical similarity to zinc. It has been recorded that zinc at low concentrations reduces the toxicity of cadmium, but our results with ZnO nanoparticles did not proved it. In contrast, ZnO nanoparticles significantly increased the negative effect of cadmium, which was reflected mainly in changes in the content of photosynthetic pigments.

Triazolide strigolactone mimics as potent selective germinators of parasitic plant Phelipanche ramosa.

BACKGROUND: Strigolactones are a unique class of plant metabolites which serve as a rhizosphere signal for parasitic plants and evocate their seed germination. The expansion of these parasitic weeds in the food crop fields urgently calls for their increased control and depletion. Simple strigolactone analogues able to stimulate seed germination of these parasitic plants may represent an efficient control measure through the induction of suicidal germination. RESULTS: Triazolide-type strigolactone mimics were easily synthesized in three steps from commercially available materials. These derivatives induced effectively seed germination of Phelipanche ramosa with EC50 as low as 5.2 × 10-10 M. These mimics did not induce seed germination of Striga hermonthica even at high concentration (≥1 × 10-5 M). CONCLUSIONS: Simple and stable strigolactone mimics with selective activity against Phelipanche ramosa were synthesized. © 2019 Society of Chemical Industry.

Metabolism of the anthelmintic drug fenbendazole in Arabidopsis thaliana and its effect on transcriptome and proteome.

Fenbendazole, a broad spectrum anthelmintic used especially in veterinary medicine, may impact non-target organisms in the environment. Nevertheless, information about the effects of fenbendazole in plants is limited. We investigated the biotransformation of fenbendazole and the effect of fenbendazole and its metabolites on gene expression in the model plant Arabidopsis thaliana. High-sensitive UHPLC coupled with tandem mass spectrometry, RNA-microarray analysis together with qPCR verification and nanoLC-MS proteome analysis were used in this study. Twelve fenbendazole metabolites were identified in the roots and leaves of A. thaliana plants. Hydroxylation, S-oxidation and glycosylation represent the main fenbendazole biotransformation pathways. Exposure of A. thaliana plants to 5 µM fenbendazole for 24 and 72 h significantly affected gene and protein expression. The changes in transcriptome were more pronounced in the leaves than in roots, protein expression was more greatly affected in the roots at a shorter period of exposure (24 h) and in leaf rosettes over a longer period (72 h). Up-regulated (>2-fold change, p < 0.1) proteins are involved in various biological processes (electron transport, energy generating pathways, signal transduction, transport), and in response to stresses (e.g. catalase, superoxide dismutase, cytochromes P450, UDP-glycosyltransferases). Some of the proteins which were up-regulated after fenbendazole-exposure probably participate in fenbendazole biotransformation (e.g. cytochromes P450, UDP-glucosyltransferases). Finally, fenbendazole in plants significantly affects many physiological and metabolic processes and thus the contamination of ecosystems by manure containing this anthelmintic should be restricted.

Resorcinol-Type Strigolactone Mimics as Potent Germinators of the Parasitic Plants Striga hermonthica and Phelipanche ramosa.

Strigolactones are a particular class of plant metabolites with diverse biological functions starting from the stimulation of parasitic seed germination to phytohormonal activity. The expansion of parasitic weeds in the fields of developing countries is threatening the food supply and calls for simple procedures to combat these weeds. Strigolactone analogues represent a promising approach for such control through suicidal germination, i.e., parasitic seed germination without the presence of the host causing parasite death. In the present work, the synthesis of resorcinol-type strigolactone mimics related to debranones is reported. These compounds were highly stable even at alkaline pH levels and able to induce seed germination of parasitic plants Striga hermonthica and Phelipanche ramosa at low concentrations, EC50 ≈ 2 × 10-7 M ( Striga) and EC50 ≈ 2 × 10-9 M ( Phelipanche). On the other hand, the mimics had no significant effect on root architecture of Arabidopsis plants, suggesting a selective activity for parasitic seed germination, making them a primary target as suicidal germinators.

Transcriptomic response of Arabidopsis thaliana roots to naproxen and praziquantel.

Exposition to pharmaceutical compounds released to the environment is considered as a potential risk for various organisms. We exposed Arabidopsis thaliana plants to naproxen (NAP) and praziquantel (PZQ) in 5 µM concentration for 2 days and recorded transcriptomic response in their roots with the aim to estimate ecotoxicity and to identify gene candidates potentially involved in metabolism of both compounds. Nonsteroidal anti-inflammatory drug NAP up-regulated 105 and down-regulated 29 genes (p-value ≤ 0.1, fold change ≥ 2), while anthelmintic PZQ up-regulated 389 and down-regulated 353 genes with more rigorous p-value ≤ 0.001 (fold change ≥ 2). High number of up-regulated genes coding for heat shock proteins and other genes involved in response to biotic and abiotic stresses as well as down-regulation of genes involved in processes such as cell proliferation, transcription and water transport indicates serious negative effect of PZQ. NAP up-regulated mostly genes involved in various biological processes and signal transduction and down-regulated mainly genes involved in signal transduction and electron transport or energy pathways. Further, two cytochrome P450s (demethylation) and one methyltransferase (methylation of carboxyl group) were identified as candidates for phase I and several glutathione- and glycosyltransferases (conjugation) for phase II of NAP metabolism. Cytochrome P450s, glutathione and glycosyltransferases seem to play role also in metabolism of PZQ. Up-regulation of several ABC and MATE transporters by NAP and PZQ indicated their role in transport of both compounds.

Synthesis and characterization of isotopically-labeled silver, copper and zinc oxide nanoparticles for tracing studies in plants.

In parallel to technological advances and ever-increasing use of nanoparticles in industry, agriculture and consumer products, the potential ecotoxicity of nanoparticles and their potential accumulation in ecosystems is of increasing concern. Because scientific reports raise a concern regarding nanoparticle toxicity to plants, understanding of their bioaccumulation has become critical and demands more research. Here, the synthesis of isotopically-labeled nanoparticles of silver, copper and zinc oxide is reported; it is demonstrated that while maintaining the basic properties of the same unlabeled ("regular") nanoparticles, labeled nanoparticles enable more sensitive tracing of nanoparticles within plants that have background elemental levels. This technique is particularly useful for working with elements that are present in high abundance in natural environments. As a benchmark, labeled and unlabeled metal nanoparticles (Ag-NP, Cu-NP, ZnO-NP) were synthesized and compared, and then exposed in a series of growth experiments to Arabidopsis thaliana; the NPs were traced in different parts of the plant. All of the synthesized nanoparticles were characterized by TEM, EDS, DLS, ζ-potential and single particle ICP-MS, which provided essential information regarding size, composition, morphology and surface charge of nanoparticles, as well as their stability in suspensions. Tracing studies with A. thaliana showed uptake/retention of nanoparticles that is more significant in roots than in shoots. Single particle ICP-MS, and scanning electron micrographs and EDS of plant roots showed presence of Ag-NPs in particular, localized areas, whereas copper and zinc were found to be distributed over the root tissues, but not as nanoparticles. Thus, nanoparticles in any natural matrix can be replaced easily by their labeled counterparts to trace the accumulation or retention of NPs. Isotopically-labeled nanoparticles enable acquisition of specific results, even if there are some concentrations of the same elements that originate from other (natural or anthropogenic) sources.