Variations in Proline Content, Polyamine Profiles, and Antioxidant Capacities among Different Provenances of European Beech (Fagus sylvatica L.).

International provenance trials are a hot topic in forestry, and in light of climate change, the search for more resilient beech provenances and their assisted migration is one of the challenges of climate-smart forestry. The main aim of the study was to determine intraspecific variability in European beech (Fagus sylvatica L.) among 11 beech provenances according to total antioxidant capacities estimated by various assays, such as DPPH (2,2-diphenyl-1-picrylhydrazyl), ABTS (2,2'-azino-bis-(3-ethylbenzothiazoline-6-sulfonic) acid), FRAP (ferric reducing antioxidant power) assay, and radical scavenging capacity against nitric oxide (RSC-NO assays), as well as osmolyte content, primarily individual polyamines (putrescine, spermidine, and spermine), and free proline content. Polyamine amounts were quantified by using HPLC coupled with fluorescent detection after dansylation pretreatment. The highest values for radical scavenger capacity assays (ABTS, DPPH, and FRAP) were measured in the German provenances DE47 and DE49. Also, the highest NO inhibition capacity was found in the provenance DE49, while the highest content of proline (PRO), total phenolic content (TPC), and total flavonoid content (TFC) was recorded in DE47. The Austrian AT56 and German provenance DE49 were most abundant in total polyamines. This research underlines the importance of the application of common antioxidant assays as well as osmolyte quantification as a criterion for the selection of climate-ready beech provenances for sustainable forest management.

In vitro selection of drought-tolerant white poplar clones based on antioxidant activities and osmoprotectant content.

Introduction: In light of upcoming climate change, there is an urgent requirement for tree improvement regarding adaptability to drought-caused stress and the development of quick and reliable screening methodologies for genotypes' drought tolerance. White poplar is, despite its high adaptability, considered to be an endangered tree species in Serbia, which gives it special importance in the preservation and improvement of biodiversity of riparian ecosystems. The main goal of this research was to evaluate the tolerance of five white poplar clones to the presence of polyethylene glycol (PEG 6000 molecular weight 6000) (different concentrations (e.g. 0 g/L, 1 g/L, 10 g/L, 20 g/L, and 50 g/L) in Aspen Culture Medium (ACM). Methods: The tolerance of the clones was evaluated by using morphological parameters (shoot fresh and dry weight, root fresh and dry weight), photosynthetic pigments (contents of chlorophyll a, chlorophyll b, carotenoids, and chlorophyll a+b), and biochemical parameters (total phenolic content, total flavonoid content, ferric reducing antioxidant power, antioxidant activities (DPPH activity and ABTS assay), free proline content and glycine betaine content. Results and Discussion: The values of morphological and photosynthetic pigments declined with an increase in the concentration of PEG 6000. At a concentration of 50 g/L, the content of shoot fresh mass decreased by 41%, the content of Chl a by 68%, Chl b by 65%, and Car by 76% compared to the control. Also, at the same medium, there was an increase in the content of total phenols, accumulation of proline, the content of glycine betaine as well as in antioxidant activity. Based on the obtained results, it can be assumed that more drought-tolerant clones are characterized by high values for biomass, high content of photosynthetic pigments, and high content of proline and glycine betaine in conditions similar to drought in vitro. Clone L-80 showed better results in most of the tested parameters, especially compared to the reference clone Villafranca.

Metal- and Organ-Specific Response to Heavy Metal-Induced Stress Mediated by Antioxidant Enzymes' Activities, Polyamines, and Plant Hormones Levels in Populus deltoides.

Besides anthropogenic factors, climate change causes altered precipitation patterns that indirectly affect the increase of heavy metals in soils due to hydrological effects and enhanced leaching (i.e., Cd and Ni), especially in the vicinity of mines and smelters. Phytoextraction is a well-known, powerful "green" technique for environmental clean-up that uses plants to extract, sequester, and/or detoxify heavy metals, and it makes significant contributions to the removal of persistent inorganic pollutants from soils. Poplar species, due to their growth features, high transpiration rate, large biomass, and feasible reproduction represent great candidates for phytoextraction technology. However, the consequences of concomitant oxidative stress upon plant metabolism and the mechanism of the poplar's tolerance to heavy metal-induced stress are still not completely understood. In this study, cuttings of poplar species (Populus deltoides W. Bartram ex Marshall) were separately exposed to two heavy metals (Cd2+ and Ni2+) that were triple the maximum allowed amount (MAA) (according to national legislation). The aim of the study was to estimate the effects of heavy metals on: (I) the accumulation of free and conjugated polyamines, (II) plant hormones (including abscisic acid-ABA and indole-3-acetic acid-IAA), and (III) the activities of different antioxidant enzymes at root and leaf levels. By using the selected ion monitoring (SIM) mode of gas chromatography with mass spectrometry (GC/MS) coupled with the isotopically labeled technique, amounts of ABA and IAA were quantified, while polyamine amounts were determined by using high-performance liquid chromatography (HPLC) with fluorometric detection after derivatization. The results showed that P. deltoides responded to elevated concentrations of heavy metals in soils by exhibiting metal- and organ-specific tolerance. Knowledge about tolerance mechanisms is of great importance for the development of phytoremediation technology and afforestation programs for polluted soils.

Ectomycorrhizal Fungi Modulate Pedunculate Oak's Heat Stress Responses through the Alternation of Polyamines, Phenolics, and Osmotica Content.

The physiological and biochemical responses of pedunculate oaks (Quercus robur L.) to heat stress (HS) and mycorrhization (individually as well in combination) were estimated. One-year-old Q. robur seedlings were grown under controlled conditions in a pot experiment, inoculated with a commercial inoculum of ectomycorrhizal (ECM) fungi, and subjected to 72 h of heat stress (40 °C/30 °C day/night temperature, relative humidity 80%, photoperiod 16/8 h) in a climate chamber, and they were compared with seedlings that were grown at room temperature (RT). An in-depth analysis of certain well-known stress-related metrics such as proline, total phenolics, FRAP, ABTS, non-protein thiols, and lipid peroxidation revealed that mycorrhized oak seedlings were more resistant to heat stress (HS) than non-mycorrhized oaks. Additionally, levels of specific polyamines, total phenolics, flavonoids, and condensed tannins as well as osmotica (proline and glycine betaine) content were measured and compared between four treatments: plants inoculated with ectomycorrhizal fungi exposed to heat stress (ECM-HS) and those grown only at RT (ECM-RT) versus non-mycorrhized controls exposed to heat stress (NM-HS) and those grown only at room temperature (NM-RT). In ectomycorrhiza inoculated oak seedlings, heat stress led to not only a rise in proline, total phenols, FRAP, ABTS, non-protein thiols, and lipid peroxidation but a notable decrease in glycine betaine and flavonoids. Amounts of three main polyamines (putrescine, spermine, and spermidine) were quantified by using high-performance liquid chromatography coupled with fluorescent detection (HPLC/FLD) after derivatization with dansyl-chloride. Heat stress significantly increased putrescine levels in non-mycorrhized oak seedlings but had no effect on spermidine or spermine levels, whereas heat stress significantly increased all inspected polyamine levels in oak seedlings inoculated with ectomycorrhizal inoculum. Spermidine (SPD) and spermine (SPM) contents were significantly higher in ECM-inoculated plants during heat stress (approximately 940 and 630 nmol g-1 DW, respectively), whereas these compounds were present in smaller amounts in non-mycorrhized oak seedlings (between 510 and 550 nmol g-1 DW for Spd and between 350 and 450 nmol g-1 DW for Spm). These findings supported the priming and biofertilizer roles of ectomycorrhizal fungi in the mitigation of heat stress in pedunculate oaks by modification of polyamines, phenolics, and osmotica content.

Species-Level Differences in Osmoprotectants and Antioxidants Contribute to Stress Tolerance of Quercus robur L., and Q. cerris L. Seedlings under Water Deficit and High Temperatures.

The general aim of this work was to compare the leaf-level responses of different protective components to water deficit and high temperatures in Quercus cerris L. and Quercus robur L. Several biochemical components of the osmotic adjustment and antioxidant system were investigated together with changes in hormones. Q. cerris and Q. robur seedlings responded to water deficit and high temperatures by: (1) activating a different pattern of osmoregulation and antioxidant mechanisms depending on the species and on the nature of the stress; (2) upregulating the synthesis of a newly-explored osmoprotectant, dimethylsulphoniopropionate (DMSP); (3) trading-off between metabolites; and (4) modulating hormone levels. Under water deficit, Q. cerris had a higher antioxidant capacity compared to Q. robur, which showed a lower investment in the antioxidant system. In both species, exposure to high temperatures induced a strong osmoregulation capacity that appeared largely conferred by DMSP in Q. cerris and by glycine betaine in Q. robur. Collectively, the more stress-responsive compounds in each species were those present at a significant basal level in non-stress conditions. Our results were discussed in terms of pre-adaptation and stress-induced metabolic patterns as related to species-specific stress tolerance features.