Dynamics of Polyamines, Proline, and Ethylene Metabolism under Increasing Cold in Winter Oilseed Rape.

Cold stress is among the most important environmental factors reducing the yield of crops. The present study aimed to investigate the impact of increasing cold stress conditions on winter oilseed rape polyamines, proline, and ethylene metabolism in acclimated and non-acclimated winter oilseed rape. This study was carried out under controlled conditions in the laboratory. The winter oilseed rape hybrid 'Visby' was used in the experiment. Acclimated and non-acclimated plants were subjected to a two-day-long increasing cold (from -1 °C to -3 °C) treatment. HPTLC, RT-qPCR, spectral analysis, and gas chromatography methods were used to analyse the levels of polyamines, gene expression, proline, and ethylene, respectively. This study showed a decrease in putrescine, spermidine, and spermine content during cold acclimation and a decrease in putrescine and spermidine levels at sub-zero temperatures. There were intensive changes in ADC2 gene expression, proline, and ethylene levels in non-acclimated plants: a substantial increase after exposure to -1 °C temperature and a sharp decrease after exposure to -3 °C temperature. The changes in these parameters were lower or absent in acclimated plants. The phenomena observed in this study add new insights to the knowledge about the plant stress response and suggest questions to be answered in the future.

Proline Enhances Resistance and Recovery of Oilseed Rape after a Simulated Prolonged Drought.

This study was carried out to evaluate the effect of exogenous proline on the growth, biochemical responses, and plant recovery of drought-stressed oilseed rape plants after renewed irrigation. The experiment was conducted under controlled laboratory conditions. After 21 days of cultivation, 3-4 leaf stage seedlings were sprayed with proline (1 mM), then subjected to prolonged drought stress for 8 days to achieve a severe water deficit, next, irrigation was resumed and recovery was assessed after 4 days. The results show that exogenous application of proline reduced the drought-induced growth inhibition of seedlings while maintaining relative water content (RWC) and growth parameters closer to those of irrigated plants. Proline had a positive effect on chlorophyll accumulation and membrane permeability while decreasing ethylene, H2O2, and MDA levels. Moreover, after 4 days of recovery, the H2O2 content of the proline-treated plants was significantly lower (2-fold) and the MDA content was close to that of continuously irrigated plants. Thus, all these biochemical reactions influenced plant survival: after drought + proline treatment, the number of surviving plants was two times higher than that of drought-treated plants. The findings show that exogenous proline has antioxidant, osmotic, and growth-promoting properties that improve the drought tolerance of winter oilseed rape plants and is, therefore, beneficial for drought adaptation in oilseed rape.

Probiotics, Proline and Calcium Induced Protective Responses of Triticum aestivum under Drought Stress.

In order to increase plants tolerance to drought, the idea of treating them with stress-protecting compounds exogenously is being considered. In this study, we aimed to evaluate and compare the impact of exogenous calcium, proline, and plant probiotics on the response of winter wheat to drought stress. The research was carried out under controlled conditions, simulating a prolonged drought from 6 to 18 days. Seedlings were treated with ProbioHumus 2 µL g-1 for seed priming, 1 mL 100 mL-1 for seedling spraying, and proline 1 mM according to the scheme. 70 g m-2 CaCO3 was added to the soil. All tested compounds improved the prolonged drought tolerance of winter wheat. ProbioHumus, ProbioHumus + Ca had the greatest effect on maintaining the relative leaf water content (RWC) and in maintaining growth parameters close to those of irrigated plants. They delayed and reduced the stimulation of ethylene emission in drought-stressed leaves. Seedlings treated with ProbioHumus and ProbioHumus + Ca had a significantly lower degree of membrane damage induced by ROS. Molecular studies of drought-responsive genes revealed substantially lower expression of Ca and Probiotics + Ca treated plants vs. drought control. The results of this study showed that the use of probiotics in combination with Ca can activate defense reactions that can compensate for the adverse effects of drought stress.

The Physiological Responses of Wheat and Maize Seedlings Grown under Water Deficit Are Modulated by Pre-Application of Auxin-Type Plant Growth Regulators.

The physiological responses of wheat and maize seedlings to exogenous auxin-type compounds 1-[2-chloroethoxycarbonyl-methyl]-4-naphthalenesulfonic acid calcium salt (TA-12) and 1-[2-dimethylaminoethoxicarbonylmethyl]naphthalene chlormethylate (TA-14) application prior to polyethyleneglycol-6000 (PEG) treatment were studied. PEG treatment inhibited seedlings growth and caused alterations in their antioxidant defence which was crop-specific. PEG increased the non-enzymatic antioxidants along with inhibition of enzymatic antioxidant activity in wheat, while in maize the opposite effects were found. The TA-12 and TA-14 applied alone increased most of the growth parameters measured in both crops, as well as the catalase activity and protein content of wheat. The growth of PEG-treated wheat and maize plants was improved by foliar spray with TA-compounds (TAs). Application of TAs before PEG treatment maintained low-molecular weight thiol-containing compounds and protein contents, and catalase and peroxidase activities close to the control levels. This was better expressed in maize than in wheat seedlings. The results showed that the preliminary application of TA-12 and TA-14 can reduce the adverse effects of moderate water deficit by crop-specific adjustment of the antioxidant defence to counteract stress.

The Influence of End-of-Day Blue Light on the Growth, Photosynthetic, and Metabolic Parameters of Lettuce at Different Development Stages.

This study evaluates the effect of end-of-day blue (EOD B) light on the physiological response of lettuce (Lactuca sativa, Lobjoits Green Cos) at different phenological development stages. Plants were grown in a controlled environment growth chamber (day/night temperature 21 ± 2 °C; relative air humidity 60 ± 5%) under the light of light-emitting diodes (LEDs) consisting of 5% blue (B; 450 nm), 85% red (R; 660 nm), and 10% green (G; 530 nm) photosynthetic photon flux density (PPFD) at 200 µmol m-2 s-1 for 16 h d-1 (BRG, control) for 8, 15, and 25 days (BBCH 12, BBCH 14, and BBCH 18, respectively). For the EOD B treatments, lettuce plants were additionally illuminated with 100% of B light at 30 and 60 µmol m-2 s-1 PPFD for 4 h d-1 (B30 and B60, respectively). The results show that EOD B light caused the elevated shoot elongation of lettuce plants regardless of their growth stages. However, leaf width increased only in more developed lettuce plants (BBCH 18). EOD B light negatively affected the development of new leaves and fresh weight, except for seedlings (BBCH 12). Most photosynthetic and spectral leaf indices also decreased when lettuce was treated with EOD B light, especially under the PPFD level of 60 µmol m-2 s-1. Moreover, the changes in metabolic parameters such as DPPH free radical activity, free proline content, and H+-ATPase activity in lettuce showed a plant response to unfavorable conditions to EOD B light.

The Application of Auxin-like Compounds Promotes Cold Acclimation in the Oilseed Rape Plant.

Cold is a major environmental key factor influencing plant growth, development, and productivity. Responses and adaption processes depend on plant physiological and biochemical modifications, first of all via the hormonal system. Indole-3-acetic acid (IAA) plays a critical role in the processes of plant functioning. To assess the influence of the auxin-like compounds 1-[2-chloroethoxycarbonylmethyl]-4-naphthalenesulfonic acid calcium salt (TA-12) and 1-[2-dimethylaminoethoxycarbonylmethyl]naphthalene chloromethylate (TA-14) in the process of cold acclimation, long-term field trials over four years were performed with two rapeseed (Brassica napus L.) plant cultivars with different wintering resistance in temperate-zone countries. In these two rapeseed cultivars, namely 'Casino' (less resistant) and 'Valesca' (more resistant), investigations were conducted in the terminal buds and root collars. The application of auxin-like compounds revealed a close interlinkage between the composition of dehydrins and the participation of the phytohormone IAA in the adaptation processes. By applying TA-12 and TA-14, the importance of the proteins, especially the composition of the dehydrins, the IAA amount, and the status of the oilseed rape cultivars at the end of the cold acclimation period were confirmed. Following on from this, when introducing oilseed rape cultivars from foreign countries, it may also be of value to assess their suitability for cultivation in temperate-zone countries.

Probiotics Enhance Cereal Yield and Quality and Modify Agrochemical Soil Properties.

The aim of this study was to determine the influence of microbial biostimulants on wheat and oat growth, grain yield, and grain quality and to evaluate the influence of these probiotics on some soil agrochemical traits in the open field. Active concentrations of ProbioHumus and NaturGel and their mixtures were selected under laboratory conditions using winter wheat as a reference plant. Probiotics had a biostimulating effect on the development of the underground and aboveground part of winter wheat when 2 µL/g was used for seed priming and 2 mL/100 mL for seedling spraying. Under field conditions, after treatment of soil (2 L/ha), wheat and oat seeds (2 L/t), and plants (2 L/ha) with ProbioHumus and NaturGel, it was found that the yield of the studied cereals increased, on average, by 0.50 t/ha to 1.09 t/ha. ProbioHumus promoted protein accumulation in the investigated cereal grains. The level of microelements in wheat and oat grains increased after treatment of plants with NaturGel. Probiotics improved soil agrochemical properties, such as total and nitrate nitrogen, total and available phosphorus, organic carbon, humic acid, and humus content. In conclusion, plant probiotics can be used as an ecological alternative for growing cereals and improving the agrochemical properties of the soil.

Effects Induced by the Agricultural Application of Probiotics on Antioxidant Potential of Strawberries.

With the recent rapid development of the functional food sector, agriculture is looking for alternatives to improve the quality of food grown by limiting chemical fertilizers. This study evaluated the effects of two commercial plant probiotics, ProbioHumus and NaturGel, on the growth and quality of strawberry fruits. Strawberry plants were sprayed with microbial probiotics twice a year: after harvesting at the beginning of dormancy and at the stage of leaf development. Spray applications of ProbioHumus, NaturGel, and NaturGel + ProbioHumus in the organic farm fields significantly increased the fresh fruit weight up to 42%, 35%, and 37%, respectively, compared to the non-treated control. An increase in the weight of fresh strawberry fruits may be associated with an increase in dry matter accumulation. The probiotics had a positive effect on the total content of phenols, anthocyanins, and especially ascorbic acid in strawberry fruits. The increase in ascorbic acid in strawberry fruits was up to 97% compared to the non-treated control. The fruits from plants inoculated with probiotics showed significantly higher antioxidant activity. In summary, ProbioHumus and NaturGel are effective tools for improving the quality of strawberries and can be exploited in sustainable agriculture as a tool for adding value to functional food.

Gibberellic Acid (GA3) Applied to Flowering Heracleum sosnowskyi Decreases Seed Viability Even If Seed Development Is Not Inhibited.

Sosnowsky's hogweed (Heracleum sosnowskyi Manden.), an important invasive species in Eastern Europe, is a monocarpic perennial plant that propagates exclusively by seeds. Hence, interfering with seed viability could help control its spread. In the present study, we investigated the effect of exogenous GA3 (25, 100 and 150 mg/L) sprayed twice onto flowering H. sosnowskyi plants on the development of fruits (mericarps) and their ability to germinate under field conditions over the growing seasons of 2018 and 2019. Mericarps from plants sprayed with GA3 failed to develop normally. The width/length ratio of mericarps decreased by 23% to 25% after 150 mg/L GA3 application and their average weight decreased between 7% and 39% under all GA3 treatments. X-ray radiographs revealed that the internal structure was malformed, with many of the mericarps lacking well-developed seeds. Proportionally fewer well-developed mericarps were produced by GA3-treated plants than water-sprayed control plants in 2018. Seed germination assessed outdoors in seeds buried in the ground was also severely reduced (from 58% to 99% after 150 mg/L GA3 application). This indicates that exogenous GA3 sprays result in incomplete seed development and a consequent decrease in viability and germination. As the highest GA3 dose used resulted in significantly reduced propagation of Sosnowsky's hogweed through seeds in the field, GA3 provides a promising approach to the control of the spread of this invasive weed species.

Effects of Elevated Temperature on Root System Development of Two Lupine Species.

The aim of this study was to assess the effect of elevated temperature on the growth, morphology and spatial orientation of lupine roots at the initial stages of development and on the formation of lupine root architecture at later stages. Two lupine species were studied-the invasive Lupinus polyphyllus Lindl. and the non-invasive L. luteus L. The plants were grown in climate chambers under 25 °C and simulated warming at 30 °C conditions. The angle of root curvature towards the vector of gravity was measured at the 48th hour of growth, and during a 4-h period after 90° reorientation. Root biometrical, histological measurements were carried out on 7-day-old and 30-day-old plants. The elevation of 5 °C affected root formation of the two lupine species differently. The initial roots of L. polyphyllus were characterized by worse spatial orientation, reduced growth and reduced mitotic index of root apical meristem at 30 °C compared with 25 °C. The length of primary roots of 30-day-old lupines and the number of lateral roots decreased by 14% and 16%, respectively. More intense root development and formation were observed in non-invasive L. luteus at 30 °C. Our results provide important information on the effect of elevated temperature on the formation of root architecture in two lupine species and suggest that global warming may impact the invasiveness of these species.

Plant Microbial Biostimulants as a Promising Tool to Enhance the Productivity and Quality of Carrot Root Crops.

The interest in studies focused on applying probiotic microorganisms is increasing due to sustainable agriculture development. In this research, we aimed to evaluate the impact of two commercial plant probiotics-ProbioHumus and NaturGel on carrot growth, yield, and quality in organic and nonorganic production systems. The research was carried out under laboratory and field conditions. Plants were treated with probiotics (2 L/ha) at the nine leaves stage. Biometrical measurements and chemical analyses were performed at a maturation stage. The average weight of carrot roots increased by 17 and 20 g in the test variant with ProbioHumus as compared to the control in the organic and nonorganic farms, respectively. Plant microbial biostimulants ProbioHumus and NaturGel had a positive effect on the quality of carrots from organic and nonorganic farms: applied in couple they promoted the accumulation of monosaccharides, ascorbic acid, carotenoids, phenols, and increased antioxidant activity. Quantitative nitrate analysis regardless of the biostimulant used revealed about twofold lower nitrate content of carrots from organic than nonorganic farms, and probiotics did not show a significant effect on nitrate accumulation. Finally, ProbioHumus and NaturGel were effective at low doses. The use of microbial biostimulants can be recommended as an element of cultivation for creating ecologically friendly technologies.

Elevated Temperature Induced Adaptive Responses of Two Lupine Species at Early Seedling Phase.

This study aimed to investigate the impact of climate warming on hormonal traits of invasive and non-invasive plants at the early developmental stage. Two different lupine species-invasive Lupinus polyphyllus Lindl. and non-invasive Lupinus luteus L.-were used in this study. Plants were grown in climate chambers under optimal (25 °C) and simulated climate warming conditions (30 °C). The content of phytohormone indole-3-acetic acid (IAA), ethylene production and the adaptive growth of both species were studied in four-day-old seedlings. A higher content of total IAA, especially of IAA-amides and transportable IAA, as well as higher ethylene emission, was determined to be characteristic for invasive lupine both under optimal and simulated warming conditions. It should be noted that IAA-L-alanine was detected entirely in the invasive plants under both growth temperatures. Further, the ethylene emission values increased significantly in invasive lupine hypocotyls under 30 °C. Invasive plants showed plasticity in their response by reducing growth in a timely manner and adapting to the rise in temperature. Based on the data of the current study, it can be suggested that the invasiveness of both species may be altered under climate warming conditions.

Biomarker identification of isolated compartments of the cell wall, cytoplasm and vacuole from the internodal cell of characean Nitellopsis obtusa.

Cells of characean algae are attractive for plant cell physiologists because of their large size and their close relation to higher plant cells. The objective of our study was to evaluate the purity of the compartments (cell wall, cytoplasm with plastids, mitochondria, nuclei and endomembrane system, and vacuole) separated mechanically from the internodal cells of Nitellopsis obtusa using enzymatic markers. These included α-mannosidase and malate dehydrogenase, vacuolar and cytoplasmic enzymes, respectively. The biomarkers applied revealed the degree of compartment contamination with the material from unwanted cell parts. The cell wall was contaminated slightly by vacuole and cytoplasm residuals, respectively by 12.3 and 1.96% of corresponding biomarker activities. Relatively high activity of vacuolar marker in the cell wall could be associated with the cell vacuoles in the multicellular structure of the nodes. The biomarkers confirmed highly purified vacuolar (99.5%) and cytoplasmic (86.7%) compartments. Purity estimation of the cell fractions enabled reevaluating nCuO related Cu concentrations in the compartments of charophyte cell. The internalisation of CuO nanoparticles in N. obtusa cell occurred already after 0.5h. In general, the approach seems to be useful for assessing the accumulation and distribution of various xenobiotics and/or metabolites within plant cell. All this justifies N.obtusa internodal cells as a model organism for modern studies in cell biology and nanotoxicology.

Modifying effects of leaf litter extracts from invasive versus native tree species on copper-induced responses in Lemna minor.

Invasive plant species tend to migrate from their native habitats under favourable climatic conditions; therefore, trophic and other relationships in ecosystems are changing. To investigate the effect of natural organic matter derived from native Alnus glutinosa tree species and from invasive in Lithuania Acer negundo tree species on copper toxicity in Lemna minor, we analysed the dynamics of Cu binding in aqueous leaf litter extracts (LLE) and plant accumulation, morphophysiological parameters, and antioxidative response. The results revealed that A. glutinosa LLE contained polyphenols (49 mg pyrogallol acid equivalent (PAE)/g DM) and tannins (7.5 mg PAE/g DM), while A. negundo LLE contained only polyphenols (23 mg PAE/g DM). The ability of LLE to bind Cu increased rapidly over 1.5-3 h to 61% and 49% of the total Cu concentration (6.0 ± 0.9 mg/L), respectively for A. glutinosa (AG) and A. negundo (AN), then remained relatively stable until 48 h. At the same time, L. minor accumulated 384, 241 or 188 µg Cu/g FW when plants were exposed to Cu (100 µM CuSO4), Cu with 100 mg/L dissolved organic carbon (DOC) from either AG LLE or AN LLE, accordingly. Catalase (CAT) and guaiacol peroxidase (POD) played a dominant role in hydrogen peroxide scavenging when plants were exposed to Cu and 10 or 100 mg/L DOCAG mixtures in both the first (up to 6h) and the second (6-48 h) response phases. Due to functioning of oxidative stress enzymes, the levels of the lipid peroxidation product malondialdehyde (MDA) reduced in concentration-dependent manner, compared to Cu treatment. When combining Cu and DOCAN treatments, the most sensitive enzymes were POD, ascorbate peroxidase and glutathione reductase. Their activities collectively with CAT were sufficient to reduce MDA levels to Cu-induced in the initial, but not the second response phase. These data suggest that leaf litter extracts of different phenolic compositions elicited different antioxidant response profiles resulting in different reductions of Cu stress, thus effecting L. minor frond and root development observed after seven days. The complex data from this study may be useful in modelling the response of the aquatic ecosystem to a changing environment.

Foliar Application of Polyamines Modulates Winter Oilseed Rape Responses to Increasing Cold.

Cold stress is one of the most common abiotic stresses experienced by plants and is caused by low temperature extremes and variations. Polyamines (PAs) have been reported to contribute in abiotic stress defense processes in plants. The present study investigates the survival and responses of PA-treated non-acclimated (N) and acclimated (A) winter oilseed rape to increasing cold conditions. The study was conducted under controlled conditions. Seedlings were foliarly sprayed with spermidine (Spd), spermine (Spm), and putrescine (Put) solutions (1 mM) and exposed to four days of cold acclimation (4 °C) and two days of increasing cold (from -1 to -3 °C). Two cultivars with different cold tolerance were used in this study. The recorded traits included the percentage of survival, H+-ATPase activity, proline accumulation, and ethylene emission. Exogenous PA application improved cold resistance, maintained the activity of plasma membrane H+-ATPase, increased content of free proline, and delayed stimulation of ethylene emission under increasing cold. The results of the current study on winter oilseed rape revealed that foliar application of PAs may activate a defensive response (act as elicitor to trigger physiological processes), which may compensate the negative impact of cold stress. Thus, cold tolerance of winter oilseed rape can be enhanced by PA treatment.

Heracleum sosnowskyi seed development under the effect of exogenous application of GA3.

Numerous studies have demonstrated the impact of exogenous gibberellin on fleshy fruit formation, but the effect on dry fruits is not yet well known. To test the role of gibberellin (GA3) in dry fruit formation, we analysed the impact of exogenous GA3 on the invasive plant Sosnowsky's hogweed (H. sosnowskyi Manden.) seed development and germination. Treatment of GA3 concentrations of 0.07 mM, 0.14 mM, 0.28 mM, 0.43 mM was applied to flowers at the early stage of development. Seeds were collected from treated satellite umbels. It was observed that GA3treatment did not have a significant effect on the size of H. sosnowskyi seeds, but caused various changes in their shape. The data on semi-thin longitudinal sections of H. sosnowskyi mericarps and SEM micrographs of embryos showed that the embryos in GA3 (0.43 mM) treated variants were at torpedo stage, while in control variants-mature embryos. The germination of seeds of each variant was estimated by burying them in the soil. Our studies indicated that GA3 application reduced the germination of H. sosnowskyi seed from 98.0% (control) to 16.5% (GA3 concentration 0.43 mM). It was assumed that exogenous application of GA3 had influence on the development of dry Sosnowsky's hogweed seeds and could be used to inhibit the spread of this invasive plant.

Ecotoxicity effects triggered in aquatic organisms by invasive Acer negundo and native Alnus glutinosa leaf leachates obtained in the process of aerobic decomposition.

The replacement of autochthonous tree species by invasive ones in coastal zones of freshwater bodies induces additional alteration of hydrochemical and microbiological characteristics due to decomposition of fallen leaves of non-indigenous species, which can lead to ecotoxic response of the littoral biota. Leaves of invasive to Lithuania boxelder maple (Acer negundo) and autochthonous black alder (Alnus glutinosa) lost more than half of biomass and released stable amount of DOC (60-70 mg/L) throughout 90-day mesocosm experiment under aerobic conditions. This, along with the relatively small BOD7 values detected after some variation within the first month confirms effective biodegradation by fungi and bacteria. The ambient water was more enriched with different forms of N and P by decomposing boxelder maple than by alder leaves. During the first month, both leachates were more toxic to charophyte (Nitellopsis obtusa) at mortality and membrane depolarization levels, while later to two crustacean species. Biomarker response, H(+)-ATPase activity in membrane preparations from N. obtusa, was stronger for A. negundo. Generally, boxelder maple leaf leachates were more toxic to tested hydrobionts and this coincides with previous study on leaves of the same pair of tree species conducted under microaerobic conditions (Krevs et al., 2013).

Ecotoxicological effects evoked in hydrophytes by leachates of invasive Acer negundo and autochthonous Alnus glutinosa fallen off leaves during their microbial decomposition.

Throughout 90-day biodegradation under microaerobic conditions, invasive to Lithuania species boxelder maple (Acer negundo) leaves lost 1.5-fold more biomass than that of autochthonous black alder (Alnus glutinosa), releasing higher contents of N(tot), ammonium and generating higher BOD(7). Boxelder maple leaf leachates were characterized by higher total bacterial numbers and colony numbers of heterotrophic and cellulose-decomposing bacteria than those of black alder. The higher toxicity of A. negundo aqueous extracts and leachates to charophyte cell (Nitellopsis obtusa), the inhabitant of clean lakes, were manifested at mortality and membrane depolarization levels, while the effect on H(+)-ATPase activity in membrane preparations from the same algae was stronger in case of A. glutinosa. Duckweed (Lemna minor), a bioindicator of eutrophic waters, was more sensitive to leaf leachates of A. glutinosa. Fallen leaves and leaf litter leachates from invasive and native species of trees, which enter water body, affect differently microbial biodestruction and aquatic vegetation in freshwater systems.

Toxicity of copper oxide nanoparticle suspensions to aquatic biota.

Toxicity effects induced by nanosuspensions of CuO (<50 nm; Sigma-Aldrich) on macrophytic algae cells of Nitellopsis obtusa (96-h median lethal concentration [LC50]), microphytic algae Chlorella (30-min median inhibitory concentration [IC50]), shrimp Thamnocephalus platyurus (24-h LC50), and rotifer Brachionus calyciflorus (24-h LC50) were investigated. No substantial differences between the effects of nonsonicated and sonicated nCuO suspensions were observed. The particle size distribution analysis accomplished by the laser diffraction technique at suspension concentration from 3 to 100 mg/L revealed rapid (within 5 min) reagglomeration of the particles after the sonication. The observed adverse effects on N. obtusa cells may be attributed to nanoparticles per se, but not to ionic Cu, because neither chemical analysis nor biological testing (algae survival in the supernatants of suspensions) confirmed the presence of cupric ions in toxic amounts. Contrary to ionic Cu form, nCuO delayed the initial phase of N. obtusa cell membrane depolarization. Lethality tests with rewash demonstrated that the least used 5-min exposure in 100 mg/L nCuO sonicated suspension induced 70% mortality in charophyte cells after 8 d, whereas the rewash after a short exposure to a noticeably toxic concentration of Cu(2+) prevented cell mortality. The obtained data suggested the possible influence of a thick charophyte cell wall on the dynamics of nanotoxicity effects.

Changes in plasmalemma K+Mg2+ -ATPase dephosphorylating activity and H+ transport in relation to seasonal growth and freezing tolerance of Festuca pratensis Huds.

Changes in plasmalemma K(+)Mg(2+)-ATPase dephosphorylating activity and H(+) transport were examined in freezing-tolerant and non-tolerant genotypes of the perennial grass species Festuca pratensis Huds. Enzyme activity and DeltamuH(+) were measured in plasmalemma fractions isolated from basal nodes and roots. Three types of experiments were undertaken: (i) a field experiment, utilizing the seasonal growth and cessation cycle of a perennial plant; (ii) a cold acclimation experiment in hydroponics; and (iii) an instant freezing test. A specific fluctuation in K(+)Mg(2+)-ATPase activity was found throughout the seasonal growth of the plants (i). The K(+)Mg(2+)-ATPase activity peaks for both the basal node and the root plasmalemma were determined early in the spring before the renewal of growth. The lowest activity values in roots occurred at the time approaching flowering, and in basal nodes at the transition into the growth cessation. The K(+)Mg(2+)-ATPase activity was approximately 50% lower in the basal node plasmalemma of freezing-tolerant plants than of non-tolerant ones, when assessed at the optimal growth stage in hydroponics. In hydroponics (ii) and in the freezing test (iii), temperature stress was followed by a more pronounced change in the level of K(+)Mg(2+)-ATPase activity than in that of H(+) transport, and this change was more clearly differentiated in the basal node plasmalemma of contrasting genotypes than in the roots. Stress response was manifested differently in freezing-tolerant and non-tolerant plants at cold acclimation (4-2 degrees C) and at freezing (-8 degrees C) temperatures. Proton transport regulation via coupled changes in the hydrolysed ATP/transported proton ratio, as an attribute of freezing-tolerant plants, is discussed.