Cellular Electrolyte Leakage Research Articles

Elucidating the underlying mechanisms of silicon to suppress the effects of nitrogen deficiency in pepper plants

In many regions, nitrogen (N) deficiency limits pepper cultivation, presenting significant cultivation challenges. This study investigates the impact of N deficiency and silicon (Si) supplementation on physiological responses and antioxidant modulation in pepper plants, focusing particularly on the homeostasis of carbon (C), nitrogen, and phosphorus (P), and their effects on growth and biomass production. Conducted in a factorial design, the experiment examined pepper plants under conditions of N sufficiency and deficiency, with and without Si supplementation (0.0 mM and 2.0 mM). Results showed that N deficiency sensitizes pepper plants, leading to increased electrolyte leakage (39.59%) and disrupted C, N, and P homeostasis. This disruption manifests as reductions in photosynthetic pigments (−64.53%), photochemical efficiency (−14.92%), and the synthesis of key metabolites such as total free amino acids (−86.97%), sucrose (−53.88%), and soluble sugars (−39.96%), ultimately impairing plant growth. However, Si supplementation was found to alleviate these stresses. It modulated the antioxidant system, enhanced the synthesis of ascorbic acid (+30.23), phenolic compounds (+33.19%), and flavonoids (+7.52%), and reduced cellular electrolyte leakage (−25.02%). Moreover, Si helped establish a new homeostasis of C, N, and P, optimizing photosynthetic and nutritional efficiency by improving the utilization of C (+17.46%) and N (+13.20%). These Si-induced modifications in plant physiology led to increased synthesis of amino acids (+362.20%), soluble sugars (+51.34%), and sucrose (77.42%), thereby supporting enhanced growth of pepper plants. These findings elucidate the multifaceted biological roles of Si in mitigating N deficiency effects, offering valuable insights for more sustainable horticultural practices.

Boosting biopesticide potential of Trichoderma harzianum for controlling the downy mildew and improving the growth and the productivity of King Ruby seedless grape

BackgroundPlasmopara viticola, the causal agent of grape downy mildew, is one of the most serious grape diseases. Application of Trichoderma harzianum grown on different potato dextrose supplemented broth media using some chemical inducers (TSDCIS), i.e., thiamine (T2), a mixture of micronutrients (T3) and potassium tartrate (T4), compared to Trichoderma without amendment (T1), was conducted under field conditions to evaluate the potential of T. harzianum against grape downy mildew, improving the growth, as well as the yield quality, and quantity.ResultsFoliar application of all TSDCIS significantly reduced the disease severity and increased the phenolic content, peroxidase, polyphenol oxidase enzyme activity, growth parameters, and yield parameters. Trichoderma growing on medium supplemented with potassium tartrate (T4) achieved the most significant reduction in the disease severity (78.9, 81.8%) than the control treatment in two growing seasons, respectively. In contrast, it decreased H2O2 content, lipid peroxidation, and cellular electrolyte leakage. Scanning electron microscopy observations revealed that the number of opened stomata, average stomatal area, and stomatal pore area decreased significantly in response to T4 treatment.ConclusionsIt can be concluded that using Trichoderma growing on medium supplemented with potassium tartrate (T4) to biosafety control the downy mildew disease of grape and improve its growth, yield, and fruit quality is recommended.

Kaolin triggers defense-responsive genes in grapevines (cv. King Ruby seedless) to downy mildew and enhances its vegetative development, production, and berries quality standards

Plasmopara viticola, the causative of downy mildew, is one of the most destructive pathogens that attack grapevines worldwide. Despite to its high productivity and good berries quality, King Ruby seedless grapevines are highly susceptible to P. viticola and suffer its adverse effects. In this study, kaolin spraying at 1 and 3% was investigated under field conditions as a likely anti-oomycete substance. Results from the field experiment showed that kaolin spraying at 3% resulted in a considerable reduction in the infection severity (80.1%). Overexpression of the responsive factor JERF3 (4-fold), and the defense-responsive genes PR1 (4.2-fold), CHI II (3.9-fold), POD (2.7-fold), and GLU (2.8-fold) was recorded. Furthermore, a promotion in the phenolic compounds in plant roots (106.5%), as well as activities of peroxidase (182.5%) and polyphenol oxidase (136.4%) enzymes was also recorded in response to this treatment. A decrease in lipid peroxidation, H2O2 content, and the cellular electrolyte leakage, as well as an increment in ascorbic acid content in grapevine leaves were observed. Scanning electron microscopy showed that kaolin treatment at 3% induced stomatal closure and led to a reduction in the stomatal area in grapevines leaves. Transmission electron microscopy indicated abnormal variations in the ultrastructure of the infected mesophyll cells of grapevine leaf. This abnormality decreased due to kaolin application at 3%. Moreover, kaolin spraying resulted in a significant enhancement in most of the evaluated growth parameters. Yield per grapevine was also improved (18.5%), as well as all evaluated berries quality standards in response to kaolin application. It can be concluded that kaolin application at 3% is a promising protective solution to control downy mildew and improve the grapevines growth and production.

Foliar-applied nanoscale zero-valent iron (nZVI) and iron oxide (Fe3O4) induce differential responses in growth, physiology, antioxidative defense and biochemical indices in Leonurus cardiaca L

The impacts of nZVI and iron oxides on growth, physiology and elicitation of bioactive antioxidant metabolites in medicinal aromatic plants must be critically assessed to ensure their safe utilization within the food chain and achieve nutritional gains. The present study investigated and compared the morpho-physiological and biochemical changes of Leonurus cardiaca L. plants as affected by various concentrations (0, 250, 500 and 1000 mg L−1) of nZVI and Fe3O4. The foliar uptake of nZVI was verified through Scanning Electron Microscopy (SEM) images and Energy Dispersive X-ray (EDX) analytical spectra. Plants exposed to nZVI at low concentration showed comparatively monotonic deposition of NPs on the surface of leaves, however, the agglomerate size of nZVI was raised as their doses increased, leading to remarkable changes in anatomical and biochemical traits. 250 mg L−1 nZVI and 500 mg L−1 Fe3O4 significantly (P < 0.05) increased plant dry matter accumulation by 37.8 and 27% over the control, respectively. The treatments of nZVI and Fe3O4 at 250 mg L−1 significantly (P < 0.01) improved chlorophyll a content by 22.4% and 15.3% as compared to the control, and then a rapid decrease (by 14.8% and 4.1%) followed at 1000 mg L−1, respectively. Both nZVI and Fe3O4 at 250 mg L−1 had no significant impact on malondialdehyde (MDA) formation, however, at an exposure of 500–1000 mg L−1, the MDA levels and cellular electrolyte leakage were increased. Although nZVI particles could be utilized by plants and enhanced the synthesis of chlorophylls and secondary metabolites, they appeared to be more toxic than Fe3O4 at 1000 mg L−1. Exposure to nZVI levels showed positive, negative and or neutral impacts on leaf water content compared to control, while no significant difference was observed with Fe3O4 treatments. Soluble sugar, total phenolics and hyperoside content were significantly increased upon optimum concentrations of employed treatments-with 250 mg L−1 nZVI being most superior. Among the extracts, those obtained from plants treated with 250–500 mg L−1 nZVI revealed the strong antioxidant activity in terms of scavenging free radical (DPPH) and chelating ferrous ions. These results suggest that nZVI (at lower concentration) has alternative and additional benefits both as nano-fertilizer and nano-elicitor for biosynthesis of antioxidant metabolites in plants, but at high concentrations is more toxic than Fe3O4.

Silica Nanoparticles as a Probable Anti-Oomycete Compound Against Downy Mildew, and Yield and Quality Enhancer in Grapevines: Field Evaluation, Molecular, Physiological, Ultrastructural, and Toxicity Investigations.

Downy mildew is the most destructive disease of grapevines in the regions of relatively warm and humid climate causing up to 50% yield losses. Application of silicon- (Si-) based products have been extensively studied against various oomycete, fungal, bacterial, and viral plant diseases, but studies on Si application in their nanosize are limited. In this study, the field application of silica nanoparticles (SiNPs) on Thompson Seedless grapevines (H4 strain) infected with downy mildew was evaluated. In addition, molecular, physiological, ultrastructural, and toxicity investigations were also conducted. The obtained results revealed that spraying of grapevines with SiNPs at 150 ppm significantly overexpressed the transcription factor jasmonate and ethylene-responsive factor 3 recording 8.7-fold, and the defense-related genes β-1,3-glucanase (11-fold), peroxidase (10.7-fold) pathogenesis-related-protein 1 (10.6-fold), and chitinase (6.5-fold). Moreover, a reduction up to 81.5% in the disease severity was achieved in response to this treatment. Shoot length and yield per grapevine were considerably enhanced recording up to 26.3 and 23.7% increase, respectively. The berries quality was also improved. Furthermore, this treatment led to an enhancement in the photosynthetic pigments, induction of phenolic and ascorbic acid contents, an increase in the activity of peroxidase and polyphenol oxidase enzymes, and a reduction in the cellular electrolyte leakage, lipid peroxidation, and H2O2 content. Scanning electron microscopy observations showed an increase up to 86.6% in the number of closed stomata and a reduction up to 55% in the average stomatal pore area in response to this treatment. Observations of the transmission electron microscopy showed ultrastructural alterations in the cells of a grapevine leaf due to the infection with downy mildew, including plasmolysis and disruption of the cellular components, abnormal chloroplasts, and thickening of the cell wall and cell membrane. These abnormal alterations were reduced in response to SiNPs spray. In contrast, this study also showed that this treatment had considerable cytotoxic and genotoxic effects at this direct dose/concentration. So, additional investigations to determine the SiNPs residue in the produced edible plant parts are urgently needed. In addition, the pre-harvest interval, toxicity index, and risk assessment should be evaluated before any recommendation for use.

Nano-fertilizers and their impact on vegetables: Contribution of Nano-chelate Super Plus ZFM and Lithovit®-standard to improve salt-tolerance of pepper

The implementation of nano-technology for the amelioration of plant tolerance against abiotic stress has been widely reported in last years. In the current study, the separate and combined effects of Nano-chelate Super Plus ZFM (A) and Lithovit®-standard (B) on salt-stressed chili pepper were evaluated. Two different concentrations of each products were tested (A1, 2.5 g L−1, A2, 5 g L−1; B1, 3 g L−1 and B2, 5 g L−1) on pepper irrigated by three NaCl solutions (1.5dS m−1, 3dS m−1 and 6dS m−1. The control (no treatment) experiments were only irrigated with the three NaCl solutions. Spraying Nano-chelate Super Plus ZFM in low concentration (A1) enhanced significantly iron, zinc and manganese content in shoots and fruits. The application of Lithovit®-standard at high concentration ameliorated calcium and magnesium content in plant parts. A1B2 caused the highest enhancement in leaf number, fresh and dry weights of plant parts. It also improved fruit number significantly, fruit fresh and dry weight, yield plant−1 and fruit dimension compared to control under all NaCl solutions. All treatments improved carotenoids content and reduced cellular electrolyte leakage and sodium content, as compared to control. Low concentration of Nano-chelate Super Plus ZFM improved chlorophyll a (Chl a), b (Chl b) and total chlorophyll (TChl) more than the high concentration (A2). Photosynthetic pigments were maximized by A1B2. Treating plants with A1B2 and A1B1 improved the most nitrogen and potassium content respectively in shoots and fruits compared to control at all EC levels. Combining both products would be an efficient method inducing salt-tolerance of pepper.

Exploring the bio-control efficacy of Artemisia fragrans essential oil on the perennial weed Convolvulus arvensis: Inhibitory effects on the photosynthetic machinery and induction of oxidative stress

Convolvulus arvensis L. (Convolvulaceae), also known as ‘field bindweed’, is a cosmopolitan weed causing important loss in crop productivity and yield worldwide. Many efforts have been made to control this weed using chemical herbicides. Thus, its eradication represents an important challenge from an economic perspective. In this respect, plant essential oils have been recently exploited as bio-herbicides due to their documented allelopathic effects. The present study was aimed to examine the possible application of the essential oil of Artemisia fragrans Willd. (Asteraceae) as a bio-herbicide to control field bindweed. Two distinct Petri dish and pot-based experiments were carried out to assess the phytotoxicity of A. fragrans essential oil. The main essential oil components detected by gas chromatography-flame ionization detector (GC-FID) and gas chromatography-mass spectrometry (GC-MS) were α-thujone (30.4 %), camphor (26.4 %), 1,8-cineole (12.6 %) and β-thujone (10.0 %). The growth and content of photosynthetic pigments in field bindweed were reduced as a function of the increasing concentrations of the essential oil in both Petri and pot experiments. Furthermore, the chlorophyll a fluorescence as a key parameter was negatively affected when the essential oil treatment was applied. Also, exposure to essential oil altered the activity of catalase (CAT), peroxidase (POD), ascorbate peroxidase (APX) and superoxide dismutase (SOD), enhanced the level of hydrogen peroxide (H2O2), malondialdehyde (MDA) and caused cellular electrolyte leakage. In conclusion, the essential oil of A. fragrans has noteworthy potential as a growth and photosynthesis disruptor as well as an oxidative stress inducer against field bindweed. Thus, this plant species may be suggested as a source of bio-herbicides on an industrial level.

Natural variation in tolerance to sub-zero temperatures among populations of Arabidopsis lyrata ssp. petraea

BackgroundTemperature is one of the most important abiotic factors limiting plant growth and productivity. Many plants exhibit cold acclimation to prepare for the likelihood of freezing as temperatures decrease towards 0 °C. The physiological mechanisms associated with enabling increased tolerance to sub-zero temperatures vary between species and genotypes. Geographically and climatically diverse populations of Arabidopsis lyrata ssp. petraea were examined for their ability to survive, maintain functional photosynthetic parameters and cellular electrolyte leakage integrity after being exposed to sub-zero temperatures. The duration of cold acclimation prior to sub-zero temperatures was also manipulated (2 and 14 days).ResultsWe found that there was significant natural variation in tolerances to sub-zero temperatures among populations of A. petraea. The origin of the population affected the acclimation response and survival after exposure to sub-zero temperatures. Cold acclimation of plants prior to sub-zero temperatures affected the maximum quantum efficiency of photosystem II (PSII) (Fv/Fm) in that plants that were cold acclimated for longer periods had higher values of Fv/Fm as a result of sub-zero temperatures. The inner immature leaves were better able to recover Fv/Fm from sub-zero temperatures than mature outer leaves. The Irish population (Leitrim) acclimated faster, in terms of survival and electrolyte leakage than the Norwegian population (Helin).ConclusionThe ability to survive, recover photosynthetic processes and cellular electrolyte leakage after exposure to sub-zero temperatures is highly dependent on the duration of cold acclimation.

Effects of 1-aminocyclopropane-1-carboxylate-deaminase–Producing Bacteria on Perennial Ryegrass Growth and Physiological Responses to Salinity Stress

The accumulation of 1-aminocyclopropane-1-carboxylate (ACC), which is a precursor for ethylene production, in plant roots exposed to salinity stress can be detrimental to plant growth. The objectives of this study were to determine whether inoculating roots with bacteria containing deaminase enzymes that break down ACC (ACC-deaminase) could improve plant tolerance to salinity in perennial ryegrass (Lolium perenne) and to examine growth and physiological factors, as well as nutrition status of plants affected by the ACC-deaminase bacteria inoculation under salinity stress. Plants of perennial ryegrass (cv. Pangea) were inoculated with either Burkholderia phytofirmans PsJN or Burkholderia gladioli RU1 and irrigated with either fresh water (control) or a 250 mm NaCl solution to induce salinity stress. The bacterium-inoculated plants had less ACC content in shoots and roots under both nonstressed and salinity conditions. Salinity stress inhibited root and shoot growth, but the bacterium-inoculated plants exhibited higher visual turf quality (TQ), tiller number, root biomass, shoot biomass, leaf water content, and photochemical efficiency, as well as lower cellular electrolyte leakage (EL) under salinity stress. Plants inoculated with bacteria had lower sodium content and higher potassium to sodium ratios in shoots under salinity stress. Shoot and root nitrogen content and shoot potassium content increased, whereas shoot and root calcium, magnesium, iron, and aluminum content all decreased due to bacterial inoculation under salinity treatment. ACC-deaminase bacteria inoculation of roots was effective in improving salinity tolerance of perennial ryegrass and could be incorporated into turfgrass maintenance programs in salt-affected soils.

Absorption of nutrients, growth and nutritional disorders resulting from ammonium toxicity in rice and spinach plants

Excess nitrogen in the ammonium form may induce different nutritional disorders in plants depending on the species cultivated. Therefore, the objective herein was to evaluate the accumulation of nutrients, growth and the visual symptoms in rice and spinach plants in function of ammonium concentrations in the nutrient solution. Two experiments were performed with rice and spinach plants in a greenhouse, at the Unesp Campus of Jaboticabal-SP, Brazil. In each experiment five ammonium concentrations were used (10, 20, 40, 80 and 100 mmol L-1), arranged in a randomized block design with five repetitions. The effects of excess ammonia were studied on the green color index, electrolyte leakage, accumulation of N, Mg, K and Ca in the shoots, leaf area, height, dry matter of root, shoots and whole plant, and also characterization of the visual symptoms of toxicity in the plants. These effects of excess ammonium reported above are reflected in the production of plant dry matter, with a linear decrease in dry matter of the roots, shoots and whole plant. Excess ammonia in the nutrient solution decreased accumulation of dry matter due to reduced leaf area and increased cellular electrolyte leakage which induced symptoms characteristic of ammonium toxicity in both species, and also decreased uptake of Ca, Mg and K in rice plants and only Ca in spinach plants.

PpCBF3 from Cold-Tolerant Kentucky Bluegrass Involved in Freezing Tolerance Associated with Up-Regulation of Cold-Related Genes in Transgenic Arabidopsis thaliana.

Dehydration-Responsive Element Binding proteins (DREB)/C-repeat (CRT) Binding Factors (CBF) have been identified as transcriptional activators during plant responses to cold stress. The objective of this study was to determine the physiological roles of a CBF gene isolated from a cold-tolerant perennial grass species, Kentucky bluegrass (Poa pratensis L.), which designated as PpCBF3, in regulating plant tolerance to freezing stress. Transient transformation of Arabidopsis thaliana mesophyll protoplast with PpCBF3-eGFP fused protein showed that PpCBF3 was localized to the nucleus. RT-PCR analysis showed that PpCBF3 was specifically induced by cold stress (4°C) but not by drought stress [induced by 20% polyethylene glycol 6000 solution (PEG-6000)] or salt stress (150 mM NaCl). Transgenic Arabidopsis overexpressing PpCBF3 showed significant improvement in freezing (-20°C) tolerance demonstrated by a lower percentage of chlorotic leaves, lower cellular electrolyte leakage (EL) and H2O2 and O2 .- content, and higher chlorophyll content and photochemical efficiency compared to the wild type. Relative mRNA expression level analysis by qRT-PCR indicated that the improved freezing tolerance of transgenic Arabidopsis plants overexpressing PpCBF3 was conferred by sustained activation of downstream cold responsive (COR) genes. Other interesting phenotypic changes in the PpCBF3-transgenic Arabidopsis plants included late flowering and slow growth or ‘dwarfism’, both of which are desirable phenotypic traits for perennial turfgrasses. Therefore, PpCBF3 has potential to be used in genetic engineering for improvement of turfgrass freezing tolerance and other desirable traits.

Ameliorative effects of squash (Cucurbita moschata Duchesne ex Poiret) leaf extracts on oxidative stress

This study screened paraquat-tolerant plants among 10 plant species, including monocots and dicots angiosperms. Squash (Cucurbita moschata Duchesne ex Poiret) and kidney bean (Phaseolus vulgaris L.) plants exhibited the highest photooxidation-tolerant phenotypes upon a foliar treatment with paraquat. A foliar treatment with paraquat pre-mixed with leaf water extracts from the squash plant significantly alleviated paraquat-induced oxidative damage in maize, but this was not the case after a treatment with the hydrophobic phase of the leaf extracts. In particular, the water extract from young leaves (4th true leaf) of squash plants conferred tenfold higher tolerance to oxidative damage in paraquat-treated leave tissues compared to paraquat-only treatment. This tolerance was tightly linked not only to the increased amounts of ascorbic acid and dehydroascorbate antioxidants in the damaged leaves, but also to the reduced chlorophyll loss, lipid peroxidation, and cellular electrolyte leakage. Moreover, the protective effects of the water extract were apparent when using another bipyridyl herbicide, diquat, but not with a diphenyl-ether herbicide, oxyfluorfen. On the other hand, pre-treatment with the extract prior to the onset of drought or cold stress had no significant antioxidative effect on the treated tissues.

Anion exchanger and the resistance against thermal haemolysis

4,4′-Diiso-thiocyanato stilbene-2,2′-disulphonic acid (DIDS) is a membrane-impermeable, highly specific covalent inhibitor and powerful thermal stabiliser of the anion exchanger (AE1), the major integral protein of erythrocyte membrane (EM). Suspensions of control and DIDS-treated (15 µM, pH 8.2) human erythrocytes were heated from 20° to 70°C using various but constant heating rates (1–8°C/min). The cellular electrolyte leakage exhibited a sigmoidal response to temperature as detected by conductometry. The critical midpoint temperature of leakage, Tmo, extrapolated to low heating rate (0.5°C/min) was used as a measure for EM thermostability. Tmo was greater for DIDS-treated erythrocytes, 63.2° ± 0.3°C, than for intact erythrocytes, 60.7° ± 0.2°C. The time, t1/2, for 50% haemolysis of erythrocytes, exposed to 53°C was used as a measure for the resistance of erythrocytes against thermal haemolysis. The t1/2 was also greater for DIDS-treated erythrocytes, 63 ± 3 min, than for intact erythrocytes, 38 ± 2 min. The fluorescent label N-(3-pyrenyl)maleimide and EPR spin label 3-maleimido-proxyl, covalently bound to sulphydryl groups of major EM proteins, were used to monitor the changes in molecular motions during transient heating. Both labels reported an intensification of the motional dynamics at the denaturation temperatures of spectrin (50°C) and AE1 (67°C), and, surprisingly, immobilisation of a major EM protein, presumably the AE1, at Tmo. The above results are interpreted in favour of the possible involvement of a predenaturational rearrangement of AE1 copies in the EM thermostability and the resistance against thermal haemolysis.

Dehydration accelerates respiration in postharvest sugarbeet roots

Sugarbeet ( Beta vulgaris L.) roots lose water during storage and often become severely dehydrated after prolonged storage and at the outer regions of storage piles which have greater wind and sun exposure. Sucrose loss is known to be elevated in dehydrated roots, although the metabolic processes responsible for this loss are unknown. To identify processes that contribute to sucrose loss in dehydrated roots, respiration rate, cellular electrolyte leakage, and sucrolytic enzyme activities were determined in roots of two varieties (VDH 66156 and Beta 4797R) during 4 weeks of 10 °C storage at high (85%) and low (40%) relative humidities. Roots stored at 40% relative humidity dehydrated significantly and lost almost 50% of their weight after 4 weeks of storage. Electrolyte leakage increased in these roots, indicating that dehydration damaged cellular membranes. Respiration rate generally increased in roots stored at 40% relative humidity compared to roots stored at 85% relative humidity. The increase in respiration rate was positively correlated with root weight loss and electrolyte leakage. Respiration rate was most closely associated with electrolyte leakage, however, suggesting that elevations in respiration rate were not due to dehydration, but to the membrane damage that occurred in response to dehydration. Activities of the sucrose-degrading enzymes, sucrose synthase, alkaline invertase and soluble acid invertase, were unaltered by dehydration. Alterations in sucrolytic enzyme activities, therefore, were not needed to provide for the increased demand for respiratory substrates in dehydrated roots. These results suggest that storage at low relative humidity alters the postharvest physiology of sugarbeet roots by increasing the rate of weight loss, reducing membrane integrity, and accelerating root respiration rate.

Increased expression of OsSPX1 enhances cold/subfreezing tolerance in tobacco and Arabidopsis thaliana

Low temperature is a major environmental stress for plants. Many important cultivated crops have limited capacity to survive below freezing/subfreezing temperatures. Low inorganic phosphate (Pi) is reportedly important in triggering cold acclimatization. SPX (SYG1/Pho81/XPR1: SYG1, suppressor of yeast gpal; Pho81, CDK inhibitor in yeast PHO pathway; XPR1, xenotropic and polytropic retrovirus receptor) domain proteins have been shown to be involved in the phosphate-related signal transduction and regulation pathways. Recently, Arabidopsis AtSPX family genes have been found to possess diverse functions in plant tolerance to phosphorus starvation, and OsSPX1 is involved in phosphate homeostasis in rice and optimizes growth under phosphate-limited conditions through a negative feedback loop. In this study, our phylogenetic and gene expression profiling approaches identified six rice OsSPX genes up-regulated during cold stress. Transgenic tobacco plants with constitutive expression of OsSPX1 were more tolerant to cold stress than were wild-type plants, and showed better seedling survival and reduced cellular electrolyte leakage. In addition, there was decreased total leaf Pi content and accumulation of free proline and sucrose in transgenic tobacco plants during cold stress. To further establish a cause-and-effect relationship between intracellular Pi level and cold acclimatization in transgenic plants, we generated transgenic Arabidopsis plants with constitutive expression of OsSPX1. Cold stress resulted in reduced leaf Pi levels in Arabidopsis transgenic relative to wild-type plants. From real-time reverse transcriptase-polymerase chain reaction analysis, several Pi starvation-related genes, such as AtSPX1 (orthologue of OsSPX1), PHO2, PLDZ2 and ATSIZ1, showed differential expression between wild-type and transgenic plants during cold stress. Our results indicate that OsSPX1 may play an important role in linking cold stress and Pi starvation signal transduction pathways.

Triazole induced drought tolerance in horse chestnut (Aesculus hippocastanum)

We determined the influence of the triazole derivatives paclobutrazol, penconazole, epixiconazole, propiconazole and myclobutanil on the drought tolerance and post drought recovery of container-grown horse chestnut (Aesculus hippocastanum L.) saplings. Myclobutanil neither conferred drought resistance, as assessed by its effects on a number of physiological and biochemical parameters, nor affected growth parameters measured after recovery from drought. Chlorophyll fluorescence (F(v)/F(m)), photosynthetic rates, total foliar chlorophyll and carotenoid concentrations, foliar proline concentration and superoxide dismutase and catalase activities were consistently higher and leaf necrosis and cellular electrolyte leakage was lower at the end of a 3-week drought in trees treated with paclobutrazol, penconazole, epixiconazole or propiconazole than in control trees. Twelve weeks after drought treatment, leaf area and shoot, root and total plant dry masses were greater in triazole-treated trees than in control trees with the exception of those treated with myclobutanil. In a separate study, trees were subjected to a 2-week drought and then sprayed with paclobutrazol, penconazole, epixiconazole, propiconazole or myclobutanil. Chlorophyll fluorescence, photosynthetic rate, foliar chlorophyll concentration and catalase activity over the following 12 weeks were 20 to 50% higher in triazole-treated trees than in control trees. At the end of the 12-week recovery period, leaf area and shoot, root and total plant dry masses were higher in triazole-treated trees than in control trees, with the exception of trees treated with myclobutanil. Application of triazole derivatives, with the exception of myclobutanil, enhanced tolerance to prolonged drought and, when applied after a 2-week drought, hastened recovery from drought. The magnitude of treatment effects was in the order epixiconazole approximately propiconazole > penconazole > paclobutrazol > myclobutanil.

Effect of delta-aminolevulinic acid on the herbicidal efficacy of foliar-applied MTB-951, a mycoherbicide to control Echinochloa crus-galli L.

The conidia of a plant pathogen (Drechslera monoceras[Drechsler] Subram. et Jain [= Exserohilum monoceras[Drechsler] Leonard et Suggs]), MTB-951, which was isolated from native Echinochloa species in Japan, were used as the herbicidal active ingredient. The effects of δ-aminolevulinic acid (ALA) on the efficacy of foliar-applied MTB-951 on Echinochloa crus-galli L. were examined. First, foliar-applied MTB-951 exhibited almost no efficacy when no dew period was provided. However, the efficacy increased depending on the length of the dew period. MTB-951 and ALA exhibited an additive effect on E. crus-galli under 3 day dew conditions, although they exhibited a significant synergistic effect under no-dew conditions. When the leaves of E. crus-galli were immersed in deionized water containing the conidia of MTB-951, cellular electrolyte leakage from the leaves was observed. The ALA increased the electrolyte leakage by MTB-951 synergistically. Also, ALA increased the number of lesions on the leaves of E. crus-galli caused by MTB-951 under light conditions but failed to increase the number under dark conditions. These results suggest that ALA promotes infection with MTB-951 for E. crus-galli through its photodynamic action.

Effect of temperature on herbicidal properties of MTB‐951, a mycoherbicide to control Echinochloa crus‐galli L.

MTB‐951 is a potential mycoherbicide using a fungal plant pathogen (Drechslera monoceras) isolated from native Echinochloa species in Japan. Conidia of this pathogen were used as the active ingredient and the effect of temperature on its properties was examined in a laboratory. The optimum temperature for conidium germination was broad between 15 and 35°C and no difference in the germination ratio existed within this temperature range. Mycelial growth was the fastest at 25°C but was slower outside of the range 15–35°C. When leaves of Echinochloa crus‐galli L. were immersed in deionized water containing the conidia, cellular electrolyte leakage from the leaves was observed. However, no electrolyte leakage occurred from leaves of rice (Oryza sativa L.) under the same conditions. The occurrence of electrolyte leakage from E. crus‐galli was dependent on the temperature and was the highest at 25°C. Herbicidal activity of MTB‐951 against E. crus‐galli was high between 25°C and 30°C, but decreased at 35°C in a glass vessel. In the present study, it was revealed that the physiological and herbicidal properties of MTB‐951 depended on the temperature.

Comparison of physiological effects of fluazifop‐butyl and sethoxydim on oat (Avena sativa L.)

Laboratory experiments were conducted to compare the physiological effects of two herbicides: fluazifop‐butyl {butyl (RS)‐2‐[4‐(5‐trifluoromethyl‐2‐pyridyloxy)phenoxy]‐propionate} and sethoxydim {(±)‐2‐[1‐(ethoxyimino)butyl]‐5‐[2‐(ethylthio)propyl]‐3‐hydroxy‐2‐cyclohexen‐1‐one} on oat (Avena sativa L. cv. Zenshin). The herbicides strongly inhibited growth of oat and induced chlorosis at the basal part of shoots and ethylene production from the seedlings. The phytotoxicity of these herbicides in oat seedlings was alleviated by 2,4‐D (2,4‐dichlorophenoxyacetic acid), but not by IAA (indole‐3‐acetic acid). Coleoptile elongation induced by 2,4‐D or IAA was inhibited by fluazifop‐butyl and sethoxydim, suggesting both herbicides possess the activity to inhibit this auxin action. Fluazifop (free acid) and sethoxydim inhibited proton excretion from oat roots but fluazifop‐butyl did not. This proton excretion was not restored by 2,4‐D or IAA. Furthermore, cellular electrolyte leakage in oat shoots was increased by both herbicides, indicating that the membrane permeability was increased. We conclude that fluazifop‐butyl and sethoxydim may have the same mechanism of action which leads to disruption of membrane integrity, although fluazifop‐butyl acts as a free acid after hydrolysis (fluazifop).

Root membrane thermostability of Cornus florida L. provenances

Flowering dogwood seed collected from native trees in 3 USDA Hardiness Zones (6b, 7a, and 8a; American Horticultural Society (AHS) Heat-Zones 7, 7, and 8, respectively) were germinated and grown in containers prior to laboratory procedures to determine root cell membrane thermostability. Unsuberized, current season, fine root tissues were subjected to temperatures ranging from 20 to 60°C for 30 min and analyzed for cellular electrolyte leakage. Electrolyte leakage from root tissue exhibited a sigmoidal response to temperature for trees from each location. Critical midpoint temperature ( T m) was greater for seedlings native to USDA Hardiness Zone 6b (AHS Heat-Zone 7), 52.4±0.6°C, than T m for seedlings originating from USDA Zone 7a (AHS Zone 7), 51.2±0.5°C. However, seedlings from USDA Zone 8a (AHS Zone 8), at 51.5±0.4°C, were similar to those collected in USDA Zones 6b (AHS Zone7) and 7a (AHS Zone 7). The results of this study find little genetic variability across this part of the native range of flowering dogwood regarding root thermotolerance. Although differences between two provenances were found, these differences may be of little ecological significance.