Effect of salinity and different nitrogen sources on the activity of antioxidant enzymes and indole alkaloid content in Catharanthus roseus seedlings

The activities of antioxidant enzymes viz. glutathione reductase, GR; superoxide dismutase, SOD; peroxidase, POD; catalase, CAT and glutathione-S-transferase, GST and alkaloid accumulation were investigated in leaf pairs (apical, middle, basal) and in roots of Catharanthus roseus seedlings under the conditions of different nitrogen sources (20 mM KNO(3) and 2 mM NH(4)Cl) and salinity, in the absence (non-saline control) and in the presence of 100 mM NaCl in the nutrient solution. Salinity caused a reduction in plant biomass. The biomass production of ammonium-fed plants was lower than that of nitrate-fed plants. The antioxidant enzymes exhibited higher activity in saline-treated plants. Changes in antioxidant enzyme activity caused by different nitrogen sources differed in all leaf pairs, as well as in roots of C. roseus. Ammonium-fed plants showed higher CAT, GR and GST activity in leaf pairs as well as in roots, while POD and SOD activity were higher in nitrate-fed plants. Higher peroxidase activity concomitant with the increased accumulation of alkaloid was found in all leaf pairs, as well as in roots of C. roseus of NO(3)(-) fed plants as compared to NH(4)(+) fed plants.     

Apoplastic superoxide production and peroxidase activity by intact and excised axenically grown seedling roots of sunflower

Excised and cold-preincubated sunflower seedling roots were compared with intact non-preincubated roots to test the effect of the injury stress and cold preincubation on the oxidative burst measured as apoplastic superoxide (O (2) (.-) ) generation and exocellular peroxidase (ECPOX) activity. Preincubated excised or intact roots released into the medium apoplastic proteins with peroxidase activity. Intact and excised roots responded to methyl jasmonate by an immediate oxidative burst that could not be induced by salicylic acid; both phytohormones also induced a slight and slow O (2) (.-) generation and ECPOX activity on excised roots, when added to the cold preincubation medium. The results with cyanide, azide, SHAM (ECPOX inhibitors) and diphenylene iodonium (inhibitor of trans-plasma membrane NAD(P)H-oxidases (NOX)-respiratory burst oxidase homologue in plants (RBOH), the trans-plasmamembrane nicotinamide adenine dinucleotide phosphate oxidase) are consistent with the hypothesis that different systems may be the origin of O (2) (.-) in intact and excised roots; ECPOX was an important component of them in both, together with NOX-RBOH in intact roots, but in excised roots the last one was replaced by an oxidase sensitive to the same inhibitors as the alternative mitochondrial oxidase. According to our hypothesis, these results could be explained if the electron flux would be deviated to different interconnected plasma membrane-redox systems, with different terminal oxidases, activated by different effectors or stresses.     

Cadmium induced oxidative stress and changes in soluble and ionically bound cell wall peroxidase activities in roots of seedling and 3–4 leaf stage plants of <Emphasis Type="Italic">Brassica juncea</Emphasis> (L.) czern

Metabolic adaptations to heavy metal toxicity in plants are thought to be related with developmental growth stage and the type of metal by which plant is affected. In the present study, changes in ionically bound CWP, soluble peroxidase activity, H(2)O(2) level and Malonaldehyde content in roots of cadmium and copper stressed seedlings and cadmium stressed 3-4 leaf stage plants of Brassica juncea were investigated. Cadmium inhibits root growth and reduces fresh biomass. The reduction in root growth and fresh biomass is correlated with increased lipid peroxidation and reduced tolerance. Treatment with cadmium resulted in an increase in ionically bound CWP activity in roots of seedlings but no significant change in its activity was found in roots of 3-4 leaf stage plants. Increased level of H(2)O(2) in roots of cadmium and copper treated seedlings, show a direct correlation with increased activity of ionically bound CWP. H(2)O(2) level in 3-4 leaf stage plant roots was found to be very low. Soluble peroxidase activity decreased in cadmium (50 and 100 mu-icroM) treated seedlings but it was ineffective to cause any change in its activity in 3-4 leaf stage plants. Copper treated seedlings showed an increase in ionically bound CWP activity, H(2)O(2) level and MDA content. Ascorbic acid (50 mM) pretreated seedlings shows significant decrease in ionically bound CWP activity when exposed to 50 muM cadmium. Hence, it is concluded that inhibition of root growth in Brassica juncea seedlings by cadmium, is associated with CWP catalyzed H(2)O(2) dependent reactions which are involved in metabolic adaptations to heavy-metal stress.     

Variation in cadmium accumulation potential and tissue distribution of cadmium in tobacco

Variation in Cd accumulation between Nicotiana species but not varieties has been observed in seedlings grown in solution culture with moderate-to-low levels of Cd. Nicotiana tabacum has been characterized as a leaf and root accumulator while Nicotiana rustica is shown to be primarily a root accumulator, having about half the leaf Cd per gram dry weight of N. tabacum. This phenotype is retained in the mature N. rustica plant. To characterize these two species which differ in their modes of Cd accumulation, tissue Cd distribution, partitioning of metal in soluble and insoluble fractions and the contribution of soluble Cd-binding proteins (peptides) to total plant Cd was assessed using mature solution cultured plants. Metal accumulation was highest in the most mature leaves and in young roots. The preponderance of young roots in N. rustica may, in part, account for low leaf/high root Cd accumulation in this species. While Cd-binding peptides appear to be a principal form of Cd in leaves and roots of seedlings and these also occur in mature leaves, Cd is equally distributed between soluble (about 80% as Cd-binding peptide) and uncharacterized insoluble forms in mature plant roots.     

Effect of <Emphasis Type="Italic">Pseudomonas</Emphasis> Bacteria on Peroxidase Activity in Wheat Plants when Infected with <Emphasis Type="Italic">Bipolaris sorokiniana</Emphasis>

We investigated the effect of treating soft wheat seeds (Triticum aestivum L.) with two Pseudomonas bacteria strains, isolated from earthworm coprolites, showing a significant antifungal and growth-promoting action in preliminary screening on the activity of guaiacol-dependant peroxidase under phytopathogenic load in the presence of Bipolaris sorokiniana (Sacc.) Shoemaker as a mechanism for inducing plant resistance to the pathogen. We established a statistically significant decrease (P < 0.05) in root rot disease incidence and severity during bacterization, which is indicative both of antifungal activity of the used bacterial isolates and of their successful colonizing the rhizosphere of wheat plants. We noted a response of free and weakly bound peroxidase of wheat plants to infection with B. sorokiniana: the enzyme activity increased during pathogenesis. Bacterization also increased peroxidase activity in plant leaves and roots, the greatest differences from non-bacterized plants being observed in wheat roots in the presence of the pathogen. We detected a direct link between peroxidase activity in wheat roots and leaf tissues in the absence of the pathogen and the feedback between peroxidase activity and plant infestation by the root rot pathogen. In the presence of the phytopathogen, there is a lack of correlation between peroxidase activity in wheat roots and leaves, and there is a shift of activity towards its increase in roots, which plays an important role in the development of systemic resistance against the root rot pathogen that penetrates into plants through the roots and root collar.     

The arbuscular mycorrhizal fungus Diversispora spurca ameliorates effects of waterlogging on growth,root system architecture and antioxidant enzyme activities of citrus seedlings

Citrus plants are often exposed to heavy rain and subsequent periods of soil waterlogging which severely restrict tree growth. We assessed the effect of one arbuscular mycorrhizal fungus species (Diversispora spurca) on growth, root system architecture (RSA), and antioxidant enzyme activities of young citrus (Citrus junos) seedlings. Waterlogging for 37 d significantly restricted mycorrhizal colonization but increased the number of entry points and vesicles. Compared with non-mycorrhizal controls, mycorrhizal seedlings had significantly greater plant height, fresh mass, total root and taproot lengths, projected and surface root areas, root volume, and numbers of lst, 2nd and 3rd order lateral roots regardless of waterlogging treatment. D. spurca significantly increased root catalase (CAT) activity in non-stressed seedlings and increased root soluble protein concentration and leaf CAT activity in waterlogged seedlings, thereby inducing lower oxidative damage. These results suggest that D. spurca ameliorates effects of waterlogging on growth, RSA and antioxidant enzyme activities.     

Cd induced changes in proline level and peroxidase activity in roots of rice seedlings

The effects of Cd on changes in proline level and peroxidase activity in roots of rice seedlings were investigated. CdCl₂ was effective in inhibiting root growth and in accumulating proline in roots. The inhibition of root growth by Cd is reversible. The reduction of root growth induced by Cd is closely associated with accumulation of proline in roots. External application of proline markedly inhibited root growth of rice seedlings in the absence of Cd. Ionically bound, but not soluble, peroxidase activity in roots was increased by CdCl₂. Proline treatment also resulted in an increase in ionically bound peroxidase activity in roots. The relationship between growth inhibition of roots induced by Cd and changes of proline level and peroxidase activity is discussed.Abbreviations POX peroxidase     

The influence of phosphorus availability and Laccaria bicolor symbiosis on phosphate acquisition,antioxidant enzyme activity,and rhizospheric carbon flux in Populus tremuloides

Many forest tree species are dependent on their symbiotic interaction with ectomycorrhizal (ECM) fungi for phosphorus (P) uptake from forest soils where P availability is often limited. The ECM fungal association benefits the host plant under P limitation through enhanced soil exploration and increased P acquisition by mycorrhizas. To study the P starvation response (PSR) and its modification by ECM fungi in Populus tremuloides, a comparison was made between nonmycorrhizal (NM) and mycorrhizal with Laccaria bicolor (Myc) seedlings grown under different concentrations of phosphate (Pi) in sand culture. Although differences in growth between NM and Myc plants were small, Myc plants were more effective at acquiring P from low Pi treatments, with significantly lower k _m values for root and leaf P accumulation. Pi limitation significantly increased the activity of catalase, ascorbate peroxidase, and guaiacol-dependent peroxidase in leaves and roots to greater extents in NM than Myc P. tremuloides. Phosphoenolpyruvate carboxylase activity also increased in NM plants under P limitation, but was unchanged in Myc plants. Formate, citrate, malonate, lactate, malate, and oxalate and total organic carbon exudation by roots was stimulated by P limitation to a greater extent in NM than Myc plants. Colonization by L. bicolor reduced the solution Pi concentration thresholds where PSR physiological changes occurred, indicating that enhanced Pi acquisition by P. tremuloides colonized by L. bicolor altered host P homeostasis and plant stress responses to P limitation. Understanding these plant–symbiont interactions facilitates the selection of more P-efficient forest trees and strategies for tree plantation production on marginal soils.     

Effects of metalaxyl enantiomers stress on root activity and leaf antioxidant enzyme activities in tobacco seedlings

The objective of this experiment was to study the effects of metalaxyl enantiomers on the activity of roots and antioxidative enzymes in tobacco seedlings. Water culture experiment was conducted to analyze the effects of different concentrations of metalaxyl enantiomers (30 and 10 mg L^?1) on root activity and leaf superoxide dismutase (SOD), peroxidase (POD), and catalase (CAT) activities and malondialdehyde (MDA) content of tobacco seedlings. The results showed that metalaxyl significantly inhibited root activity and significantly improved leaf SOD, POD, and CAT activities and MDA content. A better physiological response in tobacco seedlings was observed at 30 mg L^?1 than at 10 mg L^?1 metalaxyl. The stereoselectivity for different enantiomers had no obvious effect on root activity and the leaf POD activity, but it affected significantly the SOD and CAT activities and MDA content. The SOD activity was promoted more by R‐enantiomer than by S‐enantiomer at 30 mg L^?1 metalaxyl, and the same effect was observed on CAT activity from the beginning to the end of the stress period. The MDA content under the stress by R‐enantiomer was higher than that under the stress by S‐enantiomer at 10 mg L^?1 metalaxyl.     

土壤结构与光照水平对秋茄某些生长和生理参数的影响

在香港的３个红树森样地即黄竹湾（沙土）、西径（沙壤土）和米埔（粘壤土）进行了土壤结构对秋茄（Kandelia candel(L.)Durce)生长和生理影响的研究,并在米埔比较了林内和林外秋茄幼苗的生长和生理参数以观察光照水平的效应。在沙土和沙壤土生长的１．５年秋茄幼苗比粘壤土具有较粗的基径的较高的总生物量,说明秋茄幼苗在沙土和沙壤土中比在粘壤土中生长更好。沙土１．５ 茄幼苗的叶片厚度分别为沙壤土和粘壤土的１．７５和２．０５倍,表明沙土中的秋茄幼苗具有旱生结构以维持体内水分。然而,沙土和沙壤土４．５年秋茄幼树的叶片厚度无显著差异,沙土和沙壤土中１．５年秋茄幼苗分配于根系的生物量比例约为５０％,高于粘壤土的值（约４０％）。沙土和沙壤土中１．５年的秋茄比粘壤土具有较低的叶绿素含量、根系活力、硝酸盐还原酶活性、过氧化物酶（ＰＯＸ）活性、超氧化物歧化酶（ＳＯＤ）活性及较高的丙二醛（ＭＤＡ）含量。米埔１．５年秋茄幼苗在红树林外比林内有更好的长势,具有更大的叶面积、特殊叶面积、叶片数量及生物量。林内幼苗具有较高叶绿素含量,较低叶绿素a/b比值,较高硝酸盐还原酶活性和较强的根系活力,林外幼苗的叶片ＰＯＸ和ＳＯＤ活性比林内的值稍高,ＭＤＡ含量比林内显著要高。     

Methodological approaches for studying apoplastic redox activity: 1. Mechanisms of peroxidase release

It was demonstrated the efficiency of the application of extracellular solution, i.e., the leachate derived from roots after their incubation, for the analysis of apoplastic redox enzymes (exemplified by peroxidase) in the roots of 5-day-old seedlings of spring wheat (Triticum aestivum L., cv. Kazanskaya yubileinaya) after wounding stress. Such non-invasive approach allows the studying of enzymes without a disruption of plant tissue. The inhibitory analysis showed that newly synthesized and pre-existing in the cells peroxidases were not released from the cytoplasm at stress conditions. The proposed methodological approach for obtaining the enzymatic extracts of apoplastic redox enzymes opens up broad prospects for researches of plant immunity and stress responses.     

Hydrogel substrate amendment alleviates drought effects on young citrus plants

Water deficits affect citrus physiology, yield, fruit size and quality. Citrus can respond to drought stress conditions through endogenous hormonal regulation of water status and leaf abscission. In this work, we assayed the efficiency of an amendment to soilless media in delaying the drought stress effect in young citrus seedlings and trees. Substrate amendment promoted plant survival of citrus seedlings subjected to several cycles of drought stress and rehydration. In budded trees, the amendment increased substrate water content, leaf water potential, leaf number, root biomass, CO₂ assimilation and stomatal conductance over that of control plants growing in non-amended substrates. We conclude that the substrate amendment reduced the damaging effects of drought stress in citrus plants. The longer survival of seedlings in the amended treatment together with the reduction in leaf abscission and the improvement of physiological parameters, can account for a higher vigour of citrus grown under water stress conditions.     

Influence of age and sulfur metabolism on ATP sulfurylase activity in the soybean and a survey of selected species

ATP sulfurylase activity varied greatly among different leaves on the soybean plant [Glycine max (L.) Meer.], and high levels of activity did not appear in the leaves until the seedlings were about 3 weeks old. In general, leaves from the top of the plant had a higher activity than leaves from the bottom of the plant. A much greater activity was found in soybean leaves than in soybean roots. The absence of sulfate in the nutrient solution resulted in higher enzyme activity in leaves from young plants and in lower activity in leaves from older plants. Over the growing season, however, ATP sulfurylase activity appeared to be related to sulfur content of the leaf. Several other plant species also had measurable levels of ATP sulfurylase.     

Endogenous ascorbate restrains apoplastic peroxidase activity during sunflower leaf development

Several apoplastic enzymes have been implicated in the control of elongation growth of plant cells. Among them, peroxidases contribute to both loosening and stiffening of the cell wall. They appear to be regulated by various mechanisms, including the action of extracellular inhibitors. To obtain evidence of the role of the enzyme–inhibitor interaction during leaf development, the intercellular washing fluids from Helianthus annuus leaves of different ages were isolated using standard methods of vacuum infiltration and centrifugation. Peroxidase activities, assessed using tetramethylbenzidine as substrate, increased during leaf development, reaching a maximum value after the leaves were fully expanded. An inhibitor, chemically characterised as ascorbate, co‐localised with the enzyme in the apoplast. Moreover, there was a strong negative correlation between the action of peroxidase and the micromolar concentration of ascorbate in the apoplastic fluid. The results show that in growing leaves, the in planta ascorbate concentration is able to restrain peroxidase enzyme activity. Then, at the time of growth cessation, the loss of extracellular ascorbate relieves the inhibition on this enzyme that contributes to wall fixation.     

Role of extracellular peroxidase in the superoxide production by wheat root cells

Summary Extracellular peroxidase has been shown to contribute to superoxide production in wounded wheat (Triticum aestivum L. cv. Ljuba) root cells. The superoxide-synthesizing system of root cells was considerably inhibited by KCN and NaN₃ and activated by MnCl₂ and H₂O₂. Treatment of roots with salicylic acid and a range of di- and tri-carbonic acids (malic, citric, malonic, fumaric, and succinic acids) stimulated superoxide production in both root cells and extracellular solution. The H₂O₂-stimulated superoxide production in the extracellular solution was much higher when roots were preincubated with salicylic or succinic acid. Exogenous acids enhanced peroxidase activity in the extracellular solution. Pretreatment of root cells with the detergents trypsin and sodium dodecyl sulfate had similar effects on the peroxidase activity. Significant inhibition of both superoxide production and peroxidase activity by diphenylene iodonium suggests that the specificity of the latter as an inhibitor of NADPH oxidase is doubtful. Results obtained indicate that extracellular peroxidase is involved in the superoxide production in wheat root cells. The mobile form of peroxidase can be readily secreted to the apoplastic solution and serve as an emergency enzyme involved in plant wound response.Abbreviations DPI diphenylene iodonium - ECS extracellular solution - ROS reactive oxygen species - SA salicylic acid     

Protein and Peroxidase Modulations in Sunflower Seedlings (Helianthus annuus L.) Treated with a Toxic Amount of Aluminium

The aim of this study is to investigate the effect of aluminium treatment on peroxidases activities and protein content in both soluble and cell-wall-bound fractions of sunflower leaves, stems and roots. Fourteen-day-old seedlings, grown in a nutrient solution, were exposed to a toxic amount of aluminium (500 μM AlNO₃) for 72 h. Under stress conditions, biomass production, root length and leaf expansion were significantly reduced. Also, our results showed modulations on soluble and ionically cell-wall-bound peroxidases activities. In soluble fraction, peroxidases activities were enhanced in all investigated organs. This stimulation was also observed in ionically cell-wall-bound fraction in leaves and stems. Roots showed a differential behaviour: peroxidase activity was severely reduced. Lignifying peroxidases activities assayed using coniferyl alcohol and H₂O₂ as substrates were also modulated. Significant stimulation was shown on soluble fraction in leaves, stems and roots. In ionically cell-wall-bound fraction lignifying peroxidases were enhanced only in stems but severely inhibited in roots. Also, aluminium toxicity caused significant increase on cell wall protein content in sunflower roots.     

Comparative study of alleviating effects of GSH, Se and Zn under combined contamination of cadmium and chromium in rice (Oryza sativa)

A hydroponic experiment was conducted to study the ameliorative effects of separate or combined application of exogenous glutathione (GSH), selenium (Se) and zinc (Zn) upon 20 μM cadmium (Cd) plus 20 μM chromium (Cr) heavy metal stress (HM) in rice seedlings. The results showed that HM caused a marked reduction in seedling height, chlorophyll content (SPAD) and biomass, and activities of catalase (CAT) and ascorbate peroxidase (APX) in leaves and H⁺-ATPase in roots/leaves, but elevated superoxide dismutase (SOD) and guaiacol peroxidase (POD) activities in leaves with elevated malondialdehyde (MDA) accumulation both in leaves and roots over the control. The best mitigation effect was recorded in HM+GSH+Zn and HM+GSH (addition of GSH+Zn and GSH to HM solution), which greatly alleviated HM-induced growth inhibition and oxidative stress. Compared with HM alone, HM+GSH and HM+GSH+Zn markedly reduced Cr uptake and translocation but not affected Cd concentration; improved H⁺-ATPase activity and Fe, Zn, Mn uptake and translocation, and repressed MDA accumulation. Meanwhile exogenous GSH and GSH+Zn counteracted HM-induced response of antioxidant enzymes, via suppressing HM-induced dramatic increase of root/leaf SOD and leaf POD activities, and elevating stress-depressed leaf APX and leaf/root CAT activities.     

Changes in cucumber hypocotyl cell wall dynamics caused by Azospirillum brasilense inoculation

We previously reported that Azospirillum brasilense induced a more elastic cell wall and a higher apoplastic water fraction in both wheat coleoptile and flag leaf. These biophysical characteristics could permit increased growth. Knowledge of the biochemical effects the bacteria could elicit in plant cell walls and how these responses change plant physiology is still scarce. The objective of this work was to analyze whether A. brasilense Sp245 inoculation affected elongation and extensibility of growing cucumber (Cucumis sativus) hypocotyls and ionically bound cell wall peroxidase activities. Hypocotyl tip and basal segments were excised from A. brasilense Sp245-inoculated cucumber seedlings growing in darkness under hydroponic conditions. Elongation, cell wall extensibility, cell wall peroxidase activities against ferulic acid and guaiacol and NADH oxidase activities were analyzed. Azospirillum-inoculated cucumber seedlings grew bigger than non-inoculated ones. Dynamic cell wall differences were detected between inoculated and non-inoculated hypocotyls. They included greater acid-induced cell wall extension and in vivo elongation when incubated in distilled water. Although there was no difference between treatments in either region of the hypocotyl NADH oxidase and ferulic acid peroxidase activities were lower in both regions in inoculated seedlings. These lesser activities could be delaying the stiffening of cell wall in inoculated seedlings. These results showed that the cell wall is a target for A. brasilense growth promotion.     

林下植物根系对森林凋落物分解过程中微生物及酶活性的影响

为深入理解进入凋落物层生长的林下植物根系对森林凋落物分解的影响,本研究通过分解袋模拟试验探讨不同生物量多花黑麦草根系对中亚热带常绿阔叶林优势树种四川山矾凋落叶分解中微生物及酶活性的影响.结果表明: 在分解的240 d进程中,无根(N)、少根(L)、多根(M)3种处理下凋落叶表面细菌和真菌群落多样性指数均表现为多根＞少根＞无根处理,并且不同根生物量处理对真菌群落组成和数量的影响较细菌更为显著.随着多花黑麦草生长季结束,生长进入分解袋中的活根生物量逐渐减少,根系对真菌群落组成的影响减小.同一分解阶段,凋落叶表面酸性磷酸酶、β-葡萄糖苷酶、多酚氧化酶、过氧化物酶活性在有根条件下均高于无根条件.表明根系的生长能够改变微生物群落组成与数量,并提高微生物胞外酶活性,从而对分解产生促进作用.     

Analysis of salinity effects on basil leaf surface area, photosynthetic activity, and growth

Basil (Ocimum basilicum L.) seedlings were cultured on liquid medium in controlled conditions. Two varieties differing in leaf size were compared. When plants were 30?days old, the medium was supplemented with 50?mM NaCl. After 15?days of treatment, root, stem and leaf biomass, leaf number, and leaf surface area were measured. Ion accumulation was determined in roots, stems, and leaves. Photosynthetic parameters (CO₂ fixation rate, internal CO₂ concentration, stomatal conductance) as well as transpiration rate were determined on separate leaves. Electrolyte leakage and malondialdehyde content were used to estimate damage to membranes and lipid peroxidation, respectively. Several antioxidant enzymatic activities were used as proxies of oxidative stress. High Na⁺ concentration was reached in leaf tissues. Salt restricted whole plant biomass deposition rate by diminishing leaf number and leaf expansion, as well as photosynthetic activity were estimated from whole plant biomass production per unit leaf surface area. Diminished stomatal conductance restricted CO₂ fixation rate, and decrease in chlorophyll content presumably limited photosynthetic activity. Lipid peroxidation revealed damages to membranes. The magnitude of these responses differed between the two varieties, indicating that an intraspecific variability in salt response exists in basil.