Seed pretreatment with melatonin confers cadmium tolerance to chickpea seedlings through cellular redox homeostasis and antioxidant gene expression improvement.

Several phytoremediation strategies have been undertaken to alleviate cadmium (Cd)-mediated injury to crop yield resulting from agricultural land pollution. In the present study, the potentially beneficial effect of melatonin (Me) was appraised. Therefore, chickpea (Cicer arietinum L.) seeds were imbibed for 12 H in distilled water or Me (10 µM) solution. Then, the seeds germinated in the presence or the absence of 200 µM CdCl2 for 6 days. Seedlings obtained from Me-pretreated seeds exhibited enhanced growth traits, reflected by fresh biomass and length increase. This beneficial effect was associated with a decreased Cd accumulation in seedling tissues (by 46 and 89% in roots and shoots, respectively). Besides, Me efficiently protected the cell membrane integrity of Cd-subjected seedlings. This protective effect was manifested by the decreased lipoxygenase activity and the subsequently reduced accumulation of 4-hydroxy-2-nonenal. Melatonin counteracted the Cd-mediated stimulation of the pro-oxidant NADPH-oxidase (90 and 45% decrease compared to non-pretreated Cd-stressed roots and shoots, respectively) and NADH-oxidase activities (almost 40% decrease compared to non-pretreated roots and shoots), preventing, thus, hydrogen peroxide overaccumulation (50 and 35% lesser than non-pretreated roots and shoots, respectively). Furthermore, Me enhanced the cellular content of pyridine nicotinamide reduced forms [NAD(P)H] and their redox state. This effect was associated with the Me-mediated stimulation of the glucose-6-phosphate dehydrogenase (G6PDH) and malate dehydrogenase activities, concomitantly with the inhibition of NAD(P)H-consuming activities. These effects were accompanied by the up-regulation of G6PDH gene expression (45% increase in roots) and the down-regulation of the respiratory burst oxidase homolog protein F (RBOHF) gene expression (53% decrease in roots and shoots). Likewise, Me induced an increased activity and gene transcription of the Asada-Halliwell cycle, namely ascorbate peroxidase, monodehydroascorbate reductase, dehydroascorbate reductase, and glutathione reductase, concomitantly with a reduction of the glutathione peroxidase activity. This modulating effect led to the restoration of the redox homeostasis of the ascorbate and the glutathione pools. Overall, current results attest that seed pretreatment with Me is effective in Cd stress relief and can be a beneficial crop-protective approach.

Insights on strain 115 plant growth-promoting bacteria traits and its contribution in lead stress alleviation in pea (Pisum sativum L.) plants.

The present study aims to characterize the plant growth-promoting bacterial traits of Bacillus simplex (strain 115). This bacterium was inoculated in hydroponically conditions to improve pea (Pisum sativum L.) growth submitted to lead (Pb) toxicity. Root nodulation system was developed enough in 23-day-old plants attesting the interaction between the two organisms. In addition to its phosphate solubilization and siderophore production traits that reached 303.8 µg P mL-1 and 49.6 psu respectively, the Bacillus strain 115 exhibited Pb bio-sorption ability. Inoculation of Pb-stressed pea with strain 115 showed roots and shoots biomass recovery (+ 70% and + 61%, respectively). Similarly, water and protein contents were increased in Pb-treated plants after bacterial inoculation. In the presence of strain 115, Pb relative toxicity level decreased (- 39.3% compared to Pb stress only). Moreover, catalase and superoxide dismutase activities were upregulated in Pb-exposed plants (+ 56% and + 51%, respectively). After inoculation with strain 115, catalase and superoxide dismutase activities were restored by - 38% and - 44% respectively. Simultaneously, oxidant stress indicator (H2O2 and 4-hydroxynonenal) and osmo-regulators (proline and glycine-betaine) contents as well as lipoxygenase activity decreased significantly in Pb-treated plants after Bacillus strain's inoculation. Taken together, the results give some evidences for the plant growth-promoting capacity of strain 115 in helping alleviation of Pb stress.

Clue of zinc oxide and copper oxide nanoparticles in the remediation of cadmium toxicity in Phaseolus vulgaris L. via the modulation of antioxidant and redox systems.

The present study represents new evidence of the successful use of metal oxide nanoparticles in the remediation of heavy metals. Zinc oxide nanoparticles (ZnO NP) and copper oxide nanoparticles (CuO NP) were assessed to alleviate cadmium (Cd) toxicity in Phaseolus vulgaris L. seedlings and plants. Monitoring physiological and metabolic parameters allowed to elucidate Cd mechanism and process whereby it exerts phytotoxic effects on bean. The response of P. vulgaris seedlings is NP dose-dependent (10 mg/L, 50 mg/L, 100 mg/L, and 200 mg/L). Similarly, applied concentrations triggered a differential response of growing plants in terms of length and biomass. Our physiological data allowed to select 100 mg/L as the most appropriate concentration to apply, in order to avoid any risk of phytotoxicity. The regulatory mechanisms by which ZnO NP and CuO NP act are for the first time compared in the embryonic axes of bean seedlings under Cd stress. Both NP were able to reduce the hypergeneration of hydrogen peroxide (H2O2). They also acted via enhancing ROS scavenging enzymatic capacity, and activity of antioxidant enzymes CAT, APX, GPOX, GPX, and GR, and inhibited the activity of ROS producing enzymes such as GOX and NOX. Another mechanistic effect of NP consisted of the modulation of redox enzymes Trx, NTR, Fd, and FNR evolved in cellular homeostasis and maintaining reduced status in cells. Taken together, ZnO NP triggered more significant metabolic regulations allowing to mitigate the oxidative damage caused by Cd.

Cu and CuO Nanoparticles Affected the Germination and the Growth of Barley (Hordeum vulgare L.) Seedling.

The application of Cu and CuO nanofertilizers in horticulture has been a promising strategy to promote plants' growth. In our study, increasing concentrations (10, 25, 50, 100, 250, 500, 1000, 2000 mg/L) were assessed for their inhibitory or stimulatory effects on barley (Hordeum vulgare L.) seedlings. Our results showed an enhancement of seed germination parameters (FGP, t50, cumulative germination rate, AUC), and seedling growth parameters (roots and shoots' lengths, fresh biomasses and dry biomasses) by the low concentrations of Cu NPs and CuO NPs, while concentrations above 500 mg/L displayed inhibiting effects. CuO NPs treatment showed a significant similitude with CuSO4, which confirms that CuO NPs act mainly via released Cu ions. However, Cu NPs exhibited a different behavior since the nanosized particles contribute together with Cu ions in barley response to Cu NPs. This provides an overall picture of the way these nanoparticles may behave in plant systems.

Nitric oxide donor, sodium nitroprusside modulates hydrogen sulfide metabolism and cysteine homeostasis to aid the alleviation of chromium toxicity in maize seedlings (Zea mays L.).

The current research aimed to assess the protective role of nitric oxide (NO) against chromium (Cr) toxicity in maize seedlings. Chromium (200 µM) lowered osmotic potential in epicotyls and mostly in radicles (by 38% and 63%, respectively) as compared to the control. Sodium nitroprusside (SNP, NO donor) restored seedling biomass (+90% for both organs) and water potential, whereas application of Nω-nitro-L-arginine methylester (L-NAME, a NOS inhibitor) increased sensitivity to Cr. SNP suppressed Cr-triggered proline accumulation by inhibiting Δ1-pyrroline-5-carboxylate synthetase activity and stimulating proline dehydrogenase activity, leading to glutamate over-accumulation (~30% for both organs). Cr stimulated cysteine metabolism and this was further enhanced by SNP which stimulated serine acetyl-transferase and O-acetylserine (thiol) lyase activities. This was followed by an increase in endogenous hydrogen sulfide (H2S) generation by up-regulating L-cysteine desulfhydrase (+205%), D-cysteine desulfhydrase (+150%) and cyanoalanine synthase (+65%) activities in radicles compared to Cr-treatments plants. These positive effects were reduced in L-NAME compared to control. Combined Cr+SNP affected the levels of compounds involved in glutathione metabolism (γ-glutamyl-cysteinyl, γ-glutamyl-cysteinyl-clycine, γ-cysteinyl-glycine, and glycine.). All together, our findings indicate that NO and elicited cellular H2S act synergistically to alleviate Cr stress in maize seedlings by influencing a metabolic interplay between cysteine, proline, and glutathione.

Exogenous nitric oxide alleviates manganese toxicity in bean plants by modulating photosynthesis in relation to leaf lipid composition.

Nitric oxide (NO) is a signaling molecule controlling several steps of plant development and defense process under stress conditions. NO-induced alleviation of manganese (Mn) toxicity was investigated on bean plants submitted for 28 days to 500 µM MnCl2. Manganese excess decreased plant dry weight and elongation and increased levels of reactive oxygen species and lipid peroxidation leading to up-regulation of superoxide dismutase, catalase, and ascorbate peroxidase activities. The inhibitory effects of Mn on plant growth were associated to reduction of light-saturated carbon assimilation (Amax), stomatal conductance (gs), and transpiration (E). By contrast, Mn induced significant increase in the apparent quantum yield (ɸ) and light compensation point (LCP). Interestingly, intracellular CO2 (Ci) remains stable under Mn stress. Concomitantly, leaf membrane lipids have drastically reduced under high Mn concentration. After Mn exposition, leaf fatty acids exhibited a significant loss of linolenic acid, accompanied by an accumulation of palmitoleic, stearic, and linoleic acids leading to alteration of lipid desaturation. NO supply reversed Mn toxicity as evidenced by enhancement of growth biomass and recovery of Amax, E, ɸ, and LCP. Similarly, NO addition has positive effects on leaf lipid content and composition leading to restoration of lipid unsaturation. The modulation of fatty acid composition can be a way to reduce leaf membrane damages and maintain optimal photosynthesis and plant growth. Despite the absence of enough evidences in how NO is involved in lipid and photosynthesis recovery under Mn stress conditions, it is assumed that NO beneficial effects are attributable to NO/Mn cross-talk.

Gallic acid improves the antioxidant ability against cadmium toxicity: Impact on leaf lipid composition of sunflower (Helianthus annuus) seedlings.

In the present work, the effect of seed pre-soaking with gallic acid (GA; 3,4,5-triphydroxyl-benzoic acid) in conferring subsequent tolerance to Cd stress in sunflower (Helianthus annuus) seedlings was investigated. Exposing sunflower seedlings to increasing Cd concentrations (5, 10 and 20 µM) caused a gradual decrease in root and shoot biomass and increased the metal accumulation in both organs. Seed pretreatment with 75 µM GA significantly restricted Cd uptake, markedly alleviated Cd-induced plant growth inhibition, and mitigated the oxidative damages caused by this metal, as compared to plants directly exposed to Cd. GA pre-soaking prior to Cd stress also enhanced catalase, ascorbate peroxidase and glutathione reductase activities, while inhibiting that of superoxide dismutase. This was associated with increased levels of total thiols and glutathione along with a decreased level of oxidized glutathione in leaves. Moreover, GA pre-soaking led to changes in leaf fatty acid composition of seedlings challenged with Cd, as evidenced by the higher total lipid content and lipid unsaturation degree. As a whole, this study provides strong arguments highlighting the potential role of GA as a growth promoter for sunflower seedlings submitted to Cd stress, notably by boosting the antioxidant defense system and improving leaf membrane stability.

Calcium and ethylene glycol tetraacetic acid mitigate toxicity and alteration of gene expression associated with cadmium stress in chickpea (Cicer arietinum L.) shoots.

In the aim to estimate the protective role of calcium (Ca) and ethylene glycol tetraacetic acid (EGTA) against cadmium (Cd)-induced damage, chickpea (Cicer arietinum L.) seeds were exposed to 200 µM Cd stress for 6 days or 3 days then subjected to co-treatment of the metal with either 100 mM CaCl2 or 100 µM EGTA for 3 additional days. The addition of Ca and EGTA improved seedling growth. This protecting effect was correlated to the alleviation of the metal-induced oxidative stress, exemplified by the reduction of hydrogen peroxide (H2O2) contents. Besides, Ca and EGTA stimulated thioredoxin (Trx) and thioredoxin reductase (NTR) activities (2.75- and 1.75-fold increase when compared to Cd-stressed, respectively) protecting, thereby, protein -SH groups from the Cd-mediated oxidation, and modulated ferredoxin (Fdx) activity to a control level. Moreover, Ca and EGTA reinstated the glutathione redox steady state, mainly via preserving a high level of glutathione reduced form (GSH). This effect coincided with the maintaining of the Cd-stimulated glutathione reductase (GR) activity and the decline of glutathione peroxidase (GPX, 43% lower than Cd-stressed shoots) activity. Ca and EGTA counteracted the inhibitory effect of Cd on the activity and gene expression of Cu/Zn-superoxide dismutase (Cu/Zn-SOD) isoenzyme and modulated the activities of catalase (CAT) and ascorbate peroxidase (APX). Overall, our results provided evidence that Ca and EGTA supplement could be a promising approach in the remediation of Cd-contaminated environment.

Nitric oxide and hydrogen sulfide protect plasma membrane integrity and mitigate chromium-induced methylglyoxal toxicity in maize seedlings.

The present study aims to analyse the potential crosstalk between nitric oxide (NO) and hydrogen sulfide (H2S) in triggering resilience of maize (Zea mays L.) seedlings to hexavalent chromium (Cr VI). Exogenous application of 500 µM sodium nitroprusside (SNP, as a NO donor) or sodium hydrosulfide (NaHS, as a H2S donor) to 9-day-old maize seedlings, countered a Cr (200 µM) -elicited reduction in embryonic axis biomass. Cr caused cellular membrane injury by enhancing the levels of superoxide and hydroxyl radicals as well as methylglyoxal, and 4-hydroxy-2-nonenal. The application of SNP or NaHS considerably improved the endogenous NO and H2S pool, decreased oxidative stress and lipid peroxidation by suppressing lipoxygenase activity and improving some antioxidant enzymes activities in radicles and epicotyls. Radicles were more affected than epicotyls by Cr-stress with enhanced electrolyte leakage and decreased proton extrusion as indicated by lesser H+-ATPase activity. H2S appeared to mitigate Cr toxicity through up-regulated H+-ATPase and glyoxalase pathways and by maintaining optimal GSH levels as downstream effects of ROS and MG suppression. Hence, H2S-mediated the regeneration of GSH pool is associated with the attenuation of MG toxicity by enhancing S-lactoglutathione and D-lactate production. Taken together, our results indicate complementary roles for H2S and GSH to strengthen membrane integrity against Cr stress in maize seedlings.

Exogenous application of hydrogen sulfide reduces chromium toxicity in maize seedlings by suppressing NADPH oxidase activities and methylglyoxal accumulation.

Chromium (Cr) represents an important source of metallic stress in plants. Working with maize (Zea mays) seedlings, we characterize the suppressive effects of exogenously applied NaHS (a hydrogen sulfide; [H2S] donor) on the toxic effects of Cr (VI). Heavy metal treatment reduced radicle and epicotyl lengths and fresh weights in seedlings at 6 and 9 days following germination. The negative Cr (200 µM) effect was countered by application with NaHS (500 µM) but this countering was reduced with the co-application of the H2S generation inhibitor hydroxylamine (HA) or the H2S scavenger hypotaurine (HT). The Cr-elicited H2O2 production was suppressed by NaHS and also by an inhibitor of the reactive oxygen species (ROS) generating NADPH oxidase (NOX). These effects were correlated with relative changes in carbomyl (-CO) and thiol (-SH) groups. Nitric oxide (NO) production increased by NaHS application with associated increase in S-nitrosoglutathione (GSNO) level, but low S-nitrosoglutathione reductase (GSNOR) activities indicating an elevated S-nitrosylation. Assessment of the role of the ascorbate-glutathione antioxidant cycle indicated that whilst ascorbate played at a best minor role, glutathione was more prominent. Methylglyoxal (MG) production was increased by Cr but reduced by NaHS through a mechanism which could be based on glutathione-S-transferase (GST) detoxification. Taken together data suggest that H2S acts to counter Cr effect in maize by NOX suppression, mostly likely by the well-characterised S-nitrosylation mechanism, as well as a reduction of MG accumulation.

Alleviation of Cr(VI)-induced oxidative stress in maize (Zea mays L.) seedlings by NO and H2S donors through differential organ-dependent regulation of ROS and NADPH-recycling metabolisms.

Chromium (Cr) contamination in soil is a growing concern in relation to sustainable agricultural production and food safety. Nitric oxide (NO) and, more recently, hydrogen sulfide (H2S) are considered to be new signalling molecules with biotechnological applications in the agronomical sector. Using 9-day-old maize (Zea mays) seedlings exposed to 200µM Cr(VI), the potential mitigating effects of exogenous NO and H2S on chromium-induced stress in maize seedlings were investigated in roots, cotyledons and coleoptiles. Analysis of Cr content, lipid peroxidation, antioxidant enzymes (catalase and superoxide dismutase isozymes), peroxisomal H2O2-producing glycolate oxidase and the main NADPH-regenerating system revealed that chromium causes oxidative stress, leading to a general increase in these activities in coleptiles and roots, with the latter organ being the most affected. However, cotyledons behaved in an opposite manner. Moreover, exogenous applications of NO and H2S to Cr-stressed maize seedlings triggered a significant response, involving the virtual restoration of the values for all these activities to those observed in unstressed seedlings, although their specific impact on ROS and NADPH-recycling metabolisms depends on the seedling organ involved. Taken together, the data indicate that gas transmitters, NO and H2S, which act as a defence against the negative effects of hexavalent chromium contamination, are alternative compounds with potential biotechnological applications.