Zinc oxide nanoparticles (ZnONPs) alleviate heavy metal-induced toxicity in Leucaena leucocephala seedlings: A physiochemical analysis.

The present study describes the role of zinc oxide nanoparticles (ZnONPs) in reversing oxidative stress symptoms induced by heavy metal (Cd and Pb) exposure in Leucaena leucocephala (Lam.) de Wit. Seedling growth was significantly enhanced with the augmentation of ZnONPs following Cd and Pb exposure. Heavy metal accumulations were recorded as 1253.1 mg Cd per kg DW and 1026.8 mg Pb per kg DW for the respective treatments. Results demonstrated that ZnONPs augmentation caused an increase in photosynthetic pigment and total soluble protein contents while a significant decrease in malondialdehyde (MDA-lipid peroxidation) content in leaves. Antioxidative enzymes such as superoxide dismutase (SOD), catalase (CAT) and peroxidase (POX) were, in turn, elevated in heavy metal-exposed leaves amended with ZnONPs. The ameliorating effect of ZnO nanoparticles on oxidative stress induced toxicity was also confirmed by the reduced MDA content and the elevated level of antioxidative enzyme activities in leaf tissues of L. leucocephala seedlings. Further, addition of ZnONPs in combination with Cd and Pb metals induced distinct genomic alterations such as presence of new DNA bands and/or absence of normal bands in the RAPD pattern of the exposed plants. This study uniquely suggests a potential role of zinc oxide nanoparticles in the remediation of heavy metal contaminated media.

Enhanced plant growth promoting role of phycomolecules coated zinc oxide nanoparticles with P supplementation in cotton (Gossypium hirsutum L.).

This report focuses on application of zinc oxide nanoparticles (ZnONPs) carrying phycomolecule ligands as a novel plant growth promoter aimed at increasing the crop productivity. The present investigation examined the effect of ZnONPs on plant growth characteristics, and associated biochemical changes in cotton (Gossypium hirsutum L.) following growth in a range of concentrations (25-200 mg L-l ZnONPs) in combination with 100 mM P in a hydroponic system. Treated plants registered an increase in growth and total biomass by 130.6% and 131%, respectively, over control. Results demonstrated a significant increase in the level of chlorophyll a (141.6%), b (134.7%), carotenoids (138.6%), and total soluble protein contents (179.4%); at the same time, a significant reduction (68%) in the level of malondialdehyde (MDA) in leaves with respect to control. Interestingly, a significant increase in superoxide dismutase (SOD, 264.2%), and peroxidase (POX, 182.8%) enzyme activities followed by a decrease in the catalase (CAT) activity, in response to above treatments. These results suggest that bioengineered ZnONPs interact with meristematic cells triggering biochemical pathways conducive to an accumulation of biomass. Further investigations will map out the mode of action involved in growth promotion.

Impact assessment of mercury accumulation and biochemical and molecular response of Mentha arvensis: a potential hyperaccumulator plant.

The present study was focused on examining the effect of Hg oxidative stress induced physiochemical and genetic changes in M. arvensis seedlings. The growth rate of Hg treated seedlings was decreased to 56.1% and 41.5% in roots and shoots, respectively, compared to the control. Accumulation of Hg level in both roots and shoots was increased with increasing the concentration of Hg. Superoxide dismutase (SOD), catalase (CAT), and ascorbate peroxidase (APX) activities were found to be increased with increasing the Hg concentration up to 20 mg/L; however, it was decreased at 25 mg/L Hg concentration. The POX enzyme activity was positively correlated with Hg dose. The changes occurring in the random amplification of ploymorphic DNA (RAPD) profiles generated from Hg treated seedlings included variations in band intensity, disappearance of bands, and appearance of new bands compared with the control seedlings. It was concluded that DNA polymorphisms observed with RAPD profile could be used as molecular marker for the evaluation of heavy metal induced genotoxic effects in plant species. The present results strongly suggested that Mentha arvensis could be used as a potential phytoremediator plant in mercury polluted environment.

Genes induced in response to mercury-ion-exposure in heavy metal hyperaccumulator Sesbania drummondii.

Sesbania drummondii plants have been recognized as a potential mercury (Hg) hyperaccumulator. To identify genes modulated by Hg, two suppressive subtraction hybridization (SSH) cDNA libraries (forward and reverse) were constructed. A total of 348 differentially expressed clones were isolated and 95 of them were identified as Hg responsive. Reverse Northern results showed that 31 clones from forward library were down-regulated and 64 clones from reverse library were up-regulated in Hg-treated plants. Sixty-seven of them showed high homology to genes with known or putative function, and 28 were uncharacterized genes. Two full-length cDNAs coding for a putative metallothionein type 2 protein (SdMT2) and an auxin responsive protein (SdARP) were isolated and characterized. The expression levels of SdMT2 and SdARP increased 3- and 5-fold, respectively. Results suggest that up-regulated expression of SdARP may contribute to the survival of Sesbania plants under mercury stress, whereas SdMT2 is likely to be involved in alleviation of Hg toxicity. The possible correlation between gene expression and heavy metal tolerance of Sesbania plants is discussed.

Accumulation, speciation and cellular localization of copper in Sesbania drummondii.

Growth, accumulation and intracellular speciation and distribution of copper (Cu) in Sesbania drummondii was studied using scanning-electron microscopy (SEM), X-ray absorption near edge structure (XANES) and extended X-ray absorption fine structure (EXAFS). The growth of seedlings was assessed in terms of biomass accumulation. The growth of the seedling was enhanced by 73.5% at a low Cu concentration (50 mg l-1) compared to the control treatment. Additionally, seedling growth was inhibited by 18% at 300 mg l-1 Cu with respect to the control. Copper concentration in roots and shoots was increased with increasing Cu concentration in the growth solution. The accumulation of Cu was found to be higher in roots than in the shoots. At a concentration of 300 mg l-1 Cu, the roots accumulated 27,440 mg Cu kg-1 dry weight (dw) while shoots accumulated 1282 mg Cu kg-1 dw. Seedlings were assessed for photosynthetic activity by measuring chlorophyll a fluorescence parameters: Fv/Fm and Fv/F0 values. Photosynthetic integrity was not affected by any of the Cu treatments. The X-ray absorption spectroscopic (XAS) studies showed that Cu was predominantly present as Cu(II) in Sesbania tissue. In addition, from the XAS studies it was shown that the Cu exists in a mixture of different coordination states consisting of Cu bound to sugars and small organic acids with some possible precipitated copper oxide. From the EXAFS studies, the coordination of Cu was determined to have four equatorial oxygen(nitrogen) ligands at 1.96 A and two axial oxygen ligands at 2.31 A. Scanning-electron microscopy studies revealed the distribution of Cu within the seedlings tissues, predominantly accumulated in the cortical and vascular (xylem) regions of root tissues. In the stem, most of the Cu was found within the xylem tissue. However, the deposition of Cu within the leaf tissues was in the parenchyma. The present study demonstrates the mechanisms employed by S. drummondii for Cu uptake and its biotransformation.

Identification of lead-regulated genes by suppression subtractive hybridization in the heavy metal accumulator Sesbania drummondii.

Heavy metal contamination of soils is of widespread occurrence as a result of human, agricultural and industrial activities. Among heavy metals, lead is a potential pollutant that readily accumulates in soils and sediments. Although lead is not an essential element for plants, it gets easily absorbed and accumulated in Sesbania drummondii, which exhibits a significant level of tolerance to lead. The response of a metal tolerant plant to heavy metal stress involves a number of biochemical and physiological pathways. To investigate the overall molecular response of a metal-tolerant plant to lead exposure, suppression subtractive hybridization (SSH) was used to construct a cDNA library enriched in lead induced mRNA transcripts from lead-tolerant Sesbania. Screening the library by reverse Northern analysis revealed that between 20 and 25% of clones selected from the library were differentially regulated in lead treated plants. After differential screening, we isolated several differentially expressed cDNA clones, including a type 2 metallothionein (MT) gene which is involved in detoxification and homeostasis and shown to be differentially regulated in lead treated plants. The data from the reverse Northern analysis was further confirmed with conventional Northern analysis of a select group of genes including MT, ACC synthase/oxidase, cold-, water stress-, and other abiotic stress-induced genes, which are up-regulated rapidly in response to lead treatment. The mRNA levels of MT increased substantially after lead treatment indicating a potential role for it under lead stress in Sesbania. The present results show that SSH can serve as an effective tool for isolating genes induced in response to lead heavy metal tolerance in Sesbania. A better understanding of lead induced gene expression in Sesbania should help select candidates associated with remediation of heavy metal toxicity. The possible link between this result and the heavy-metal response of plants is discussed.

Cadmium accumulation and antioxidative responses in the Sesbania drummondii callus.

The effect of cadmium (Cd) on growth, accumulation, and antioxidative response was studied in Sesbania drummondii callus, cultivated on different concentrations of Cd (0-250 microM) for four weeks. Callus growth was comparable to that of the control for concentrations up to 50 microM Cd; however, concentrations higher than 50 microM affected growth. A concentration of 100 microM Cd inhibited growth by 16%, with respect to control. Cd concentration in callus increased with increasing Cd concentrations in the growth medium. Callus accumulated 530 mg Cd kg(-1) of their dry weight at 100 microM Cd concentration. Sesbania callus responded to Cd-induced oxidative stress by modulating antioxidants (glutathione and other non-protein thiols) level and antioxidative enzymes: superoxide dismutase (SOD), ascorbate peroxidase (APX), and glutathione reductase (GR). The content of the glutathione (GSH) and GSH/GSSG ratio first increased up to a concentration of 50 microM Cd and then decreased. The content of other non-protein thiols significantly increased with increasing Cd concentrations in the growth medium. The activities of antioxidative enzymes, SOD, APX, and GR, followed the same trends as antioxidants first increasing up to a concentration of 50 microM Cd and then decreasing. These results suggest that antioxidative defense mechanisms play a significant role in Cd detoxification and accumulation in Sesbania drummondii.

Photoautotrophic tobacco cells adapted to grow at high salinity.

Photoautotrophic tobacco (Nicotiana tabacum var. Wisconsin 38) cell cultures were gradually adapted to grow in media containing the normally inhibitory concentration of 20 g l-1 NaCl. Both salt-adapted cultures maintained in 20 g l-1 NaCl (P20) and salt-unadapted (P0) cultures demonstrated similar chloroplast morphology and similar growth characteristics on a dry weight basis, but P20 cells showed reduced growth on a fresh weight basis compared to P0 cells. Compared to P0 cells, intracellular sucrose levels were significantly higher in P20 cells while starch levels in P0 cells were significantly higher than in P20 cells. Levels of intracellular and extracellular reducing sugars, and chlorophyll accumulated to the same degree in P20 and P0 cells, but accumulation was delayed by approximately 13 days in P20 cells. O2 evolution and14[CO2] fixation was more resistant to inhibition by NaCl in P20 cells than in P0 cells. However, significant changes in the abundance of thylakoid membrane proteins could not be demonstrated between P20 and P0 cells although higher levels of Rubisco on a per milligram chlorophyll basis were observed in P0 compared to P20 chloroplasts.

A thin layer chromatographic technique for detecting inducers of Agrobacterium virulence genes in corn, wheat and rye.

A method is described for detecting plant metabolites capable of inducing the virulence genes of Agrobacterium tumefaciens. The method uses A. tumefaciens containing a plasmid with an inducible virulence gene fused to a galactosidase gene (virE::lacZ). Thin layer chromatography plates are overlayed with agar containing the indicator bacterium and a chromogenic galactoside (X-gal). Virulence gene inducing plant metabolites induce galactosidase which releases an aglycone readily oxidized by air to a blue pigmented zone at the Rf of the inducer. The method has been used to demonstrate the presence of virulence gene inducers in corn, wheat and rye. The uninduced background level of galactosidase also permits detection of bacterial growth inhibitors after a longer incubation period.

Corn metabolites affect growth and virulence of Agrobacterium tumefaciens.

Homogenates of corn seedlings inhibit both growth of Agrobacterium tumefaciens and induction of its Ti plasmid virulence (vir) genes by acetosyringone (AS). The heat-labile inhibitor has been identified as 2,4-dihydroxy-7-methoxy-2H-1,4-benzoxazin-3(4H)-one (DIMBOA), present in 2-week-old seedlings (B73) at a concentration of 1.5 mM or greater. A concentration of 0.3 mM DIMBOA is sufficient to block growth of A. tumefaciens completely for 220 hr. DIMBOA at 0.1 mM concentration completely inhibited vir gene induction by 100 microM AS and reduced growth rate by 50%. Thus, DIMBOA can be expected to have a significant effect on attempts to transform corn by using A. tumefaciens as a vector.