Co-inoculation of Urea and DAP Tolerant <Emphasis Type="Italic">Sinorhizobium meliloti</Emphasis> and <Emphasis Type="Italic">Pseudomonas aeruginosa</Emphasis> as Integrated Approach for Growth Enhancement of <Emphasis Type="Italic">Brassica juncea</Emphasis>

Two plant growth promoting rhizobacteria––Sinorhizobium meliloti RMP1 and Pseudomonas aeruginosa GRC₂ were studied for integrated nutrient management to obtain improved yield of Brassica juncea. Low concentrations of urea and diammonium phosphate (DAP) stimulated the growth of both S. meliloti RMP1 and P. aeruginosa GRC₂. 1 M of urea and 0.35 M of DAP was found lethal for RMP1, while 1.3 M and 0.37 M concentrations of urea and DAP proved to be toxic for GRC₂. Lc₅₀ was observed as 0.49 M of urea and 0.15 M of DAP for RMP1, and 0.66 M urea and 0.18 M of DAP for GRC₂. Urea and DAP adaptive variants of RMP1 and GRC₂ was isolated. Adaptive bacterial variants had better growth rates at sub-lethal (Lc₅₀) concentrations of urea and DAP as compared to non-adaptive variants. They also retained plant growth promoting attributes similar to non adaptive variants. GRC₂ and RMP1 did not affect the growth of each other and were chemotactically active for DAP, urea as well as root exudates of B. juncea. Both the isolates colonized well in the rhizosphere of B. juncea, as their populations were recorded ≈5 log₁₀ cfu g⁻¹ after 120 days. Interestingly, the colonization ability was found even better when both strains were co-inoculated, as their population was recorded in the range of ≈6 log₁₀ cfu g⁻¹ after 120 days. In field trials, application of RMP1 and GRC₂ resulted in significant increase in biomass and yield of B. juncea as compared to control. However, yield was better with application of half dose and full dose of recommended fertilizers. Interestingly, the biomass as well as yield improved further when both isolates were applied together along with half dose of recommended fertilizers.     

Cloning,characterization, and expression of the <Emphasis Type="Italic">BjEXPA1</Emphasis> gene and its promoter region from <Emphasis Type="Italic">Brassica juncea</Emphasis> L.

To explore the regulation of expansin expression under heavy metal stresses, a novel expansin gene, BjEXPA1, and its upstream region were obtained from Indian mustard (Brassica juncea L.). BjEXPA1 encodes a cell wall protein of the α-subgroup of the expansin family. The expression of BjEXPA1 is regulated by exogenous phytohormones, NaCl, and heavy metals. Sequence analysis of its upstream region revealed the presence of several putative cis-acting elements, including phytohormone response, abiotic stress response, and tissue-specific elements. GUS activity under the control of the BjEXPA1 promoter allowed observation of BjEXPA1 spatial and temporal expression patterns. The reporter construct indicated that BjEXPA1 is induced by exogenous gibberellin and auxin, and inhibited by abscisic acid, CdCl₂ and PEG6000. Transgenic tobacco seedlings overexpressing BjEXPA1 showed an altered phenotype and enhanced sensitivity to cadmium. Higher H₂O₂ accumulation and extensive plasma membrane damage were detected in the leaves of transgenic seedlings when exposed to Cd, compared with wild-type plants. Our findings support the conclusion that overexpression of BjEXPA1 in tobacco renders plants susceptible to Cd stress, probably due to disruption of the growth process and/or the breakdown of cell walls.     

Drought-induced oxidative damage and antioxidant responses in peanut (<Emphasis Type="Italic">Arachis</Emphasis><Emphasis Type="Italic">hypogaea</Emphasis> L.) seedlings

Two cultivars of peanut (Arachis hypogaea L.) which were designated as resistant (Florispan) and sensitive (Gazipasa) according to their growth retardation under drought stress conditions were compared for their oxidative damage and antioxidant responses. Sixteen days-old peanut seedlings were subjected to PEG-6000 solutions of two different osmotic potentials; −0.4 and −0.8 MPa, and various growth parameters, photosystem II activity, changes in malondialdehyde (MDA), hydrogen peroxide (H₂O₂) and proline levels, activities of ascorbate peroxidase (APX), catalase (CAT), peroxidase (POX) and gluthatione reductase (GR) enzymes were determined. Both cultivars exhibited water deficit at −0.8 MPa osmotic potential of PEG-6000 and H₂O₂ levels significantly increased during exposure to −0.4 MPa osmotic potential. However, H₂O₂ levels were under control in both cultivars at exposure to −0.8 MPa osmotic potential. Significant proline accumulation was observed in the tissues of cv. Florispan at −0.8 MPa osmotic potential, whereas proline accumulation did not appear to be an essential part of the protection mechanism against drought in cv. Gazipasa. No significant variation in chlorophyll fluorescence values were detected in neither of the cultivars. Enzyme activity measurements revealed that Gazipasa copes well with lesser magnitudes of drought stress by increasing the activity of mainly APX, and during harsh stress conditions, only APX maintains its activity in the tissues. In cultivar Florispan, GR activity appears to take role in lesser magnitudes of drought stress, whereas CAT and APX activities appear to be very crucial antioxidative defenses during intense stress conditions. The results indicate that, the level of proline and activities of the enzymes CAT and APX are important mechanisms for the maintenance of drought tolerance in peanut plants.     

Characterization of <Emphasis Type="Italic">Brassica juncea</Emphasis> antioxidant potential under salinity stress

Present study characterizes the anti-oxidative defense potential of four Brassica juncea varieties, Pusa Jaikisan, Varuna, RLM-198, and CS-52, differing in their ability to withstand salinity stress. 7-day-old seedlings raised in MS medium supplemented with 0, 50, 100, and 150 mM NaCl were used to monitor changes in the growth profile, level of stress marker molecules, and activities of important antioxidant enzymes. Increasing NaCl concentration resulted in a significant (P ≤ 0.05) reduction of shoot fresh and dry mass and vigor index in all the varieties tested. Maximum reduction in growth was recorded for RLM-198 while CS-52 maintained better growth characteristics. Varuna and RLM-198 exhibited a limited increase in superoxide dismutase, ascorbate peroxidase, and total peroxidase activity under increasing salinity. These varieties also recorded maximum salt stress-induced damage in terms of increased lipid peroxidation, H₂O₂ content, and electrolyte leakage. On the other hand, CS-52 recorded maximum proline accumulation with minimum levels of H₂O₂, electrolyte leakage, and malondialdehyde contents. With increasing salinity stress, CS-52 recorded maximal increase in the activity of antioxidant enzymes. However, catalase activity did not correlate with alterations in H₂O₂ levels under stress. Interestingly, a lower superoxide dismutase:ascorbate peroxidase ratio in CS-52 correlated with stress tolerance trait, while a comparatively higher superoxide dismutase:ascorbate peroxidase ratio in RLM-198 marked the susceptible nature of the variety. Our results propose that superoxide dismutase:ascorbate peroxidase ratio is the critical factor, determining the degree of stress tolerance in Brassica juncea.     

Induction of phytochelatins and antioxidant defence system in <Emphasis Type="Italic">Brassica juncea</Emphasis> and <Emphasis Type="Italic">Vigna radiata</Emphasis> in response to chromium treatments

Growth performance, chromium (Cr) accumulation potential and induction of antioxidative defence system and phytochelatins (PCs) were studied in hydroponically grown Brassica juncea (Indian mustard) and Vigna radiata (mungbean) at various levels of Cr treatments (0, 50, 100, 200 μM Cr). B. juncea accumulated twofolds and threefolds higher Cr in root and shoot, respectively than in V. radiata. Compared to B. juncea, V. radiata was found to be particularly sensitive to Cr as observed by the severity and development of Cr toxicity symptoms and decreased growth. Induction of PC and enzymes of antioxidant defence system were monitored as plant’s primary and secondary metal detoxifying responses, respectively. There was induction of PC and enzymes of antioxidant defence system in both the plants. PCs were induced significantly in roots and shoot of both the plants at all the levels of Cr treatments. Significantly higher activities of superoxide dismutase (SOD), ascorbate peroxidase (APX), catalase (CAT) and glutathione reductase (GR) were observed in shoot of B. juncea than V. radiata at all the levels of Cr treatments. Induction of PCs along with antioxidant defence system in response to Cr stress suggests the cumulative role of PCs and antioxidants in conferring tolerance against accumulated Cr in B. juncea, and thereby signifies the suitability of this plant as one of the potential remediators of Cr.     

Changes in growth,photosynthetic capacity and ionic relations in spring wheat (<Emphasis Type="Italic">Triticum aestivum</Emphasis> L.) due to pre-sowing seed treatment with polyamines

The influence of pre-sowing seed treatment with polyamines (2.5 mM putrescine, 5.0 mM spermidine and 2.5 mM spermine) on growth, photosynthetic capacity, and ion accumulation in two spring wheat (Triticum aestivum L.) cultivars MH-97 (intolerant) and Inqlab-91 (tolerant) was examined. The primed seeds of each treatment and non-primed seeds were sown in a field containing 15 dS m⁻¹ NaCl. Although all three polyamines were effective in improving shoot growth and grain yield in both cultivars under saline conditions, the effect of spermine was very pronounced particularly in improving grain yield. Different priming agents did not affect the net CO₂ assimilation rate and transpiration rate of either cultivar. However, pre-treatment with spermidine increased stomatal conductance (g_s) in the tolerant cultivar, whereas with spermine stomatal conductance decreased in the intolerant cultivar under salt stress. Priming agents had different effects on the accumulation of different ions in wheat plant tissues. When spermidine and distilled water were used as priming agents, they were effective in reducing shoot [Na⁺] in the tolerant and intolerant cultivars, respectively under saline conditions. Although all priming agents caused an increase in shoot [K⁺], distilled water was more effective in improving shoot [K⁺] in both cultivars under salt stress. Pre-treatment with spermidine was very effective in reducing shoot [Cl⁻] under saline conditions particularly in the tolerant cultivar. However, the pattern of accumulation of different ions in roots due to different seed priming treatments was not consistent in either cultivar except that root Na⁺ decreased due to priming with spermine and spermidine in the intolerant and tolerant cultivars under saline conditions. In conclusion, although all three priming agents, spermine, spermidine and putrescine, were effective in alleviating the adverse effect of salt stress on wheat plants, their effects on altering the concentration of different ions and growth were different in the two cultivars differing in salt tolerance.     

Effect of salt stress on physiological and antioxidative responses in two species of <Emphasis Type="Italic">Salicornia</Emphasis> (<Emphasis Type="Italic">S. persica</Emphasis> and <Emphasis Type="Italic">S. europaea</Emphasis>)

The effects of salt stress on growth parameters, free proline content, ion accumulation, lipid peroxidation, and several antioxidative enzymes activities were investigated in S. persica and S. europaea. The seedlings were grown for 2 months in half-strength Hoagland solution and treated with different concentrations of NaCl (0, 85, 170, 340, and 510 mM) for 21 days. The fresh and dry weights of both species increased significantly at 85 and 170 mM NaCl and decreased at higher concentrations. Salinity increased proline content in both the species as compared to that of control. Sodium (Na⁺) content in roots and shoots increased, whereas K⁺ and P_i content in both organs decreased. At all NaCl concentrations, the total amounts of Na⁺ and K⁺ were higher in shoots than in roots. Malondialdehyde (MDA) content declined at moderate NaCl concentrations (85 and 170 mM) and increased at higher levels. With increased salinity, superoxide dismutase (SOD), catalase (CAT), and guaiacol peroxidase (GPX) activities also increased gradually in both species. In addition, it seems that GPX, CAT, and SOD activities play an essential protective role in the scavenging reactive oxygen species (ROS) in both species. Native polyacrylamide gel electrophoresis (PAGE) indicated different isoform profiles between S. persica and S. europaea concerning antioxidant enzymes. These results showed that S. persica exhibits a better protection mechanism against oxidative damage and it is more salt-tolerant than S. europaea possibly by maintaining and/or increasing growth parameters, ion accumulation, and antioxidant enzyme activities.     

Effects of pH on NaCl tolerance of American elm (<Emphasis Type="Italic">Ulmus americana</Emphasis>) seedlings inoculated with <Emphasis Type="Italic">Hebeloma crustuliniforme</Emphasis> and <Emphasis Type="Italic">Laccaria bicolor</Emphasis>

In the present study, we investigated the effects of pH treatments on NaCl tolerance in mycorrhizal and non-mycorrhizal American elm. American elm (Ulmus americana) seedlings were inoculated with Hebeloma crustuliniforme, Laccaria bicolor or with both mycorrhizal fungi and subsequently subjected to different pH solutions (pH 3, 6 and 9) containing 0 mM (control) and 60 mM NaCl for 4 weeks. Inoculation with the mycorrhizal fungi did not have a large effect on seedling dry weights when the pH and NaCl treatments were considered independently. However, when the inoculated seedlings were treated with 60 mM NaCl at pH 3 or 6, shoot to root ratios and root hydraulic conductivity were higher compared with non-inoculated plants, likely reflecting changes in seedling water flow properties. At pH 6, transpiration rates were about twofold lower in non-inoculated plants treated with NaCl compared with non-treated controls. For NaCl-treated H. crustuliniforme- and L. bicolor-inoculated plants, the greatest reduction of transpiration rates was at pH 9. Treatment with 60 mM NaCl reduced leaf chlorophyll concentrations more in non-inoculated compared with inoculated plants, with the greatest, twofold, decrease occurring at pH 6. At pH 3, root Na concentrations were higher in inoculated than non-inoculated seedlings; however, there was no effect of inoculation on root Na concentrations at pH 6 and 9. Contrary to the roots, the leaves of inoculated plants had lower Na concentrations at pH 6 and 9, but not at pH 3. The results point to an interaction between ECM fungi and root zone pH for salt tolerance of American elm.     

Molecular cloning and characterization of the promoter for the multiple stress-inducible gene <Emphasis Type="Italic">BjCHI1</Emphasis> from <Emphasis Type="Italic">Brassica juncea</Emphasis>

We have previously isolated a Brassica juncea cDNA encoding a novel chitinase BjCHI1 with two chitin-binding domains (Zhao and Chye in Plant Mol Biol 40:1009–1018, 1999). The expression of BjCHI1 was highly inducible by methyl jasmonate (MeJA) treatment, wounding, caterpillar feeding, and pathogenic fungal infection. These observations suggest that the promoter of BjCHI1 gene might contain specific cis-acting elements for stress responses. Here, we report the cloning and characterization of the BjCHI1 promoter. A 1,098 bp BjCHI1 genomic DNA fragment upstream of the ATG start codon was isolated by PCR walking and various constructs were made by fusing the BjCHI1 promoter or its derivatives to β-glucuronidase reporter gene. The transgenic Arabidopsis plants showed that the BjCHI1 promoter responded to wounding and MeJA treatment, and to treatments with either NaCl or polyethyleneglycol (PEG 6000), indicating that the BjCHI1 promoter responses to both biotic and abiotic stresses. A transient gene expression system of Nicotiana benthamiana leaves was adopted for promoter deletion analysis, and the results showed that a 76 bp region from −695 to −620 in the BjCHI1 promoter was necessary for MeJA-responsive expression. Furthermore, removal of a conserved T/G-box (AACGTG) at −353 to −348 of the promoter greatly reduced the induction by MeJA. This is the first T/G-box element identified in a chitinase gene promoter. Gain-of-function analysis demonstrated that the cis-acting element present in the 76 bp region requires coupling with the T/G-box to confer full magnitude of BjCHI1 induction by MeJA.     

Chloroplast-located <Emphasis Type="Italic">BjFer1</Emphasis> together with anti-oxidative genes alleviate hydrogen peroxide and hydroxyl radical injury in cytoplasmic male-sterile <Emphasis Type="Italic">Brassica juncea</Emphasis>

We studied how plant cell modulated redox homeostasis in cytoplasmic male-sterility (CMS) Brassica juncea. The CMS Brassica juncea was identified to be mutated in several mitochondrial genes that suggested the changes of cell redox homeostasis. We observed that it was not associated with increased oxidative stress as shown by decreased H₂O₂ and ^∙OH contents in this type of CMS. The expressions of several anti-oxidative genes were up-regulated in 5-day-old seedlings of CMS than MF lines under light and dark conditions. The mitochondrial alternative oxidase pathway was not activated, as indicated by no increased expression of AOX1a gene in CMS. Interestingly, the expression of Ferritin1 gene was markedly activated in 5-day-old seedlings of CMS than MF line under light and dark conditions. Consequently, we detected increased content of total iron in 30-day-old leaves in CMS than MF line. We isolated Ferritin1 orthologous gene from Brassica juncea, which was targeted to the chloroplast and induced by Fe-citrate and H₂O₂, not ABA. Taken together, we proposed that increased expressions of BjFer1 and several antioxidant genes protected cell from oxidative stress in CMS Brassica juncea.     

Seed priming with sorghum extracts and benzyl aminopurine improves the tolerance against salt stress in wheat (<Emphasis Type="Italic">Triticum aestivum</Emphasis> L.)

Salt stress impedes the productivity of wheat (Triticum aestivum L.) in many parts of the world. This study evaluated the potential role of benzyl aminopurine (BAP) and sorghum water extract (SWE) in improving the wheat performance under saline conditions. Seeds were primed with BAP (5 mg L^?1), SWE (5% v/v), BAP + SWE, and distilled water (hydropriming). Soil filled pots maintained at the soil salinity levels of 4 and 10 dS m^?1 were used for the sowing of primed and non-primed seeds. Salt stress suppressed the wheat growth; seed priming treatments significantly improved the wheat growth under optimal and suboptimal conditions. Total phenolics, total soluble sugars and proteins, α-amylase activity, chlorophyll contents, and tissue potassium ion (K⁺) contents were increased by seed priming under salt stress; while, tissue sodium ion (Na⁺) contents were decreased. Seed priming with SWE + BAP was the most effective in this regard. Under salt stress, the tissue Na⁺ contents were reduced by 5.78, 28.3, 32.2, 36.7% by hydropriming, seed priming with SWE, seed priming with BAP, and seed priming with SWE + BAP, respectively over the non-primed control. Effectiveness of seed priming techniques followed the order SWE + BAP > BAP > SWE > Hydropriming. In conclusion, seed priming with SWE + BAP may be opted to improve the tolerance against salt stress in wheat.     

Effect of Drought on the Growth and Survival of the Stress-Tolerant Bacterium <Emphasis Type="Italic">Rhizobium</Emphasis> sp. NBRI2505 <Emphasis Type="Italic">sesbania</Emphasis> and Its Drought-Sensitive Transposon Tn<Emphasis Type="Italic">5</Emphasis> Mutant

Studies were conducted to elucidate the nature of drought tolerance in the bacterium Rhizobium sp. NBRI2505 sesbania and its transposon Tn5 induced mutant to assess the role of salt, pH, and temperature stresses in contributing to drought tolerance, and to correlate drought tolerance and symbiotic effectiveness. Rhizobium sp. NBRI2505 sesbania tolerated yeast extract mannitol broth (YEB) containing 28% salt (NaCl; wt/vol) for up to 18 h of incubation at 30°C, survived a 2-h incubation in YEB at 65°C, and when subjected to drought stress, tolerated YEB containing 45% polyethylene glycol 6000 (PEG; wt/vol) for up to 5 days of incubation at 30°C. One drought-sensitive mutant Rhizobium sp. NBRI2505 sesbania T112 (T112) containing a single Tn5 insertion was selected after screening about 10,000 clones. T112 was specifically defective in its tolerance for drought: when subjected to drought stress, it tolerated YEB containing 45% PEG for up to 2 days of incubation at 30°C. T122 mutant was also more sensitive to the heat and desiccation stresses, compared with Rhizobium sp. NBRI2505 sesbania in the presence of 45% PEG. Our results demonstrated a positive effect of calcium on the survival of Rhizobium sp. sesbania under acidic stress conditions. The observed enhanced survival at pH 3 of Rhizobium sp. NBRI2505 sesbania and T112 in the presence of 5% CaCO₃ suggests the requirement of calcium for growth and survival, which may have an ecological significance in acidic soils. Mutant strain T112 produced ineffective symbiosis with the plant host in the presence of 2.5 and 5% PEG, indicating that drought tolerance is required for effective symbiosis. Received: 11 January 2002 / Accepted: 18 February 2002     

Role of osmotic stress in ion accumulation and physiological responses of mycorrhizal white spruce (<Emphasis Type="Italic">Picea glauca</Emphasis>) and jack pine (<Emphasis Type="Italic">Pinus banksiana</Emphasis>) to soil fluoride and NaCl

To examine the mechanisms of earlier reported alleviation of fluoride injury in ectomycorrhizal plants by NaCl, jack pine (Pinus banksiana) and white spruce (Picea glauca) seedlings were subjected to 1 mM and 5 mM KF in the presence of either 60 mM NaCl or 10% polyethylene glycol 3350 (PEG) for 2 weeks. Before the treatments, seedlings had either been inoculated with the ectomycorrhizal fungus Suillus tomentosus or remained non-inoculated. The inoculation with S. tomentosus reduced Na uptake by shoots and roots of jack pine seedling and by roots of white spruce that were treated with 60 mM NaCl. Mycorrhizal associations also drastically decreased fluoride uptake by jack pine seedlings, but did not affect shoot fluoride concentrations in white spruce. When NaCl was replaced by PEG in the 5 mM KF treatment solution, shoot fluoride concentrations were reduced by more than twofold without corresponding reductions in transpiration rates in mycorrhizal and non-mycorrhizal white spruce seedlings. When fluoride was present in the treatment solution, Na concentrations were lower in shoots and roots of both jack pine and white spruce mycorrhizal and non-mycorrhizal seedlings. The results suggest that Suillus tomentosus may help alleviate the effects of soil fluoride and salinity in jack pine and that fluoride uptake in white spruce is sensitive to osmotic stress.     

<Emphasis Type="Italic">n</Emphasis>-Alkanes variability in the diazotrophic cyanobacterium <Emphasis Type="Italic">Anabaena cylindrica</Emphasis> in response to NaCl stress

n-Alkanes pattern in response to NaCl stress has been studied in the cyanobacterium Anabaena cylindrica. Saturated hydrocarbons were separated and identified by gas chromatography-mass spectrometry (GC-MS) using serially coupled capillary column. Light chain n-alkanes in the range of C₉–C₁₇ (43%) and heavy chain n-alkanes in range of C₁₇–C₂₃ (34%) and C₂₃–C₃₁ (23%) were identified as the major components of total hydrocarbons in the NaCl adapted cells of A. cylindrica. In contrast, NaCl-untreated cells of A. cylindrica had dominance of only long chain n-alkanes in the range of C₂₃–C₃₁ comprising about 94% of its total n-alkanes. The persistence of high level (43%) of short chain n-alkanes (C₉–C₁₇) in NaCl adapted cells of A. cylindrica as compared to its negligible level (0.2%) in NaCl untreated counterpart clearly indicates that NaCl stress causes the A. cylindrica to shift towards the synthesis of short chain n-alkanes.     

Molecular Characterization of <Emphasis Type="Italic">Arabidopsis</Emphasis> and <Emphasis Type="Italic">Brassica juncea</Emphasis> Cu/Zn-Superoxide Dismutases Reveals Their Regulation of Shoot Regeneration

Superoxide dismutases (SODs) are ubiquitous metalloenzymes that catalyze the dismutation of superoxide radicals (O₂^-) to molecular oxygen (O₂) and hydrogen peroxide (H₂O₂). In this study we characterized an Arabidopsis thaliana CuZnSOD (CSD1), a close ortholog of a previously identified Brassica juncea CuZnSOD (MSOD1). CSD1 and other two homologs CSD2 and CSD3 were spatially regulated in Arabidopsis, and CSD1 exhibited distinct expression patterns in response to different stress treatments. To investigate the in vivo function of SOD, transgenic Arabidopsis plants, expressing sense and antisense MSOD1 RNAs, were generated and those with altered SOD activity were selected for further characterization. Although SOD transgenic plants exhibited normal phenotypes, the shoot regeneration response in transgenic explants was significantly affected by the modulated SOD activity and the corresponding H₂O₂ levels. Transgenic explants with downregulated SOD activity were poorly regenerative, whereas those with upregulated SOD activity were highly regenerative. These results suggest that shoot regeneration in vitro is regulated by the SOD activity.     

Phosphomannose-isomerase (<Emphasis Type="Italic">pmi</Emphasis>) gene as a selectable marker for <Emphasis Type="Italic">Agrobacterium</Emphasis>-mediated transformation of rapeseed

Nodal segments (4 ± 1 mm long) of Hibiscus moscheutos (hardy hibiscus) were excised from in vitro proliferating microshoots and utilized to evaluate initial factors involved in bulk alginate encapsulation. The factors evaluated were; storage vessel type, volume and multiple rinse effects of CaCl₂ solutions, and sodium alginate concentrations (2.5, 2.75, 3.0 or 3.25%) for bulk alginate encapsulation. Results indicate that vessels utilized for bulk alginate encapsulation should have a lower base area (L × W) to height ratio to reduce the amount of alginate matrix shrinkage resulting in exposure of nodal segments. Increased volumes and multiple 50 mM CaCl₂ solution rinses did not have an effect on alginate solidification. Shoot length, root number, and root length significantly decreased in a linear manner from nodal explants stored for 4 weeks with increasing concentrations of sodium alginate. This research suggests an innovative technique for alginate encapsulation of H. moscheutos utilizing bulk methods as an alternative to single bead alginate encapsulation.     

Interaction of silicon and cadmium in <Emphasis Type="Italic">Brassica juncea</Emphasis> and <Emphasis Type="Italic">Brassica napus</Emphasis>

The objective of this study was to determine the effect of silicon (Si) and cadmium (Cd) on root and shoot growth and Cd uptake in two hydroponically cultivated Brassica species (B. juncea (L.) Czern. cv. Vitasso and B. napus L. cv. Atlantic). Both species are potentially usable for phytoextraction. Inhibitory effects of Cd on root elongation were diminished by the impact of Si. Primary roots elongation in the presence of Cd + Si compared with Cd was stronger and the number of lateral roots was lower in B. juncea than in B. napus. Cd content per plant was higher in B. napus roots and shoots compared with B. juncea. Suberin lamellae were formed closer to the root apex in Cd + Si than in Cd treated plants and this effect was stronger in B. napus than in B. juncea. Accelerated maturation of endodermis was associated with reduced Cd uptake. Cd decreased the content of chlorophylls and carotenoids in both species, but Si addition positively influenced the content of photosynthetic pigments which was higher in B. napus than in B. juncea. Si enhanced more substantially translocation of Cd into the shoot of B. napus than of B. juncea. Based on our results B. napus seems to be more suitable for Cd phytoextraction than B. juncea because these plants produce more biomass and accumulate higher amount of Cd. The protective effect of Si on Cd treated Brassica plants could be attributed to more extensive development of suberin lamellae in endodermis.     

Effect of NaCl and CaCl<Subscript>2</Subscript> on the antioxidant mechanism of leaves and stems of the rootstock CAB-6P (<Emphasis Type="Italic">Prunus cerasus</Emphasis> L.) under in vitro conditions

The effect of salinity on the non-enzymic and enzymic antioxidant activity, shoot proliferation and nutrient accumulation was studied in in vitro cultures of the rootstock CAB-6P (Prunus cerasus L.). Three concentrations (0, 30 and 60 mM) of NaCl or CaCl₂ were added to a modified MS medium. Between the two salt treatments used, only the explants treated with CaCl₂ presented significant decrease in growth parameters. The concentrations of Na⁺ and Cl⁻ in the explants treated with NaCl were increased, as NaCl in the culture medium increased. Furthermore, in the explants treated with CaCl₂ the concentrations of Ca²⁺ and Cl⁻ were increased while that of K⁺ decreased, as CaCl₂ concentration increased. The activity of peroxidase in leaves as well as the number of its anionic isoforms was increased under 30 mM CaCl₂ as well as 60 mM NaCl or CaCl₂. On the contrary, increasing salinity, from 0 to 60 mM CaCl₂, resulted in a reduction of the catalase activity in leaves followed by disappearance of the only one catalase isoform that was detected in leaves (60 mM CaCl₂). In the stems of the explants treated with NaCl the peroxidase activity was reduced. In the stems and leaves of the explants grown in saline substrate the non-enzymic antioxidant activity was significantly increased. The results suggest that the stems and leaves of CAB-6P explants presented variable antioxidant responses that were depended on the salt form used. The contribution of enzymic and non-enzymic protection mechanisms to the adaptation of CAB-6P explants under salinity stress is discussed.