Accumulation of low-molecular-weight organic acids in roots and leaf segments of Zea mays plants treated with cadmium and copper

Maize seedlings grown in a nutrient solution were treated with Cd (50, 100 µM) or Cu (50, 100 µM). Roots and separated leaf sections (from the youngest––basal, through the middle––mature, to the oldest––apical) were analysed. Shoot and root fresh weight, and root net growth rates were reduced significantly after Cu application in comparison to Cd. Copper (50, 100 µM) and cadmium (100 µM) decreased the sum of chlorophyll and total carotenoid pools mostly in mature and old leaf sections. The concentration of Cu and Cd increased in the old and mature leaf sections. Analysis of organic acids showed that both metals differently influenced the low-molecular-weight organic acids (LMWOAs) content in maize leaf sections. In mature sections, the excess Cu increased the succinate and tartrate contents, whereas in the young ones mainly the tartrate level was elevated. Cadmium elevated citrate accumulation in mature and old sections, compared to the control plants. Malate, the main LMWOA in maize leaves, increased only after addition of 100 µM of Cd (mature and old sections) or 50 µM of Cu (old sections). Analysis of LMWOAs in roots showed that the excess of Cd or Cu induced higher accumulation of tartrate and malate and, additionally, copper increased the citrate content.     

Nickel and zinc effects,accumulation and distribution in ruderal plants Lepidium ruderale and Capsella bursa-pastoris

Ruderal plants can grow in polluted areas, but little is known about heavy metal accumulation and distribution in them. Here Ni and Zn accumulation, distribution and effects were investigated in Lepidium ruderale and Capsella bursa-pastoris grown at 5–30 µM Ni(NO₃)₂ or 10–80 µM Zn(NO₃)₂. Metal contents were measured by flame atomic absorption spectrophotometry and tissue distribution of metals was studied histochemically. Ni was more toxic than Zn for both plants. When metal-induced growth-inhibiting effects were compared at various metal concentrations in solution, L. ruderale was more tolerant to Ni, whereas C. bursa-pastoris to Zn. However, when compared at similar Zn or Ni contents in roots, root growth of C. bursa-pastoris was more tolerant than that of L. ruderale. On the contrary, at similar Zn or Ni contents in shoots, shoot growth of L. ruderale was more tolerant. Both plants are excluders maintaining low metal levels in shoots. In roots, Ni located in protoplasts while Zn was also detected in cell walls. Metal accumulation in root apices resulted in growth inhibition. Ni accumulation in root cortex constrained metal translocation into central cylinder and then to shoots, where it located only in conductive tissues and epidermis, particularly in leaf trichomes of C. bursa-pastoris. Zn was translocated to shoots more actively and distributed in all shoot tissues, being accumulated in leaf vascular bundles and epidermis. To conclude, these patterns of Ni and Zn distribution are aimed at metal sequestration in roots and leaf epidermis, thus keeping mesophyll from metal penetration and pigment degradation.     

Copper phytotoxicity affects root elongation and iron nutrition in durum wheat (Triticum turgidum durum L.)

This work investigated how copper (Cu) phytotoxicity affected iron (Fe) nutrition and root elongation in hydroponically grown durum wheat (Triticum turgidum durum L., cv Acalou) in order to establish the critical level of Cu concentration in roots above which significant Cu phytotoxicity occurs. This was assessed at two levels of Fe supply (2 and 100 μM). Severe symptoms of Cu phytotoxicity were observed at Cu²⁺ concentration above 1 μM, i.e. interveinal chlorosis symptoms and global root growth alteration. Total root Cu concentration of about 100, 150 and 250–300 mg kg^?1 corresponded to 10%, 25% and 50% reduction in root elongation, respectively. Copper and Fe concentrations as well as amounts of Cu and Fe accumulated in shoots varied inversely which suggested an antagonism between Cu and Fe leading to Fe deficiency. In addition, the root-induced release of complexing compounds increased significantly with increasing Cu concentration in nutrient solution and was positively correlated with Cu uptake without significant difference between the two Fe treatments (high and low Fe supply). This work suggests that total root Cu concentration might be a simple, sensitive indicator of Cu rhizotoxicity. It also indicated that Cu phytotoxicity which may have resulted in Fe deficiency and significant increase in root-induced release of complexing compounds (presumably phytosiderophores) was independent of the level of Fe supply provided that the threshold values of phytotoxicity were based on the free Cu-ion concentration.     

Copper tolerance in the cuprophyte Haumaniastrum katangense (S. Moore) P.A. Duvign. & Plancke

Cu tolerance and accumulation have been studied in Haumaniastrum katangense, a cuprophyte from Katanga (DR Congo), previously described as a copper hyperaccumulator. Nicotiana plumbaginifolia, a well-known non-tolerant and non-accumulator species, was used as a control. The germination rate of H. katangense was enhanced by copper and fungicide addition, suggesting that fungal pathogens, which restrain germination in normal conditions, are limiting. In hydroponic culture in the Hoagland medium, H. katangense did not grow well, in contrast to N. plumbaginifolia. Better growth was achieved by adding fungicide or higher copper concentrations. The maximal non-effective concentration (NEC) was 12 µM CuSO₄ for H. katangense grown in hydroponics, i.e. 24 times greater than Cu concentration in the Hoagland medium. By comparison, copper concentrations greater than 0.5 µM had a negative effect on the growth of N. plumbaginifolia. EC₅₀ (50% effective concentration) in hydroponics was 40 µM CuSO₄ for H. katangense and 6 µM CuSO₄ for N. plumbaginifolia. EC₁₀₀ (100% effective concentration) was 100 µM CuSO₄ for H. katangense and 15 µM CuSO₄ for N. plumbaginifolia. In soil, growth was also stimulated by Cu addition up to 300 mg kg^-1 CuSO₄. Surplus copper was also required for cultivating H. katangense in sterile conditions, suggesting that Cu excess may be necessary for needs other than pathogen defence. Cu accumulation in the shoot has been measured for N. plumbaginifolia and H. katangense at their respective NEC. Cu allocation in the two species showed a similar response to increasing Cu concentrations, i.e. root/shoot concentration ratio well above 1. In conclusion, H. katangense is highly tolerant to copper and has elevated copper requirement even in the absence of biotic interactions. Its accumulation pattern is typical of an excluder species.     

Toxicities of soluble Al, Cu, and La include ruptures to rhizodermal and root cortical cells of cowpea

Elevated levels of many metals are toxic to plant roots, but their modes of action are not well understood. We investigated the toxicities of aluminium (Al), copper (Cu), and lanthanum (La) in solution on the growth and external morphology of 3-d-old cowpea (Vigna unguiculata L.) roots for periods of up to 48 h. Root elongation rate decreased by 50% at ca. 30 μM Al, 0.3 μM Cu, or 2.0 μM La, accompanied by a decrease in the distance from the root tip to the proximal lateral root. Kinks developed in some roots 2.0 ± 0.4 mm from the root apex on exposure to Al or La (but not Cu). Light and scanning electron microscopy showed that soluble Al, Cu, or La caused similar transverse ruptures to develop > 1 mm from the root apex through the breaking and separation of the rhizodermis and outer cortex from inner-layers. The metals differed, however, in the range in concentration at which they had this effect; developing in solutions containing 54 to‑600 μM Al, but only from 0.85 to 1.8 μM Cu or 2.0 to 5.5 μM La. These findings suggest that Al, Cu, and La bind to the walls of cells, causing increased cell wall rigidity and eventual cell rupturing of the rhizodermis and outer cortex in the elongating zone. We propose that this is a major toxic effect of Al, and that Cu and La also have additional toxic effects.     

Physiological responses and antioxidant enzyme changes in <Emphasis Type="Italic">Sulla coronaria</Emphasis> inoculated by cadmium resistant bacteria

Plant growth promoting bacteria (PGPB) may help to reduce the toxicity of heavy metals on plants growing in polluted soils. In this work, Sulla coronaria inoculated with four Cd resistant bacteria (two Pseudomonas spp. and two Rhizobium sullae) were cultivated in hydroponic conditions treated by Cd; long time treatment 50 µM CdCl₂ for 30 days and short time treatment; 100 µM CdCl₂ for 7 days. Results showed that inoculation with Cd resistant PGPB enhanced plant biomass, thus shoot and root dry weights of control plants were enhanced by 148 and 35% respectively after 7 days. Co-inoculation of plants treated with 50 and 100 µM Cd increased plant biomasses as compared to Cd-treated and uninoculated plants. Cadmium treatment induced lipid peroxidation in plant tissues measured through MDA content in short 7 days 100 µM treatment. Antioxidant enzyme studies showed that inoculation of control plants enhanced APX, SOD and CAT activities after 30 days in shoots and SOD, APX, SOD, GPOX in roots. Application of 50 µM CdCl₂ stimulated all enzymes in shoots and decreased SOD and CAT activities in roots. Moreover, 100 µM of CdCl₂ increased SOD, APX, CAT and GPOX activities in shoots and increased significantly CAT activity in roots. Metal accumulation depended on Cd concentration, plant organ and time of treatment. Furthermore, the inoculation enhanced Cd uptake in roots by 20% in all treatments. The cultivation of this symbiosis in Cd contaminated soil or in heavy metal hydroponically treated medium, showed that inoculation improved plant biomass and increased Cd uptake especially in roots. Therefore, the present study established that co-inoculation of S. coronaria by a specific consortium of heavy metal resistant PGPB formed a symbiotic system useful for soil phytostabilization.     

Somatic embryogenesis and plant regeneration of blackberry using the thin cell layer technique

Limonium ‘Misty Blue’ is an interspecific hybrid of Limonium latifolium and L. bellidifolium and has a huge demand in floriculture business as both fresh and dry flowers with stunning purple-blue blooms. The propagation only through vegetative means restrict the popularization of this plant to the flower growers. We therefore optimized an efficient micropropagation protocol for direct organogenesis from root explants, as leaf is not conducible to respond in culture. 61.43% of root explants directly formed shoot buds on their surface after 4-weeks of culture in media containing ½ MS, 43.82 mM sucrose 2.22 µM BA and 1.07 µM NAA. The shoot buds failed to differentiate into healthy shoots unless the previous medium was replaced by full strength MS, and 87.64 mM sucrose along with 0.44 µM BA and 1.07 µM NAA. Encapsulations of juvenile shoots were carried out by 3% sodium alginate and 100 mM CaCl₂ which were again successfully stored at 4?°C for 30 days along with 56.79% of plant recovery in MS?+?0.44 µM BA?+?4.5 µM IBA?+?87.64 mM sucrose containing medium. 150 synthetic seed derived full grown plants were successfully acclimatized in green house, where a total of 101 plants survived after secondary hardening. The ISSR analysis revealed genetic homogeneity of synthetic seed derived hardened plants.     

Does Inoculation with Glomus mosseae Improve Salt Tolerance in Pepper Plants?

A pot experiment was conducted to determine the effects of Glomus mosseae inoculation on growth and some biochemical activities in roots and shoots of pepper (Capsicum annuum L. cv. Zhongjiao 105) plants subjected to four levels of NaCl [0 (control), 25 (low), 50 (medium), and 100 (high) mM] for 30 days, after 30 days of establishment under non-saline conditions. In mycorrhizal (M) plants, root colonization varied from 48 to 16 %. M plants had higher root and shoot dry weight and leaf area compared with non-mycorrhizal (NM) plants. Under salinity stress, M plants accumulated higher amounts of leaf photosynthetic pigments as well as soluble sugar, soluble protein, and total free amino acids in roots and shoots than those of NM plants. In contrast, the accumulation of proline was less intense in M plants than NM plants. Salt stress induced oxidative stress by increasing malondialdehyde (MDA) content; however, the extent of oxidative damage in M plants was less compared with NM plants due to G. mosseae-enhanced activity of superoxide dismutase (SOD) and peroxidase (POD). We concluded that inoculation with G. mosseae improved growth performance and enhanced salt tolerance of pepper plants via improving photosynthetic pigments and the accumulation of organic solutes (except proline), reducing oxidative stress, and enhancing antioxidant activities of the SOD-POD system.     

<Emphasis Type="Italic">In vitro</Emphasis> regeneration of <Emphasis Type="Italic">Melothria maderaspatana</Emphasis> via indirect organogenesis

An effective protocol was developed for in vitro regeneration of the Melothria maderaspatana via indirect organogenesis in liquid and solid culture systems. Organogenesis was achieved from liquid culture calluses derived from leaf and petiole explants of mature plants. Organogenic calluses (98.2?±?0.36 and 94.8?±?0.71%) were induced from both leaf and petiole explants on Murashige and Skoog (MS) liquid medium containing 6.0 µM 2,4-dichlorophenoxyacetic acid (2,4-D) and 0.5 µM thidiazuron (TDZ); and 6.0 µM 2,4-D and 1.0 µM benzyladenine (BA) combinations, respectively. Adventitious shoot regeneration (68.2?±?0.06 shoots per explant) was achieved on MS medium supplemented with 2.0 µM BA, 4.0 µM TDZ, 10% v/v coconut water and 0.06 mM glutamine from leaf-derived calluses. Petiole-derived calluses produced adventitious shoots (45.4?±?0.09 shoots per explant) on MS medium fortified with 2.0 µM BA, 4.0 µM TDZ, 10% v/v coconut water, and 0.08 mM glutamine. Elongation of shoots occurred in MS medium with 2.0 µM gibberellic acid (GA₃). Regenerated shoots (2–3 cm in length) rooted (74.2?±?0.38%) and hardened (85?±?1.24%) when they were transferred to 1/2-MS medium supplemented with 3.0 µM indole-3-butyric acid (IBA) followed by garden soil, vermiculate, and sand (2:1:1 ratio) mixture. The elongated shoots (4–5 cm in length) were exposed simultaneously for rooting as well as hardening (100%) in moistened [(1/8-MS basal salt solution with 5 µM IBA and 100 mg l^?1 Bavistin® (BVN)] garden soil, vermiculate, and sand (2:1:1 ratio) mixture. Subsequently, the plants were successfully established in the field. The survival percentage differed with seasonal variations.     

Plant growth regulator induced phytochemical and antioxidant variations in micropropagated and acclimatized Eucomis autumnalis subspecies autumnalis (Asparagaceae)

Eucomis species is a valuable plant with both medicinal and horticultural potential. The current study evaluated the role of plant growth regulator (PGR) on growth, phytochemicals, and antioxidant activity in Eucomis autumnalis subspecies autumnalis. Five cytokinins including topolins and benzyladenine (BA) at 2 µM in combination with varying (0–15 µM) concentrations of naphthalene acetic acid (NAA) were tested. In vitro regenerants were acclimatized in the greenhouse for 4 months. Highest number of shoots (9 shoots/explant) was observed with 15 µM NAA alone or when combined with BA. Acclimatized plants derived from the 15 µM NAA treatment had the highest number of roots, largest leaf area and biggest bulbs. While applied PGRs increased the iridoids and condensed tannins in the in vitro regenerants, total phenolics and flavonoids were higher in the PGR-free treatment. Among the in vitro regenerants, 5 µM NAA and 2 µM BA treatments produced the best antioxidant activity in the DPPH (55 %) and beta-carotene (88 %) test systems, respectively. A remarkable carry-over effect of the PGR was conspicuous in the phytochemical levels and antioxidant activity of the 4-month-old plants. In addition to the optimized micropropagation protocols, the current findings present a promising potential for manipulating the type and concentration of applied PGRs to improve phytochemical production and hence medicinal value in E. autumnalis subspecies autumnalis.     

Effect of anti-ethylene compounds on isoenzyme patterns and genome stability during long term culture of <Emphasis Type="Italic">Moringa oleifera</Emphasis>

Multiplication of Moringa oleifera shoots on MS medium supplemented with 2.5 µM BAP for 3 weeks resulted in shoot vitrification which led to chlorosis, retardation of shoot formation, reduction in shoot length, necrosis of shoot tips and formation of friable calli on the base of cultured explants. Vitrification symptoms decreased when MS medium containing 2.5 µM BAP in combination with 10 µM AgNO₃, 50 µM salicylic acid (SA) or 200 µM CoCl₂ was used. Studying isoenzyme patterns of SOD, POX, CAT, GOT and EST indicated that moringa shoots multiplied without obvious variation in isoenzyme patterns up to 7 subcultures. Moringa shoots subjected to 14 subcultures and anti-ethylene compounds showed variation in isoenzyme patterns and were associated with the disappearance of vitrification which facilitated root formation and acclimatization. Under long term cultures, RAPD, ISSR and SSR indicated that AgNO₃ was the optimal anti-ethylene substance for avoidance of vitrification in moringa but it resulted in high somaclonal variation. Application of SA decreased vitrification as well as somaclonal variation compared to CoCl₂ under long term culture. Consequently, SA was recommended for moringa clonal multiplication.     

In vitro induction of tetraploid <Emphasis Type="Italic">Ziziphus jujuba</Emphasis> Mill. var. <Emphasis Type="Italic">spinosa</Emphasis> plants from leaf explants

The genetic manipulation of Capsicum has been unsuccessful, and a large bottleneck to transferring the desired genes is due to the difficulty in regenerating whole plants through tissue culture because of its highly recalcitrant and high genotype specificity. This study aimed to investigate and establish rapid shoot regeneration from the proximal ends of the leaves of Capsicum frutescens KT-OC and BOX-RUB varieties. A maximum of 8–10 shoot buds were obtained from the margins of the proximal portion of a cotyledonary leaf explant of C. frutescens variety KT-OC on medium I containing 44.44 µM 6-benzylaminopurine (BA), 5.71 µM indole-3-acetic acid (IAA), 10 µM silver nitrate (AgNO₃) and 1.98 mg L^?1 2-(N-morpholine) ethane sulphonic acid within 4 weeks of incubation, of which 60% of explants responded in terms of shoot buds. Petiole explants (40%) cultured on the same medium produced 2–4 shoots per explant from the distal portion. The cut portions of the cotyledonary leaf proximal portions responded well to shoot bud formation in the presence of 22.20 µM BA and 14.68 µM phenyl acetic acid (PAA), wherein 100% of explants responded in terms of shoot bud formation, with an average of 10?±?1.7 and 8?±?1.9 shoot buds per explant in KT-OC and BOX-RUB varieties, respectively. The differentiated shoots grew well and proliferated in the presence of 14.68 µM PAA?+?22.20 µM BA and 10 µM AgNO₃. Shoot elongation was obtained in presence of 1.44 µM gibberellic acid (GA₃) and 10 µM AgNO₃. These shoots were rooted on plant growth regulator-free half-strength MS medium and upon hardening; field survival rate was 70%. This reproducible regeneration method for C. frutescens, especially the Indian high pungent variety, from proximal portion of cotyledonary leaf and petiole explants, can be used for biotechnological improvement.     

Toxicity of Cd to signal grass (Brachiaria decumbens Stapf.) and Rhodes grass (Chloris gayana Kunth.)

Given that Cd accumulates within plant tissues to levels that are toxic to animals, it is necessary to understand the role of plants in highly Cd-contaminated systems and their subsequent impact on the health of animals. A solution culture experiment was conducted to elucidate the effects of increasing Cd²⁺ activity ({Cd²⁺}) on growth of Rhodes grass (Chloris gayana Kunth.) and signal grass (Brachiaria decumbens Stapf.). The shoot and root fresh mass of both Rhodes grass and signal grass was reduced by 50% at ca. 0.5 µM {Cd²⁺}. Elevated {Cd²⁺} resulted in a significant decrease in the tissue Mn concentration for both the shoots and roots, and caused a chlorosis of the veins in the shoots. Root hair growth was prolific even at high {Cd²⁺}, thus root hair growth appeared to be less sensitive to elevated Cd than was root growth per se. The critical shoot tissue concentrations (50% reduction in growth), 230 µg g^?1 for Rhodes grass and 80 µg g^?1 for signal grass, exceeded the maximum level of Cd tolerated in the diet of animals (ca. 5 µg g^?1). When assessing the risk associated with the revegetation of Cd-contaminated sites with Rhodes grass or signal grass, careful consideration must be given, therefore, to the transfer of toxic concentrations of Cd to grazing animals and through the wider food chain.     

TDZ induces shoot regeneration in various Kalanchoë blossfeldiana Poelln. cultivars in the absence of auxin

In order to optimize shoot regeneration in Kalancho? blossfeldiana, leaf and internode explants of seven cultivars including one inter-specific were studied. The effects of various combinations of α-naphthalene acetic acid (NAA) (0, 0.57 M) and thidiazuron (TDZ) (0, 0.45, 4.5, 22.5, 67.5 μM) on MS medium were examined. In all cultivars shoot regeneration frequency and number of shoots per explant were enhanced by increasing TDZ concentration. Supplementing the media with NAA did not improve shoot regeneration. Maximum regeneration frequency and optimum concentration of TDZ for shoot regeneration depended significantly on the cultivar. Internode explants, but not leaf explants, of some cultivars, were able to produce adventitious shoots without treatment with growth regulator.     

Tissue partitioning of cadmium in transgenic tobacco seedlings and field grown plants expressing the mouse metallothionein I gene

Since agricultural crops contribute >70% of human cadmium (Cd) intake, modification of crops to reduce accumulation of this pollutant metal during plant growth is desirable. Here we describe Cd accumulation characteristics of seedlings and field grown tobacco plants expressing the Cd-chelating protein, mouse metallothionein I. The objective of the transformation is to entrap Cd in roots as Cd-metallothionein and thereby reduce its accumulation in the shoot. Transformed and control seedlings were exposed for 15 days in liquid culture at a field soil-solution-like Cd concentration of 0.02 μm. Transformed seedlings ofNicotiana tabacum cultivar KY 14 contained about 24% lower Cd concentration in shoots and about 5% higher Cd concentration in roots than control seedlings. Dry weights of transformed and control tissues did not differ significantly. In the field in 1990, mature transformedN. tabacum cv. KY 14 plants exposed only to endogenous soil Cd contained about 14% lower leaf lamina Cd concentration than did controls. Differences were significant at thep≤0.1 level in 13 of 16 leaf positions. Leaf dry weight did not differ significantly but transformed field plants had 12% fewer leaves and were 9% shorter than the controls. Copper (Cu) concentration was significantly higher (ca10%) in the bottom nine leaf positions of transformed plants suggesting that reduced leaf number and plant height may be due to Cu deficiency or toxicity. Alternatively, somaclonal variation or gene position effects may be involved. No differences were found in zinc levels. WithN. tabacum cv. Petit Havana, transformed seedlings contained no less Cd in shoots but 48% higher Cd concentration in roots. However, dry weights of shoots and roots of transformed seedlings were 25% and 26%, respectively, greater than in controls. In the field, transformed and control plants of this cultivar showed little significant differences in leaf Cd content, plant height or leaf number. Although comparison of additional metallothionein-expressing tobaccos and other plants is needed, results obtained with cultivar KY 14 support the hypothesis that sequestration of Cd in roots as Cd-metallothionein may have potential for reducing Cd content of above root tissues of certain plants.     

Zinc ameliorates copper-induced oxidative stress in developing rice (Oryza sativa L.) seedlings

Rice (Oryza sativa L.) seedlings were treated with different concentrations of copper (Cu) either in presence or absence of zinc (Zn), and different events were investigated to evaluate the ameliorative effect of Zn on Cu stress. In presence of high Cu concentration, growth of both root and shoots were considerably reduced. Decline in elongation and fresh mass was observed in root and shoot. Zn alone did not show any considerable difference as compared to control, but when supplemented along with high concentration Cu, it prompted the growth of both root and shoot. After 7 days, root growth was 9.36 and 9.59 cm, respectively, at 200 and 500 μM of Cu alone as compared to 10.59 and 12.26 cm at similar Cu concentrations, respectively, in presence of Zn. Cu accumulation was considerably high after 7 days of treatment. In absence of Zn, significant accumulation of Cu was observed. Zn supplementation ameliorated the toxic impact of Cu and minimized its accumulation. Cu treatment for 1 and 7 days resulted in a dose-dependent increase in hydrogen peroxide (H₂O₂). When Cu was added in presence of Zn, the H₂O₂ production in root and shoot was reduced significantly. The increase in H₂O₂ production under Cu stress was accompanied by augmentation of lipid peroxidation. In absence of Zn, Cu alone enhanced the malondialdehyde (MDA) production in both root and shoot after 1 and 7 days of treatment. The MDA content drastically reduced in root and shoot as when Zn was added during Cu treatment. The activities of antioxidant enzymes like superoxide dismutase (SOD), catalase (CAT), and guaiacol peroxidase (GPX) were elevated under Cu stress both in root and shoot. Addition of Zn further stimulated the activities of these enzymes. Both ascorbate (AsA) and glutathione (GSH) contents were high under Cu stress either in presence or absence of Zn. The results suggests that Zn supplementation improves plant survival capacity under high Cu stress by modulating oxidative stress through stimulation of antioxidant mechanisms and restricts the accumulation of toxic concentrations of Cu.     

Tolerance and potential for bioaccumulation of <Emphasis Type="Italic">Alternanthera tenella</Emphasis> Colla to cadmium under in vitro conditions

High concentrations of cadmium (Cd) in the environment can threaten the local biota and one of its main sources is anthropic activities such as zinc (Zn) mining. Some plant species are able to tolerate high Cd concentrations, using anatomical and physiological strategies to avoid the absorption or accumulation of this element in their biomass. The in vitro assessment of these strategies is an efficient way to control variables external to the experiment. We aimed to investigate the anatomical and physiological changes in Alternanthera tenella exposed to Cd and its potential for accumulation in controlled microenvironmental conditions. We evaluated changes in the leaf and root anatomy, antioxidant system, and biomass of A. tenella grown in a culture medium containing increasing Cd concentrations (0, 50, 100, and 150 μM), in the presence of 1500 μM Zn. Alternanthera tenella was able to accumulate Cd and Zn and these elements competed for absorption by the species. Increase in Cd in the medium led to a progressive thickening of the root tissues, which was also observed on the leaves, albeit only at concentrations below 100 μM Cd. The concentration of 150 μM Cd was toxic to the leaf tissue and stimulated the formation of hydrogen peroxide, interfering with the antioxidant system and reducing plant biomass and the chlorophyll levels. Therefore, in vitro cultivated A. tenella can accumulate Cd and tolerate up to 100 μM Cd by modifying its anatomy and physiology in order to cope with Cd stress.     

Characterization and Identification of Hepatic mRNA Related to Copper Metabolism and Homeostasis in Cattle

Copper is an essential trace mineral required for growth and development. Copper homeostasis within the cell is mediated by the expression of the Cu transporter protein (CTR1), ATPase7A (ATP7A), ATPase7B (ATP7B), Cox17, and Cu chaperone for Cu–Zn superoxide dismutase (CCS) which helps to regulate Cu uptake, export, and intracellular compartmentalization in non-ruminants. Copper also serves as a cofactor of antioxidant, superoxide dismutase1 (SOD1). Liver tissue from eighteen Holstein bull calves (average BW 201?±?58.5 kg, 7.3?±?1.9 months) from a previous experiment were utilized to characterize and identify hepatic mRNA related to Cu metabolism and homeostasis in cattle. Hepatic Cu concentration was determined via flame atomic absorption, and total RNA was extracted using TRI reagent and purified using RNeasy. Hepatic Cu concentrations ranged from 86 to 801 mg of Cu/kg DM. Real-time polymerase chain reaction analysis revealed that CTR1, ATP7A, and ATP7B mRNA expressions were negatively correlated with hepatic Cu concentration, while CCS (P?=?0.0887) and SOD1 had a tendency (P?=?0.0733) to be negatively correlated to hepatic Cu concentration. These data indicate that higher than normal hepatic Cu concentration downregulates gene expression of CTR1, ATP7A, ATP7B, and Cox17, which are involved in bovine liver copper homeostasis.     

Copper-Induced Modulation of Biomass Growth,Physiological Parameters,Bioactive Centellosides,and Expression of Biosynthetic Pathway Genes in an Important Medicinal Herb, <Emphasis Type="Italic">Centella asiatica</Emphasis>

Hydroponically established Centella asiatica plants were studied under different concentrations of copper (0, 0.32,1.6, and 3.2 µM) for their morphological, physiological, and biochemical characteristics. The plants grown in the medium without any exogenous supply of copper showed improved biomass accumulation with maximum fresh weight (FW?=?24.7 g plant^?1) and dry weight (DW?=?2.35 g plant^?1). The total chlorophyll content, leaf number, and the leaf area were higher (0.27 mg g^?1 DW, 20 and 100.54 cm²) in the plants grown under copper-deficient conditions. The toxic effects of increased levels of copper were evident by significant inhibition in growth and other morphological parameters. Copper treatment showed an increase in malondialdehyde (MDA) content and SOD activity. Bioactive phytochemical profiling using HPLC analysis revealed that higher levels of copper (1.6 and 3.2 µM) inhibit the accumulation of total centelloside content. The differential accumulation of centelloside content was further validated by the consistent pattern of expression of key pathway genes related to centelloside production in copper-treated C. asiatica plants. The study unveiled some of the important facts associated with the mechanism of copper tolerance in this important medicinal herb C. asiatica. The findings of the present study can be further used to provide better production of high value, in demand centellosides.     

Efficient plant regeneration from shoot apex explants of maize (Zea mays) and analysis of genetic fidelity of regenerated plants by ISSR markers

An efficacious regeneration system was developed from shoot apex explants of Zea mays using a two-step culture procedure. Seventeen Indian genotypes were assessed for their regeneration potential. The maximum response of shoot induction was obtained from explants cultured on Murashige and Skoog medium supplemented with 4.5 µM thidiazuron and 26.7 µM glycine. Maximum mean number of shoots (17.2) was observed in genotype COH (m)-5 while NPK was the least responsive (6.7). Shoot clumps transferred from shoot induction medium to multiplication (second) medium amended with 1.1 µM thidiazuron and 0.88 µM N ₆ -benzylaminopurine showed increased number of shoots in COH (m)-5 (36.1 shoots); NPK was the least responsive with an average of 9.5 shoots. The best response in root induction, with a larger number of roots (10.5) and longer roots (6.6 cm), was observed in Murashige and Skoog medium supplemented with 7.3 µM indole-3-butyric acid and 7.9 µM phloroglucinol. Analysis of variance indicated that plant regeneration response varied greatly among the genotypes. In vitro raised plants were successfully transferred to the field after hardening, with a 99 % survival rate. Inter simple sequence repeats analysis revealed that the similarity matrix pair-wise value was 1, the Mantel test value was p 1.0; Analysis of molecular variance genetic variances were 93 % within the population and 7 % between populations; Principal component jolliffe cut off was 0.15, Principal component and Principle coordinate analysis % variance was 13.19. These values were congruent for both the mother and the in vitro-raised plants, confirming genetic integrity.