Localisation of nickel and mineral nutrients Ca,K, Fe,Mg by Scanning Electron Microscopy microanalysis in tissues of the nickel-hyperaccumulator Alyssum bertolonii Desv. and the non-accumulator Alyssum montanum L.

Plants of the nickel-hyperaccumulator Alyssum bertolonii Desv. and of the non-accumulator A. montanum L. growing on a serpentine site in Tuscany, Italy, and plants of A. montanum from a nearby non-serpentine site were analysed for metal concentration and localisation. The leaves of A. bertolonii contained 160 times more nickel than those of A. montanum from the same site, thus demonstrating its hyperaccumulation capacity towards this metal. On the other hand, both species showed an inversion of the Ca/Mg ratio in their organs relative to the soil. Nickel localisation in plant tissues was examined by Scanning Electron Microanalysis (SEM/EDX). In A. bertolonii, a specific pattern of nickel distribution was detected, with the highest concentrations present in parenchyma and sclerenchyma cells for the roots; in the shoots, the highest amounts of nickel were found in the stem epidermis, the leaf epidermal surface, and the leaf trichome base. This particular nickel tissue distribution pattern was not found in the non-accumulator A. montanum growing on serpentine soil. Other mineral nutrients, namely Mg, Ca, K, Fe, instead, had a similar distribution in the two species. The A. montanum plants from the non-serpentine site had very low nickel levels in their tissues, and these were of the same magnitude as those found in A. bertolonii plants grown in a greenhouse on commercial horticultural soil with low nickel concentration. In A. bertolonii plants, the tissue-specific allocation patterns appeared to depend on the degree of nickel hyperaccumulation, which is, in turn, directly linked to the soil characteristics.     

Ynt is the primary nickel import system used by Proteus mirabilis and specifically contributes to fitness by supplying nickel for urease activity

Proteus mirabilis is a Gram-negative uropathogen and frequent cause of catheter-associated urinary tract infection (CAUTI). One important virulence factor is its urease enzyme, which requires nickel to be catalytically active. It is, therefore, hypothesized that nickel import is critical for P. mirabilis urease activity and pathogenesis during infection. P. mirabilis strain HI4320 encodes two putative nickel import systems, designated Nik and Ynt. By disrupting the substrate-binding proteins from each import system (nikA and yntA), we show that Ynt is the primary nickel importer, while Nik only compensates for loss of Ynt at high nickel concentrations. We further demonstrate that these are the only binding proteins capable of importing nickel for incorporation into the urease enzyme. Loss of either nickel-binding protein results in a significant fitness defect in a murine model of CAUTI, but YntA is more crucial as the yntA mutant was significantly outcompeted by the nikA mutant. Furthermore, despite the importance of nickel transport for hydrogenase activity, the sole contribution of yntA and nikA to virulence is due to their role in urease activity, as neither mutant exhibited a fitness defect when disrupted in a urease-negative background.     

Nickel localization in tissues of different hyperaccumulator species of Euphorbiaceae from ultramafic areas of Cuba

Pantropical species of the genera Phyllanthus and Euphorbia and the Cuban endemic genus Leucocroton from the Euphorbiaceae family, were selected for nickel localization microanalysis. Scanning Electron Microscopy coupled with Energy Dispersive X-ray Microanalysis (SEM-EDX) was used for qualitative detection of nickel in the selected Ni-hyperaccumulator species: Euphorbia helenae, Leucocroton linearifolius, L. flavicans Phyllanthus orbicularis, P. discolor and P. xpallidus, all collected from Cuban ultramafic soils. The leaves and stems from the Euphorbiaceae species analyzed were the organs with higher nickel accumulation. Elemental mapping of leaves and stem tissues from these species have been compared. The highest Ni concentrations were found in the laticifer tubes of stems and the epidermis tissues of leaves in all the analyzed species, suggesting a general pattern of the Euphorbiaceae family for nickel accumulation. The high nickel concentrations and its rather homogeneous distribution found in leaves of these Ni-hyperaccumulating plants suggest a possible role in protection mechanisms against environmental stress, such as UV irradiation.     

Biodiversity of Hydrogenases in Frankia

Eighteen Frankia strains originally isolated from nine different host plants were used to study the biodiversity of hydrogenase in Frankia. In the physiological analysis, the activities of uptake hydrogenase and bidirectional hydrogenase were performed by monitoring the oxidation of hydrogen after supplying the cells with 1% hydrogen and the evolution of hydrogen using methyl viologen as an electron donor, respectively. These analyses were supported with a study of the immunological relationship between Frankia hydrogenase and other different known hydrogenases from other microorganisms. Uptake hydrogenase activity was recorded from all the Frankia strains investigated. A methyl-viologen-mediated hydrogen evolution was recorded from only four Frankia strains irrespective of the source of Frankia. From the immunological and physiological studies, we here report that there are at least three types of hydrogenases in Frankia: Ni-Fe uptake hydrogenase, hydrogen-evolving hydrogenase, and [Fe]-hydrogenase. An immunogold localization study, by cryosection technique, of the effect of nickel on the intercellular distribution of hydrogenase proteins in Frankia indicated that nickel affects the transfer of hydrogenase proteins into the membrane.     

The influence of Nickel on population dynamics and on some demographic parameters of Daphnia magna

Daphnia magna was cultured with two different techniques at sublethal nickel concentrations from 0 up to 200 µg Ni l^-1. The results of the experiments run with cohorts of single individuals in static cultures were compared with the results obtained with populations held in flow through cultures. The most sensitive demographic parameters in cohorts were life span, the total number of progeny and the achievable maximum body length. The investigated nickel concentrations lowered the mean animal density in populations. Size class distribution was affected and the oscillation of the percentage of ovigorous adults increased with increasing nickel concentrations. In 200 µg Ni l^-1 one population extincted. Individuals in static cultures exposed to the same nickel concentration produced at least two neonates. Both employed techniques are discussed.     

Binding of Ni2+ to a histidine- and glutamine-rich protein, Hpn-like

Hpn-like (Hpnl) protein, encoded by the hpnl gene in Helicobacter pylori and featuring a histidine-rich and two glutamine-rich motifs, can render nickel tolerance to H. pylori when the external nickel level reaches toxic limits. We found that the recombinant Hpnl exists as an oligomer in the native state and binds to two molar equivalents of nickel ions per monomer with a dissociation constant of 3.8 μM. Nickel could be released from Hpnl either at acidic pH (pH_1/2 4.6) or in the presence of chelate ligands, such as EDTA (t _1/2 = 220, 355, and 716 min at pH 6.0, 7.0, and 7.5, respectively). Our combined spectroscopic data show that nickel ion coordinates to a nitrogen of a histidine residue possibly with a coordination number of four (square-planar geometry) or five. The growth of Escherichia coli cells with or without the hpnl gene implied a protective role of Hpnl under higher concentrations of external nickel ions. Hpnl may serve a role in binding/storage or detoxification of excess nickel ions. Electronic supplementary material  The online version of this article (doi:) contains supplementary material, which is available to authorized users.     

Response to cytosolic nickel of Slow Vacuolar channels in the hyperaccumulator plant <Emphasis Type="Italic">Alyssum bertolonii</Emphasis>

We applied the patch-clamp technique to investigate the transport properties of the Slow Vacuolar (SV) channel identified in leaf vacuoles of Alyssum bertolonii Desv., a nickel hyperaccumulator plant growing in serpentine soil of the northern Apennines (Italy). SV currents recorded in vacuoles from adult plants collected in their natural habitat showed high sensitivity towards cytosolic nickel. Dose-response analyses indicated half-maximal current inhibition at submicromolar concentrations, i.e. up to three orders of magnitude lower than previously reported values from other plant species. The voltage-dependent increase of residual currents at saturating nickel concentrations could be interpreted as relief of channel block by nickel permeation at high positive membrane potentials. Including young plants of A. bertolonii into the study, we found that SV channels from these plants did not display elevated nickel sensitivity. This difference may be related to age-dependent changes in nickel hyperaccumulation of A. bertolonii leaf cells.     

The Effect of Recombinant Heavy Metal-Resistant Endophytic Bacteria on Heavy Metal Uptake by Their Host Plant

The ncc-nre nickel resistance system of Ralstonia metallidurans 31A was efficiently expressed in Burkholderia cepacia L.S.2.4 and Herbaspirillum seropedicae LMG2284. The heterologous expression of ncc-nre encoded nickel resistance was accompanied by nickel removal from the culture medium. B. cepacia L.S.2.4:: ncc-nre and H. seropedicae LMG2284::ncc-nre were able to remove 35 and 15% nickel, respectively. The capacity to remove nickel through sequestration or bio-precipitation processes and consequently lowering the free nickel concentration could offer interesting benefits for these endophytic bacteria and their host plants. Once inoculated in their host plant, they could possibly alter the nickel speciation and therefore decrease the free ions and thus toxic concentration for the plant metabolism. Lupinus luteus L, when grown on a nickel enriched substrate and inoculated with B. cepacia L.S.2.4::ncc-nre, showed a significant increase (30%) of nickel concentration in the roots, whereas the nickel concentration in the shoots remained comparable with that of the control plants. The inoculation of Lolium perenne (cv Atlas) with the nickel resistance derivative of H. seropedicae LMG2284::ncc-nre resulted in a significant decrease of the nickel concentration in the roots (11%) as well as in the shoots (14%). As this phenomenon was also observed in the Lolium perenne plants inoculated with the wild-type strain LMG2284, the nickel resistance characteristics probably are not responsible for the altered nickel uptake observed.     

Serpentine and non-serpentine Silene dioica plants do not differ in nickel tolerance

Serpentine soils are hostile to plant life. They are dry, contain high concentrations of nickel and have an unfavorable calcium/magnesium ratio. The dioecious plant Silene dioica (L.) Clairv. (Caryophyllaceae) is the most common herb on serpentine soils in the Swedish mountains. It also commonly grows on non-serpentine soils in the subalpine and coastal area. I have compared the germination frequency, plant establishment and growth of serpentine and subalpine non-serpentine populations in serpentine soil under greenhouse conditions. Further more I have studied the specific effect of nickel on root and shoot growth of serpentine and non-serpentine plants from the subalpine and coastal area in solutions with different concentrations of nickel. Plants from serpentine and non-serpentine populations grew well and in a similar fashion in serpentine soil. Moreover, S. dioica plants, irrespective of original habitat, tolerated enhanced concentrations of nickel when grown in solutions. An analysis of metal content in serpentine plants from natural populations shows that S. dioica has a higher nickel concentration in the roots than in the shoots. The growth studies show that S. dioica is constitutively adapted to serpentine, and that all populations have the genetic and ecological tolerance to grow on serpentine.     

Allergic contact dermatitis to nickel is associated with a Taq I HLA-DQA allelic restriction fragment

RFLP analysis of the HLA class II genes DRA, DQA, and DQB was performed in 33 patients with allergic contact eczema to nickel. A significant association with a Taq I HLA-DQA allelic restriction fragment was found. Twenty-two of 33 patients compared to 31 of 100 healthy controls had a 4.5-kb DQA fragment (corrected P value <0.05, relative risk 4.5, and etiologic fraction 0.52). In order to study whether the magnitude of the response in a lymphocyte proliferative assay to nickel sulfate was controlled by HLA class II genes or not, the patients were divided into low, intermediate, and high responders. No significant differences were found in the distribution of Taq I HLA class II allelic patterns between the groups.     

Tunable Free‐Standing Ultrathin Porous Nickel Film for High Performance Flexible Nickel–Metal Hydride Batteries

The nickel matrix has a significant impact on the structure and performance of a nickel–metal hydride (NiMH) battery. However, few studies have focused on the nickel matrix thus far due to the difficulty of fabricating controllable porous nickel materials. In addition, conventional nickel matrices show poor flexibility, making it difficult to fabricate flexible NiMH batteries. To achieve a high performance flexible NiMH battery, the fabrication of a thin, free‐standing, and flexible nickel matrix with an optimized pore structure is a key prerequisite. Here, a novel flexible porous nickel matrix with a controllable pore size, density, and distribution of pore position is developed by nickel electrodeposition on templates that are produced by silkscreen printing different insulating ink microarrays on stainless steel sheets. Benefitting from the excellent structure of the porous nickel matrix, flexible NiMH batteries are fabricated, which show excellent flexibility and very high energy densities of 151.8 W h kg^?1 and 508.5 W h L^?1 as well as high energy efficiencies of 87.9–98.5%. These batteries outperform conventional NiMH batteries and many other commercial batteries, holding great promise for their future practical application. The present strategy provides a new route to promote the development of nickel‐based alkaline rechargeable batteries.     

Nickel absorption and distribution from rat small intestine In situ

The aim of this work was to study the absorption of nickel chloride in rats by means of the intestinal perfusion in situ technique at nickel concentrations of 1, 5, 10, 25, and 100 mg/L. Active transport and facilitated diffusion seem to play an important role in the intestinal absorption of nickel at concentrations≤10 mg/L. At higher concentrations, the absorption rate would be limited by saturation of the carriers. The distribution of the absorbed nickel was studied by intestinal perfusion of a 10-mg Ni/L solution for 30 or 60 min. Both in concentration and amount, the jejunum showed the higher values of absorbed nickel, followed by the kidneys and liver. When all of the collected organs (brain, heart, liver, lungs, spleen, kidneys, and testicles) and blood, but not the small intestine, are analyzed following a 60-min perfusion, it was found that 1% of the initial concentration had passed through the intestinal barrier.     

Nickel enhances telomeric silencing in Saccharomyces cerevisiae

Certain nickel compounds including crystalline nickel sulfide (NiS) and subsulfide (Ni₃S₂) are potent human and animal carcinogens. In Chinese hamster embryo cells, an X-linked senescence gene was inactivated following nickel-induced DNA methylation. Nickel also induced the inactivation of the gpt reporter gene by chromatin condensation and a DNA methylation process in a transgenic gpt+ Chinese hamster cell line (G12), which is located near a heterochromatic region. To determine if nickel can cause gene silencing independently of DNA methylation, based only on the induction of changes in chromatin structure, we measured its effect on gene silencing in Saccharomyces cerevisiae. Growth of yeast in the presence of nickel chloride repressed a telomeric marker gene (URA3) and resulted in a stable epigenetic switch. This phenomenon was dependent on the number of cell doubling prior to selection and also on the distance of the marker gene from the end of the chromosome. The level of TPE (telomeric position effect) increased linearly with elevations of nickel concentration. Addition of magnesium inhibited this effect, but magnesium did not silence the reporter gene by itself. The level of silencing was also assessed following treatment with other transition metals: cobalt, copper and cadmium. In the sublethal range, cobalt induced similar effects as nickel, while copper and cadmium did not change the basal level of gene expression. Silencing by copper and cadmium were evident only at concentrations of those metals where the viability was very low.     

Comparison of two serpentine species with different nickel tolerance strategies

The effects of nickel were studied in two serpentine species with different metal tolerance strategies:Silene italica L., which limits nickel uptake and translocation, andAlyssum bertolonii Desv., a serpentine endemic, which accumulates nickel mostly in the leaves. InS. italica, nickel 7.5 μM inhibited root growth and depressed mitotic activity in root tips. Peroxidase activity and phenol concentration both in roots and shoots were increased; under the same conditions nickel did not produce any relevant effect onA. bertolonii. InS. italica an adequate calcium concentration (25 mM) was able to reverse the effects of nickel on root growth and metabolism. InA. bertolonii the same calcium concentration reduced root growth, confirming this species adaptation also to low calcium concentrations, typical of serpentines.     

Nickel deficiency alters nickel flux in rat everted intestinal sacs

This study was conducted to determine nickel absorption in nickel-deficient rats. Jejunal segments obtained from dietary nickeldepleted (13 μg nickel/kg diet) and nickel-control (1 mg nickel/kg diet) adult rats from the first generation, and suckling pups from the second offspring were used. The nickel transfer across the intestinal epithelium and nickel uptake into the intestine were measured by use of everted jejunal sacs using a wide range of nickel concentrations administered on the luminal side (1.1 x 10^-8 M til 1.0 x 10^-4 M). Both the intestinal nickel transfer and nickel uptake were influenced by the dietary nickel supply in rat offspring, but not in the adult rats from the first generation. However, in nickel-deficient offspring, the nickel transfer across the small intestine was higher than in nickelcontrol offspring. This difference was greater using low intraluminal nickel concentrations than high nickel concentrations, and was significant at 1.1 x 10^-8 M, 6.1 x 10^-8 M, 5.1 x 10^-7 M, 1.0 x 10^-6 M, and 5.0 x 10^-6 M. Also, nickel uptake into the intestine was somewhat greater in nickel-deficient rat pups than in nickel-control pups, and significant using 1.1 x 10^-7 M and 1.0 x 10^-6 M nickel. A definite saturation type kinetic for the intestinal nickel absorption in relation to the intraluminal nickel concentration could not be observed.     

Bioleaching of nickel from equilibrium fluid catalytic cracking catalysts

Summary This study investigates the possibility of reusing metal-contaminated equilibrium fluid catalytic cracking (FCC) catalyst after bioleaching. Leaching with Aspergillus niger culture was found to be more effective in the mobilization of nickel from the catalyst particles compared to chemical leaching with citric acid. Bioleaching achieved 32% nickel removal whereas chemical leaching achieved only 21% nickel removal from catalyst particles. The enhanced nickel removal from the catalysts in the presence of A. niger culture was attributed to the biosorption ability of the fungal mycelium and to the higher local concentration of citric acid on the catalyst surface. It was found that 9% of solubilized nickel in the liquid medium was biosorbed to fungal biomass. After nickel leaching with A. niger culture, the hydrogen-to-methane molar ratio and coke yield, which are the measures of dehydrogenation reactions catalysed by nickel during cracking reactions, decreased significantly.     

Effects of nickel on Frankia and its symbiosis with Alnus glutinosa (L.) Gaertn

The tolerance of nickel by Frankia in culture and in symbiosis with Alnus was determined. Yield of three Frankia strains was not affected significantly by 2.25 mM nickel when cultured in propionate medium containing hydolysed casein as nitrogen source. Yield of two strains in medium without combined nitrogen, and thus reliant on fixed nitrogen, was stimulated markedly by the same nickel concentration. Utilisation of nickel for synthesis of uptake hydrogenases is presumed to be the cause of enhanced nitrogenase activity.Although growth was reduced, treatment of 2-month-old seedlings with 0.025 mM nickel for 4 weeks did not affect nodulation significantly while nitrogenase activity was doubled. Nodulation and nitrogenase activity of seedlings receiving 0.075 mM nickel were inhibited markedly, while 0.5 mM nickel was lethal to all seedlings after 4 weeks of treatment. A few small, ineffective nodules were initiated early on some of the latter seedlings, suggesting that effects of nickel on host plant processes rather than Frankia are the primary cause of inhibition of nodulation. This interpretation is supported by the retention of substantial nitrogenase activity in 10-month-old plants 1 day after the treatment with 0.59 mM nickel, when the nickel content of roots and nodules was already maximal. No nitrogenase activity was detected after 3 days, by which time the leaves were almost completely necrotic. Over a 4 day period, most nickel was retained in the roots and nodules. Supplying histidine simultaneously at concentrations equal to, or in excess of, nickel prevented wilting and leaf necrosis, but did not increase translocation of nickel to the shoot.     

Nicotianamine Over-accumulation Confers Resistance to Nickel in <Emphasis Type="Italic">Arabidopsis thaliana</Emphasis>

Nicotianamine is a methionine derivative involved in iron homeostasis, able to bind various other metals in vitro. To investigate its role in vivo, we expressed a nicotianamine synthase cDNA (TcNAS1) isolated from the polymetallic hyperaccumulator Thlaspi caerulescens in Arabidopsis thaliana. Transgenic plants expressing TcNAS1 over-accumulated NA, up to 100-fold more than wild type plants. Furthermore, increased NA levels in different transgenic lines were quantitatively correlated with increased nickel tolerance. The tolerance to nickel is expressed at the cellular level in protoplast experiments and is associated with an increased NA content. We have also shown that the most NA-over accumulating line showed a high tolerance to nickel and a significant Ni accumulation in the leaves when grown on nickel-contaminated soil. Our results highlight a new potential role for nicotianamine in heavy metal tolerance at the cellular but also at the whole plant level, easily transposable to a non-tolerant non-hyperaccumulator species. These results open new perspectives for the modulation of nicotianamine content in plants for phytoremediation.