Nickel hyperaccumulation defends Streptanthus polygaloides (Brassicaceae) against pathogens

The Ni-hyperaccumulating annual, Streptanthus polygaloides, may contain as much as 16,400 ppm Ni (dry weight) in its tissues. The function of Ni hyperaccumulation is not known. We tested the hypothesis that one function of Ni hyperaccumulation in S. polygaloides is defense against pathogens. Growth of pathogenic organisms on Ni-hyperaccumulating plants (averaging 5,630 ppm Ni, produced by growing plants on high-Ni soil) was compared to pathogen growth on nonhyper-accumulating plants (averaging 124 ppm Ni, produced by growing plants on low-Ni soil). Plants containing hyperaccumulated Ni were more slowly infected by a powdery mildew (Erysiphe polygoni) than low-Ni plants. Two strains of the bacterial pathogen Xanthomonas campestris pv. campestris (one a genetically engineered bioluminescent strain) grew in low-Ni plants but not high-Ni plants. Growth of X. campestris pv. campestris was markedly inhibited by Ni concentrations of 400 ppm in artificial media. Growth of the fungus Alternaria brassicicola, which was necrotrophic on S. polygaloides, also was inhibited on high-Ni leaves relative to low-Ni leaves. These results demonstrated negative effects of hyperaccumulated Ni on a taxonomically wide range of pathogenic organisms, supporting the hypothesis that Ni hyperaccumulation defends S. polygaloides against plant pathogens.     

Metal Tolerance and Accumulation Ability of the Ni Hyperaccumulator Streptanthus polygaloides Gray (Brassicaceae)

High concentrations of metals occur in some plant species (termed hyperaccumulators), such as the Ni hyperaccumulator Streptanthus polygaloides. We determined the tolerance of S. polygaloides to, and its accumulation abilities for, six metals (Ni, Zn, Cu, Co, Mn, and Pb). Potting mix concentrations used for all metals ranged from 0 to 1200 μg/g dry weight. For Ni, a treatment of 1600 μg/g was included. For Mn, treatments of 1600, 2000, and 2500 μg/g also were used, and for Pb these concentrations plus 3500 μg/g were included. Germination, plant number per pot, and size at days 30 and 39, number of plants at the end of the experiment (day 49), flower production, and metal concentration in the aboveground biomass were documented. Lead and Ni showed no consistent effects on plant performance, but yielded increased tissue metal concentrations. Streptanthus polygaloides was more sensitive to Co, Cu, and Zn, as ≥ 400 mg/g significantly suppressed plant growth, survival, and flower production. Tissue metal concentrations also were increased to maxima of 1500 μg Co/g, 120 μg Cu/g, and 6000 μg Zn/g. Manganese affected S. polygaloides less markedly, as ≥ 800 mg/kg decreased growth, survival, and flower production. Maximum tissue Mn concentration was 2900 μg/g. We concluded that S. polygaloides would be an appropriate phytoextractor for soils contaminated with Ni or low levels of Co but would not be useful for Cu, Zn, Mn, and Pb.     

Rhizosphere Microbial Densities and Trace Metal Tolerance of the Nickel Hyperaccumulator Alyssum Serpyllifolium Subsp. Lusitanicum

In this study we determine culturable microbial densities (total heterotrophs, ammonifiers, amylolytics and cellulolytics) and bacterial resistance to Co, Cr, and Ni in bulk and rhizosphere soils of three populations of the Ni-hyperaccumulator Alyssum serpyllifolium subsp. lusitanicum and the excluder Dactylis glomerata from ultramafic sites (two populations in Northeast (NE) Portugal (Samil (S), Morais (M)) and one population in Northwest (NW) Spain (Melide (L)). The relationship between bioavailable metal concentrations (H₂O-soluble) and microbial densities were analysed. Significant differences in microbial densities and metal-resistance were observed between the two species and their three populations. The hyperaccumulator showed higher microbial densities (except cellulolytics) and a greater rhizosphere effect, but this was only observed in S and M populations. These populations of A. serpyllifolium also showed selective enrichment of Ni-tolerant bacteria at the rhizosphere where Ni solubility was enhanced (densities of Ni-resistant bacteria were positively correlated with H₂O-soluble Ni). These rhizobacteria could solubilise Ni in the soil and potentially improve phytoextraction strategies.     

Nickel Hyperaccumulation in the Serpentine Flora of Cuba

Extraordinary uptake (hyperaccumulation) of nickel (Ni), reachingconcentrations of 0.1–5.0%, about 1000-times greater thanthose usually found in flowering plants, has been reported overthe period 1948–1996 in about 190 species that grow onNi-rich serpentine soils derived from ultramafic rocks in variousparts of the world. A recent study of the families Buxaceaeand Euphorbiaceae identified a further 80 Ni hyperaccumulatorsfrom the very large ultramafic flora of Cuba, the largest numberfound to date in any one country. A much wider investigationof the elemental content of plants from the Cuban ultramaficflora, reported here with representative analyses of the correspondingsoils, has revealed Ni hyperaccumulation in an additional 50taxa (in 16 genera and eight families). The number of hyperaccumulatorsis greatest on the oldest serpentine soils, which are believedto have been available for colonization for the last 10–30million years. Both Ni hyperaccumulators, and serpentine endemicspecies generally, are much more frequent on these old soils,occurring in the eastern and western extremities of Cuba, thanon those developed within the last million years in the centralpart of the country. Hyperaccumulating plants of the familiesAcanthaceae, Asteraceae, Clusiaceae, Myrtaceae, Ochnaceae, Oleaceae,Rubiaceae and Tiliaceae are discussed.Copyright 1999 Annalsof Botany Company. Nickel, hyperaccumulation, serpentine flora, Cuba.     

Evolutionary lineages of nickel hyperaccumulation and systematics in European Alysseae (Brassicaceae): evidence from nrDNA sequence data

Background and Aims

Nickel (Ni) hyperaccumulation is a rare form of physiological specialization shared by a small number of angiosperms growing on ultramafic soils. The evolutionary patterns of this feature among European members of tribe Alysseae (Brassicaceae) are investigated using a phylogenetic approach to assess relationships among Ni hyperaccumulators at the genus, species and below-species level.

Methods

Internal transcribed spacer (ITS) sequences were generated for multiple accessions of Alysseae. Phylogenetic trees were obtained for the genera of the tribe and Alyssum sect. Odontarrhena. All accessions and additional herbarium material were tested for Ni hyperaccumulation with the dimethylglyoxime colorimetric method.

Key Results

Molecular data strongly support the poorly known hyperaccumulator endemic Leptoplax (Peltaria) emarginata as sister to hyperaccumulator species of Bornmuellera within Alysseae. This is contrary to current assumptions of affinity between L. emarginata and the non-hyperaccumulator Peltaria in Thlaspideae. The lineage Bornmuellera–Leptoplax is, in turn, sister to the two non-hyperaccumulator Mediterranean endemics Ptilotrichum rupestre and P. cyclocarpum. Low ITS sequence variation was found within the monophyletic Alyssum sect. Odontarrhena and especially in A. murale sensu lato. Nickel hyperaccumulation was not monophyletic in any of three main clades retrieved, each consisting of hyperaccumulators and non-hyperaccumulators of different geographical origin.

Conclusions

Nickel hyperaccumulation in Alysseae has a double origin, but it did not evolve in Thlaspideae. In Bornmuellera–Leptoplax it represents an early synapomorphy inherited from an ancestor shared with the calcicolous, sister clade of Mediterranean Ptilotrichum. In Alyssum sect. Odontarrhena it has multiple origins even within the three European clades recognized. Lack of geographical cohesion suggests that accumulation ability has been lost or gained over the different serpentine areas of south Europe through independent events of microevolutionary adaptation and selection. Genetic continuity and strong phenotypic plasticity in the A. murale complex call for a reduction of the number of Ni hyperaccumulator taxa formally recognized.     

Nickel localization on tissues of hyperaccumulator species of Phyllanthus L. (euphorbiaceae) from Ultramafic Areas of Cuba

Two species of perennial Phyllanthus (Euphorbiaceae) (Phyllanthus orbicularis and Phyllanthus discolor, both endemic to ultramafic areas of Cuba, and their natural hybrid, Phyllanthus xpallidus) were selected for metal localization microanalysis. Different plant tissues were analyzed by X-ray fluorescence, inductively coupled plasma—atomic emission spectroscopy, and scanning electron microscopy coupled with an energy-dispersive X-ray probe. All of the studied taxa are nickel (Ni) hyperaccumulators and significant concentrations of this element were found in different leaf and stem tissues. The highest Ni content was found in the laticifer tubes, whereas leaf epidermis Ni content resulted to be much more relevant in terms of total metal storage. Calcium and magnesium were found more evenly distributed in leaf and stem tissues.     

Nickel levels in arthropods associated with Ni hyperaccumulator plants from an ultramafic site in New Caledonia

Arthropods (mainly insects) were collected from a forest site that contained at least six species of Ni hyperaccumulators. Whole body Ni analysis was performed for 12 arthropod taxa, two of which were studied at different life cycle stages. We found two Nitolerant insects. The pentatomid heteropteran Utana viridipuncta, feeding on fruits of the Ni hyperaccumulator Hybanthus austrocaledonicus, contained a mean of 2 600 μNi/g in nymphs and 750 μNi/g in adults. The tephritid fly Bactrocera psidii, feeding on pulp of Sebertia acuminata fruits that contained 6 900 μNi/g, contained 420 μNi/g as larvae that had evacuated their guts and significantly less (65 μNi/g) as adults. European honeybees (Apis mellifera) visiting flowers of the Ni hyperaccumulator H. austrocaledonicus contained significantly more Ni (8‐fold more) than those collected from flowers of Myodocarpus fraxinifolius, a non‐hyperaccumulator. Our results show that some insects feed on Ni hyperaccumulator plants and that their feeding mobilizes Ni into local food webs.     

NICKEL UPTAKE BY CALIFORNIAN STREPTANTHUS AND CAULANTHUS WITH PARTICULAR REFERENCE TO THE HYPERACCUMULATOR S. POLYGALOIDES GRAY (BRASSICACEAE)

The nickel content of herbarium specimens of Californian Streptanthus and Caulanthus has been determined. Serpentine-tolerant species had nickel values of 10–100 μg/g (dry mass) except in the case of S. polygaloides Gray which had nickel values in the range 3,300–14,800 μg/g (0.33%–1.48%). This is the first hyperaccumulator of nickel found in the Americas. It is suggested that the later classification of S. polygaloides as the monotypic Microsemia polygaloides (Gray) Greene may be justified if the nickel accumulation data are used to reinforce the existing marked morphological differences from other Streptanthus.     

Crop rotation associating a legume and the nickel hyperaccumulator <Emphasis Type="Italic">Alyssum murale</Emphasis> improves the structure and biofunctioning of an ultramafic soil

Nickel (Ni) agromining aims to phytoextract heavy metals using hyperaccumulators whilst at the same time rehabilitating ultramafic soils. After removing the bioavailable metal, ultramafic soils are improved in terms of their agronomic properties with the aim of future agricultural uses. The low fertility of ultramafic soils can be compensated by integrating legumes already used in traditional agro-systems because of their importance in soil nitrogen enrichment. However, few studies have evaluated the potential profits of legumes on Ni agromining and their potential benefits on soil biological fertility. Here, we characterized the effect of a crop rotation with two plants, a legume (Vicia sativa) and a hyperaccumulator (Alyssum murale), on the phytoextraction efficiency and on soil structure and biofunctioning. A pot experiment was set up in controlled conditions to grow A. murale and four treatments were tested: rotation with V. sativa (Ro), fertilized mono-culture (FMo), non-fertilized mono-culture (NFMo) and bare soil without plants (BS). No significant difference was found between the Ro and NFMo treatments for the dry biomass yield. However, the Ro treatment showed the highest Ni concentrations ([Ni]) in A. murale shoots compared to FMo and NFMo treatments. The Ro treatment plants had more than twice as many leaves [Ni] compared to FMo. Soil physico-chemical analyses showed that the Ro treatment was better structured and showed the highest presence of bacterial micro-aggregates, as well as less non-aggregated particles. Legumes integration in Ni-agromining systems could be a pioneering strategy to reduce chemical inputs and to improve soil biofunctioning and thus fertility.     

Interaction of nickel and manganese in accumulation and localization in leaves of the Ni hyperaccumulators Alyssum murale and Alyssum corsicum

The genus Alyssum contains >50 Ni hyperaccumulator species; many can achieve >2.5% Ni in dry leaf. In soils with normal Mn levels, Alyssum trichome bases were previously observed to accumulate Ni and Mn to high levels. Here we report concentration and localization patterns in A. murale and A. corsicum grown in soils with nonphytotoxic factorial additions of Ni and Mn salts. Four leaf type subsets based on size and age accumulated Ni and Mn similarly. The greatest Mn accumulation (10 times control) was observed in A. corsicum with 40 mmol Mn kg^?1 and 40 mmol Ni kg^?1 added to potting soil. Whole leaf Ni concentrations decreased as Mn increased. Synchrotron X-ray fluorescence mapping of whole fresh leaves showed localized in distinct high-concentration Mn spots associated with trichomes, Ni and Mn distributions were strongly spatially correlated. Standard X-ray fluorescence point analysis/mapping of cryofractured and freeze-dried samples found that Ni and Mn were co-located and strongly concentrated only in trichome bases and in cells adjacent to trichomes. Nickel concentration was also strongly spatially correlated with sulfur. Results indicate that maximum Ni phytoextraction by Alyssum may be reduced in soils with higher phytoavailable Mn, and suggest that Ni hyperaccumulation in Alyssum species may have developed from a Mn handling system.     

Bacterially Induced Weathering of Ultramafic Rock and Its Implications for Phytoextraction

The bioavailability of metals in soil is often cited as a limiting factor of phytoextraction (or phytomining). Bacterial metabolites, such as organic acids, siderophores, or biosurfactants, have been shown to mobilize metals, and their use to improve metal extraction has been proposed. In this study, the weathering capacities of, and Ni mobilization by, bacterial strains were evaluated. Minimal medium containing ground ultramafic rock was inoculated with either of two Arthrobacter strains: LA44 (indole acetic acid [IAA] producer) or SBA82 (siderophore producer, PO₄ solubilizer, and IAA producer). Trace elements and organic compounds were determined in aliquots taken at different time intervals after inoculation. Trace metal fractionation was carried out on the remaining rock at the end of the experiment. The results suggest that the strains act upon different mineral phases. LA44 is a more efficient Ni mobilizer, apparently solubilizing Ni associated with Mn oxides, and this appeared to be related to oxalate production. SBA82 also leads to release of Ni and Mn, albeit to a much lower extent. In this case, the concurrent mobilization of Fe and Si indicates preferential weathering of Fe oxides and serpentine minerals, possibly related to the siderophore production capacity of the strain. The same bacterial strains were tested in a soil-plant system: the Ni hyperaccumulator Alyssum serpyllifolium subsp. malacitanum was grown in ultramafic soil in a rhizobox system and inoculated with each bacterial strain. At harvest, biomass production and shoot Ni concentrations were higher in plants from inoculated pots than from noninoculated pots. Ni yield was significantly enhanced in plants inoculated with LA44. These results suggest that Ni-mobilizing inoculants could be useful for improving Ni uptake by hyperaccumulator plants.     

Nickel bioavailability in an ultramafic toposequence in the Vosges Mountains (France)

A serpentinised harzburgite outcrop located in the Vosges Mountains hosts a population of the Ni-hyperaccumulator Thlaspi caerulescens J. & C. Presl. A complete study was undertaken to relate the variability of Ni availability along the ultramafic toposequence to pedogenesis, soil mineralogy and functioning with X-Ray Diffraction, Transmission Electron Microscope observations coupled with Isotopic Exchange Kinetics and diethylenetriamine pentaacetic acid extraction of Ni. The soil profiles ranging from Dystric Cambisol to Hypermagnesic Hypereutric Cambisol were distributed unevenly along the toposequence probably due to geochemical variability of the bedrock and also complex quaternary erosion features. The richest soils were characterised by slight mineral weathering leading to Ni, Cr and Fe accumulation in the B horizons whereas the lowest saturated soils had very low-metal contents. Most soil minerals were inherited from the parent materials and there were only few traces of formation of secondary minerals. Primary minerals (e.g. serpentine, chlorite) contained low Ni concentrations (0.2%) whereas neoformed goethite, mainly in the B horizons of the richest soils, contained up to 4.3% Ni. Ni was probably sorbed onto amorphous Fe oxy−hydroxide particles (oxalate extraction) rather than incorporated within the crystal lattice of goethite. Ni availability in the B horizon of Hypereutric Cambisols was extremely high and so was the oxalate extractable Fe. At the toposequence level, there was a high level of Ni availability in the least weathered soils and a very low-availability level in the more intensively weathered soils (strongly acidic pH). Ni availability was unexpectedly positively correlated to pH and was controlled by soil mineralogy and Ni-bearing mineral phases. Ni hyperaccumulation (above 1,000 mg kg⁻¹) by native T. caerulescens was only reached in the Ni-rich soils as a consequence of the local edaphic factors. Ni uptake by T. caerulescens is strongly regulated by Ni availability in soils and therefore related to pedogenesis.     

Nickel distribution in the hyperaccumulator Alyssum serpyllifolium Desf. spp. from the Iberian Peninsula

Abstract

The relative concentration and distribution of nickel (Ni) in vegetative tissues (leaves, stems and trichomes) and reproductive organs (seeds) was studied using energy-dispersive X-ray microanalysis (EDXS) and scanning electron microscopy (SEM) in two previously studied Ni-hyperaccumulator subspecies of Alyssum serpyllifolium Desf. growing naturally in ultramafic soils of the Iberian Peninsula: A. serpyllifolium ssp. lusitanicum Dudley & P. Silva and A. serpyllifolium ssp. malacitanum Rivas Goday ex G. López. Both taxa showed that Ni accumulates preferentially in the leaves, exhibiting a homogeneous distribution on both epidermis surfaces. The highest Ni concentrations were found inside the epidermal cells and at the base of trichome stalks. Ni accumulation in seeds was lower than in the vegetative organs. The location of Ni in these plants allows us to postulate that its accumulation is a protection mechanism against external stress.     

Effects of calcium on nickel tolerance and accumulation in Alyssum species and cabbage grown in nutrient solution

Nickel (Ni) phytoextraction using hyperaccumulator plant species to accumulate Ni from mineralized and contaminated soils rich in Ni is undergoing commercial development. Serpentinite derived soils have a very low ratio of Ca/Mg among soils due the nature of the parent rock. In crop plants, soil Ca reduces Ni uptake and phytotoxicity, so it is possible that the low Ca of serpentine soils could limit hyperaccumulator plant tolerance of serpentine soils used for commercial phytomining. In this study, we investigated the effects of varied Ca concentration in the presence of high Mg characteristic of serpentine soils on Ni uptake and tolerance by serpentine-endemic species Alyssum murale Waldst. et Kit. and A. pintodasilvae T.R. Dudley in comparison with cabbage (Brassica oleracea L. var. capita) in a nutrient solution study. The levels of Ca and Mg used were based on serpentine and normal soils, and Ni was based on achieving over 1% Ni in Alyssum shoots in preliminary tests. Varied solution concentrations of Ni (31.6–1,000 μM for Alyssum, 1.0–10 μM for cabbage) and Ca (0.128–5 mM) were used in a factorial experimental design; 2 mM Mg was used to mimic serpentine soils. Alyssum spp. showed much greater tolerance to high Ni, high Mg, and low Ca solution concentrations than cabbage. For Alyssum spp., Ni induced phytotoxicity was only apparent at 1,000 μM Ni with relatively low and high Ca/Mg quotient. In the 1,000 μM Ni treatment, shoot Ni concentrations ranged from 8.18 to 22.8 g kg^?1 for A. murale and 7.60 to 16.0 g kg^?1 for A. pintodasilvae. Normal solution Ca concentrations (0.8–2 mM) gave the best yield across all Ni treatments for the Alyssum species tested. It was clear that solution Ca levels affected shoot Ni concentration, shoot yield and Ni translocation from root to shoot, but the relation was non-linear, increasing with increasing Ca up to 2 mM Ca, then declining at the highest Ca. Our results indicate that Ca addition to high Mg serpentine soils with very low Ca/Mg ratio may reduce Ni phytotoxicity and improve annual Ni phytoextraction by Alyssum hyperaccumulator species. Removal of shoot biomass in phytomining will require Ca application to maintain full yield potential.     

Nickel hyperaccumulation by Streptanthus polygaloides protects against the folivore Plutella xylostella (Lepidoptera: Plutellidae)

We determined the effectiveness of Ni as an elemental defence of Streptanthus polygaloides (Brassicaceae) against a crucifer specialist folivore, diamondback moth (DBM), Plutella xylostella. An oviposition experiment used arrays of S. polygaloides grown on Ni-amended (high-Ni) soil interspersed with plants grown on unamended (low-Ni) soil and eggs were allowed to hatch and larvae fed freely among plants in the arrays. We also explored oviposition preference by allowing moths to oviposit on foil sheets coated with high- or low-Ni plant extract. This was followed by an experiment using low-Ni plant extract to which varying amounts of Ni had been added and an experiment using sheets coated with sinigrin (allyl glucosinolate) as an oviposition stimulant. Diamondback moths laid 2.5-fold more eggs on low-Ni plants than on high-Ni plants and larval feeding was greater on low-Ni plants. High-Ni plants grew twice as tall, produced more leaves, and produced almost 3.5-fold more flowers. Low-Ni plants contained more allyl glucosinolate than high-Ni plants and moths preferred to oviposit on foil sheets dipped in low-Ni plant extract. Moths showed no preference when Ni concentration of low-Ni extract was varied and overwhelmingly preferred sinigrin coated sheets. We conclude that Ni hyperaccumulation is an effective elemental defence against this herbivore, increasing plant fitness through a combination of toxicity to DBM larvae and decreased oviposition by adults.     

The Importance of Nickel Phytoavailable Chemical Species Characterization in Soil for Phytoremediation Applicability

In this work Ni speciation in natural and spiked soils (with similar total concentration) was studied. Spiked soils were prepared by addition of NiSO₄.6H₂O to obtain concentration similar to the one in natural soils. In soils mobile species were determined with a simplified sequential extraction as follows: H₂O for water-soluble metal, KNO₃ for exchangeable species, DTPA for complexed/adsorbed species. Results show that in spiked soils the exchangeable and adsorbed Ni concentrations are considerably higher than in natural soils. A study of plant uptake was carried out in order to evaluate the relation between mobile species and phyto-availability. Alfalfa (Medicago sativa L.), even though it is not a hyperaccumulator, was selected for its tolerance to high metal concentrations in soil. Preliminary results show a very high correlation between Ni mobile species and Ni uptake by alfalfa. Significant differences were found between spiked and natural soils. In the latter, high levels of total Ni did not correspond to relevant uptake as in the case of spiked soil. Results stress the importance of evaluating preliminarly heavy metal speciation in soil before planning phytoremediation procedures.     

Bacterial community diversity and functional roles in the rhizosphere of Rinorea cf. bengalensis and Phyllanthus rufuschaneyi under a nickel concentration gradient

Plant and Soil - The tropical nickel (Ni) hyperaccumulator plants, Rinorea cf. bengalensis and Phyllanthus rufuschaneyi, are locally common on ultramafic soils in the Malaysian state of Sabah on...     

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.     

Arbuscular mycorrhizal fungi from New Caledonian ultramafic soils improve tolerance to nickel of endemic plant species

In order to improve knowledge about the role of arbuscular mycorrhizal fungi (AMF) in the tolerance to heavy metals in ultramafic soils, the present study investigated the influence of two Glomus etunicatum isolates from New Caledonian ultramafic maquis (shrubland), on nickel tolerance of a model plant species Sorghum vulgare, and of two ultramafic endemic plant species, Alphitonia neocaledonica and Cloezia artensis. In a first step, plants were grown in a greenhouse, on sand with defined concentrations of Ni, to appreciate the effects of the two isolates on the alleviation of Ni toxicity in controlled conditions. In a second step, the influence of the AMF on A. neocaledonica and C. artensis plants grown in a New Caledonian ultramafic soil rich in extractable nickel was investigated. Ni reduced mycorrhizal colonization and sporulation of the fungal isolates, but the symbionts increased plant growth and adaptation of endemic plant species to ultramafic conditions. One of the two G. etunicatum isolates showed a stronger positive effect on plant biomass and phosphorus uptake, and a greater reduction in toxicity symptoms and Ni concentration in roots and shoots. The symbionts seemed to act as a barrier to the absorption of Ni by the plant and reduced root-to-shoot Ni translocation. Results indicate the potential of selected native AMF isolates from ultramafic areas for ecological restoration of such degraded ecosystems.     

Bacterial Communities Associated with Flowering Plants of the Ni Hyperaccumulator Thlaspi goesingense

Thlaspi goesingense is able to hyperaccumulate extremely high concentrations of Ni when grown in ultramafic soils. Recently it has been shown that rhizosphere bacteria may increase the heavy metal concentrations in hyperaccumulator plants significantly, whereas the role of endophytes has not been investigated yet. In this study the rhizosphere and shoot-associated (endophytic) bacteria colonizing T. goesingense were characterized in detail by using both cultivation and cultivation-independent techniques. Bacteria were identified by 16S rRNA sequence analysis, and isolates were further characterized regarding characteristics that may be relevant for a beneficial plant-microbe interaction—Ni tolerance, 1-aminocyclopropane-1-carboxylic acid (ACC) deaminase and siderophore production. In the rhizosphere a high percentage of bacteria belonging to the Holophaga/Acidobacterium division and α-Proteobacteria were found. In addition, high-G+C gram-positive bacteria, Verrucomicrobia, and microbes of the Cytophaga/Flexibacter/Bacteroides division colonized the rhizosphere. The community structure of shoot-associated bacteria was highly different. The majority of clones affiliated with the Proteobacteria, but also bacteria belonging to the Cytophaga/Flexibacter/Bacteroides division, the Holophaga/Acidobacterium division, and the low-G+C gram-positive bacteria, were frequently found. A high number of highly related Sphingomonas 16S rRNA gene sequences were detected, which were also obtained by the cultivation of endophytes. Rhizosphere isolates belonged mainly to the genera Methylobacterium, Rhodococcus, and Okibacterium, whereas the majority of endophytes showed high levels of similarity to Methylobacterium mesophilicum. Additionally, Sphingomonas spp. were abundant. Isolates were resistant to Ni concentrations between 5 and 12 mM; however, endophytes generally tolerated higher Ni levels than rhizosphere bacteria. Almost all bacteria were able to produce siderophores. Various strains, particularly endophytes, were able to grow on ACC as the sole nitrogen source.