Growth and mineral element composition in two ecotypes of Thlaspi caerulescens on Cd contaminated soil

The heavy metal hyperaccumulator Thlaspi caerulescens occurs both on heavy metal polluted soils (metallicolous ecotype MET) and on soils with normal heavy metal content (non-metallicolous ecotype: NMET). In order to assess the extent and structure of variation in growth, shoot accumulation of Cd, Zn and mineral element (Ca, Mg, K, Fe), a MET ecotype from Belgium and a NMET ecotype from Luxembourg were studied. Seven maternal families from two populations of each ecotype were grown on both Cd and Zn contaminated soil. Although both ecotypes presented a similar heavy metal tolerance in the experimental conditions tested, they differed in several points. The MET populations had markedly higher biomass and higher root:shoot ratio compared to NMET populations. The Zn, and at lesser extent, the Cd hyperaccumulation capacity tended to be higher in the NMET populations. The same trend was observed for the foliar concentrations of Mg, Ca and Fe with NMET populations having higher concentrations compared to MET ones. Cd and Zn concentrations were negatively correlated with the biomass of both ecotype. However, the negative correlation between the Zn and biomass was much lower in MET ecotype suggesting a tighter control of internal Zn concentration in this ecotype. Finally, although the Cd phytoextraction capacity was similar in both ecotype, a higher Zn phytoextraction capacity was detected in NMET ecotype when these plants grow on moderate Cd and Zn concentrations.     

Impact of cadmium and zinc on growth and water status of Zygophyllum fabago in two contrasting metallicolous populations from SE Spain: comparison at whole plant and tissue level

Cadmium and zinc accumulation and toxicity were assessed in whole plants and callus culture of two Zygophyllum fabago populations originating from two metallicolous sites in Murcia (southeast Spain), La Peña and Mazarrón, the first containing 2.8‐times more Cd and five‐times more Zn than the second. Seedlings from both ecotypes were exposed for 3 weeks to 1 or 10 μm Cd, and to 10 or 100 μm Zn in nutrient solution in a controlled environment. Calli from both ecotypes were exposed to 0.01, 0.1 or 1 mm Cd, and to 0.1, 1 or 5 mm Zn. Plants from both populations exhibited similar tolerance to Zn, while tolerance to Cd appeared more important in plants from La Peña than those from Mazarrón. Only minor differences were recorded in final Cd accumulation, with higher Cd retention in roots and stems of plants from La Peña. In both populations, transient decreases in the rate of Zn intake and translocation from root to shoot were recorded. This reduction in ion uptake was not more efficient for the population from the most contaminated area compared to the less contaminated area. Similar concentrations of Cd were found in cotyledon‐derived calli from the two populations, but absorbed Cd induced conspicuous water stress in calli issues from Mazarrón but not in those from La Peña. It is concluded that, beside comparable levels of heavy metal concentration in tissues, the physiological strategy of tolerance may differ according to the metal and according to the considered population.     

Culturable bacteria from Zn‐ and Cd‐accumulating Salix caprea with differential effects on plant growth and heavy metal availability

Aims: To characterize bacteria associated with Zn/Cd‐accumulating Salix caprea regarding their potential to support heavy metal phytoextraction. Methods and Results: Three different media allowed the isolation of 44 rhizosphere strains and 44 endophytes, resistant to Zn/Cd and mostly affiliated with Proteobacteria, Actinobacteria and Bacteroidetes/Chlorobi. 1‐Aminocyclopropane‐1‐carboxylic acid deaminase (ACCD), indole acetic acid and siderophore production were detected in 41, 23 and 50% of the rhizosphere isolates and in 9, 55 and 2% of the endophytes, respectively. Fifteen rhizosphere bacteria and five endophytes were further tested for the production of metal‐mobilizing metabolites by extracting contaminated soil with filtrates from liquid cultures. Four Actinobacteria mobilized Zn and/or Cd. The other strains immobilized Cd or both metals. An ACCD‐ and siderophore‐producing, Zn/Cd‐immobilizing rhizosphere isolate (Burkholderia sp.) and a Zn/Cd‐mobilizing Actinobacterium endophyte were inoculated onto S. caprea. The rhizosphere isolate reduced metal uptake in roots, whereas the endophyte enhanced metal accumulation in leaves. Plant growth was not promoted. Conclusions: Metal mobilization experiments predicted bacterial effects on S. caprea more reliably than standard tests for plant growth‐promoting activities. Significance and Impact of the Study: Bacteria, particularly Actinobacteria, associated with heavy metal‐accumulating Salix have the potential to increase metal uptake, which can be predicted by mobilization experiments and may be applicable in phytoremediation.     

Rhizosphere bacteria affect growth and metal uptake of heavy metal accumulating willows

A variety of plants growing on metalliferous soils accumulate metals in their harvestable parts and have the potential to be used for phytoremediation of heavy metal polluted land. There is increasing evidence that rhizosphere bacteria contribute to the metal extraction process, but the mechanisms of this plant–microbe interaction are not yet understood. In this study ten rhizosphere isolates obtained from heavy metal accumulating willows affiliating with Pseudomonas, Janthinobacterium, Serratia, Flavobacterium, Streptomyces and Agromyces were analysed for their effect on plant growth, Zn and Cd uptake. In plate assays Zn, Cd and Pb resistances and the ability of the bacteria to produce indole-3-acetic acid (IAA), 1-amino-cyclopropane-1-carboxylic acid deaminase (ACC deaminase) and siderophores were determined. The isolates showed resistance to high Zn concentrations, indicating an adaptation to high concentrations of mobile Zn in the rhizosphere of Salix caprea. Four siderophore producers, two IAA producers and one strain producing both siderophores and IAA were identified. None of the analysed strains produced ACC deaminase. Metal mobilization by bacterial metabolites was assessed by extracting Zn and Cd from soil with supernatants of liquid cultures. Strain Agromyces AR33 almost doubled Zn and Cd extractability, probably by the relase of Zn and Cd specific ligands. The remaining strains, immobilized both metals. When Salix caprea plantlets were grown in γ-sterilized, Zn/Cd/Pb contaminated soil and inoculated with the Zn resistant isolates, Streptomyces AR17 enhanced Zn and Cd uptake. Agromyces AR33 tendentiously promoted plant growth and thereby increased the total amount of Zn and Cd extracted from soil. The IAA producing strains did not affect plant growth, and the siderophore producers did not enhance Zn and Cd accumulation. Apparently other mechanisms than the production of IAA, ACC deaminase and siderophores were involved in the observed plant–microbe interactions.     

The effects of heavy metal pollution on genetic diversity in zinc/cadmium hyperaccumulator Sedum alfredii populations

The genetic diversity and population structure of seven populations of Sedum alfredii growing in lead/zinc (Pb/Zn) mine spoils or in uncontaminated soils from eastern and southern China were investigated using random amplified polymorphic DNA (RAPD) technology. Four of the sampled sites were heavily contaminated with heavy metals (Zn, Cd, Pb), and extremely high concentrations of Zn, Cd, and Pb were found among these corresponding populations. A significant reduction of genetic diversity was detected in the mining populations. The reduction of genetic diversity could be derived from a bottleneck effect and might also be attributed to the prevalence of vegetative reproduction of the mining populations. Analysis of molecular variance (AMOVA) and the unweighted pair group method with arithmetic mean (UPGMA) tree derived from genetic distances further corroborated that the genetic differentiation between mine populations and uncontaminated populations was significant. Polymorphism with the heavy metal accumulation capability of S. alfredii probably due to the genetic variation among populations and heavy metal contamination could have more impact on the genetic diversity and population structure of S. alfredii populations than geographic distance.     

Demographic history of the trace metal hyperaccumulator Noccaea caerulescens (J. Presl and C. Presl) F. K. Mey. in Western Europe

Noccaea caerulescens (Brassicaceae) is a major pseudometallophyte model for the investigation of the genetics and evolution of metal hyperaccumulation in plants. We studied the population genetics and demographic history of this species to advance the understanding of among‐population differences in metal hyperaccumulation and tolerance abilities. Sampling of seven to 30 plants was carried out in 62 sites in Western Europe. Genotyping was carried out using a combination of new chloroplast and nuclear neutral markers. A strong genetic structure was detected, allowing the definition of three genetic subunits. Subunits showed a good geographic coherence. Accordingly, distant metallicolous populations generally belonged to distinct subunits. Approximate Bayesian computation analysis of demographic scenarios among subunits further supported a primary isolation of populations from the southern Massif Central prior to last glacial maximum, whereas northern populations may have derived during postglacial recolonization events. Estimated divergence times among subunits were rather recent in comparison with the species history, but certainly before the establishment of anthropogenic metalliferous sites. Our results suggest that the large‐scale genetic structure of N. caerulescens populations pre‐existed to the local adaptation to metalliferous sites. The population structure of quantitative variation for metal‐related adaptive traits must have established independently in isolated gene pools. However, features of the most divergent genetic unit (e.g. extreme levels of Cd accumulation observed in previous studies) question the putative relationships between adaptive evolution of metal‐related traits and subunits isolation. Finally, admixture signals among distant metallicolous populations suggest a putative role of human activities in facilitating long‐distance genetic exchanges.     

Morphological and genetic distinctiveness of metallicolous and non-metallicolous populations of Armeria maritima s.l. (Plumbaginaceae) in Poland

Patterns of morphological, genetic and epigenetic variation (DNA methylation pattern) were investigated in metallicolous (M) and non‐metallicolous (NM) populations of Armeria maritima. A morphological study was carried out using plants from six natural populations grown in a greenhouse. Morphological variation was assessed using seven traits. On the basis of this study, three representative populations were selected for molecular analyses using metAFLP to study sequence‐ and methylation‐based DNA variation. Only one morphological trait (length of outer involucral bracts) was common to both metallicolous populations studied; however, the level of variation was sufficient to differentiate between M and NM populations. Molecular analyses showed the existence of naturally occurring epigenetic variation in A. maritima populations, as well as structuring into distinct between and within population components. We show that patterns of population genetic structure differed depending on the information used in the study. Analysis of sequence‐based information data demonstrates the presence of three well‐defined and genetically differentiated populations. Methylation‐based data show that two major groups of individuals are present, corresponding to the division into M and NM populations. These results were confirmed using different analytical approaches, which suggest that the DNA methylation pattern is similar in both M populations. We hypothesise that epigenetic processes may be involved in microevolution leading to development of M populations in A. maritima.     

Metal accumulation and arbuscular mycorrhizal status in metallicolous and nonmetallicolous populations of <Emphasis Type="Italic">Pteris vittata</Emphasis> L. and <Emphasis Type="Italic">Sedum alfredii</Emphasis> Hance

Although Pteris vittata L. and Sedum alfredii Hance have been identified as an As hyperaccumulator and a Zn/Cd hyperaccumulator, respectively, for a few years, variations in metal accumulation among populations and their arbuscular mycorrhizal (AM) status have not been fully explored. Six populations of P. vittata and four populations of S. alfredii from southeast China were investigated. Up to 1,373 As, 680 Pb, 376 Zn, 4.8 Cd, 169 Cu mg kg⁻¹ in fronds of P. vittata and 358 As, 2,290 Pb, 23,403 Zn, 708 Cd, 342 Cu mg kg⁻¹ in shoots of S. alfredii were detected. Constitutive properties of As and Zn hyperaccumulation in metallicolous populations of P. vittata and S. alfredii, respectively, were confirmed. However, Cd hyperaccumulation in S. alfredii varied among populations. The two hyperaccumulators varied in efficiency in taking up other heavy metals. Different metal tolerance strategies adopted by the two hyperaccumulators varied among plant species and metal species. Low to moderate levels of AM colonization in P. vittata (4.2–12.8%) and S. alfredii (8.5–45.8%) were observed at uncontaminated and metal-contaminated sites. The relationship between metal concentrations and AM colonization in the two hyperacumulators was also examined. The abundance of AM fungal spores ranged from 16 to 190 spores per 25 g soil. Glomus microaggregatum, Glomus mosseae, Glomus brohultii and Glomus geosporum were the most common species associated with both P. vittata and S. alfredii. To our knowledge, this is the first report of AM fungal status in rhizosphere of P. vittata and S. alfredii.     

Different strategies of Cd tolerance and accumulation in Arabidopsis halleri and Arabidopsis arenosa

Pseudometallophytes are commonly used to study the evolution of metal tolerance and accumulation traits in plants. Within the Arabidopsis genus, the adaptation of Arabidopsis halleri to metalliferous soils has been widely studied, which is not the case for the closely related species Arabidopsis arenosa. We performed an in-depth physiological comparison between the A. halleri and A. arenosa populations from the same polluted site, together with the geographically close non-metallicolous (NM) populations of both species. The ionomes, growth, photosynthetic parameters and pigment content were characterized in the plants that were growing on their native site and in a hydroponic culture under Cd treatments. In situ, the metallicolous (M) populations of both species hyperaccumulated Cd and Zn. The NM population of A. halleri hyperaccumulated Cd and Zn while the NM A. arenosa did not. In the hydroponic experiments, the NM populations of both species accumulated more Cd in their shoots than the M populations. Our research suggests that the two Arabidopsis species evolved different strategies of adaptation to extreme metallic environments that involve fine regulation of metal homeostasis, adjustment of the photosynthetic apparatus and accumulation of flavonols and anthocyanins.     

Metal concentration and metal mass of metallicolous, non metallicolous and serpentine Noccaea caerulescens populations, cultivated in different growth media

Aims

Evaluate the genetic and environmental variability of metal concentration and metal mass of Noccaea caerulescens, from metalliferous (MET), non metalliferous (NMET) and serpentine (SERP) soils.

Methods

18 populations were cultivated in 18 different growth conditions, such as a soil mine tailing, soils amended with zinc (Zn), cadmium (Cd) and nickel (Ni) salts (in mixtures or in monometallic salts) and a hydroponic solution with two Zn concentrations.

Results

MET populations had Zn concentrations lower than NMET and SERP in the different soils but higher Cd mass (the product of aerial biomass and foliar metal concentration). SERP had the highest Ni concentration and Ni mass values. The addition of Cd or Ni to a Zn-contaminated soil significantly decreases Zn concentration. In hydroponics, MET and NMET had equivalent Zn concentrations but these were three times higher than those obtained in soil experiments. Zn mass of NMET was significantly lower than MET with the latter having Zn mass values largely above those obtained in mine soil.

Conclusions

Results showed a large heterogeneity of responses among populations depending on the substrate used, and it was not possible to correctly assign a single population to its accurate origin with only one experiment. Finally, data on metal concentration obtained in culture soils are closer to those in field soils than those from hydroponics so that they could give a more accurate information on the accumulating capacity of Noccaea caerulescens and its use in phytoextraction of metals in field conditions.     

Bioaccumulation and Physiological Response of Five Willows to Toxic Levels of Cadmium and Zinc

We evaluated the phytoremediation potential of Salix spp. exposed to high cadmium (Cd) and zinc (Zn) concentrations to select feasible plant materials for restoration and revegetation of mining soil contaminated by heavy metals on the basis of their Cd and Zn accumulation, Cd-Zn interaction on bioaccumulation, and the changes of photosynthetic parameters. The Cd and Zn concentrations were in the order of root > leaf > stem, regardless of the species. In the combined Cd and Zn treatment, the leaf and stem Cd concentration in all species were higher relative to Cd-alone treatment. In contrast, the Zn concentration in plant tissues when exposed to the combined Cd + Zn treatment decreased relative to the Zn-alone treatment. The translocation factor (TF) of Cd and Zn from root to leaf was generally higher compared to TF from root to stem than those in the single treatment. The Cd + Zn treatments resulted in enhanced translocation of Cd from root to aboveground tissue (synergistic), while the same treatment suppressed the Zn translocation from root to leaf and stem (antagonistic). The reduction of photosynthetic parameters in Zn alone and Cd + Zn treatments was generally higher than that of Cd-alone treatment. Among the different species, S. caprea and P. alba×glandulosa have the lowest photosynthetic reduction relative to the control. Overall, S. caprea could be a potential candidate for phytoremediation of Cd- and Zn-contaminated sites.     

ASSOCIATED BACTERIA INCREASE THE PHYTOEXTRACTION OF CADMIUM AND ZINC FROM A METAL-CONTAMINATED SOIL BY MYCORRHIZAL WILLOWS

In order to enhance phytoremediation efficiency, we investigated the effects of dual inoculation with ectomycorrhizal fungi and the ectomycorrhiza associated bacteria Micrococcus luteus and Sphingomonas sp. on the growth and metal accumulation of willows (Salix viminalis x caprea) on contaminated soil. The bacterial strains were previously collected from sporocarps of ectomycorrhizal fungi. The bacteria increased plant growth and the mycorrhizal dependency of willows colonized with the ectomycorrhizal fungus Hebeloma crustuliniforme. The total cadmium (Cd) and zinc (Zn) accumulation in the shoot biomass was increased after inoculation with the fungal strain Hebeloma crustuliniforme in combination with Micrococcus luteus up to 53% and in combination with Sphingomonas sp. up to 62%, respectively. The dual inoculation in combination with Laccaria laccata did not increase the accumulation of Cd and Zn in the willows. We conclude that associated bacteria can enhance the ectomyorrhiza formation and growth of willows and, thereby, the Cd and Zn accumulation in the plant biomass. The results suggest that bacterial support of root growth promoting ectomycorrhizal fungi may be a promising approach to improve the remediation of metal-contaminated soils by using willows.     

Differential effects of AM fungal isolates on Medicago truncatula growth and metal uptake in a multimetallic (Cd, Zn, Pb) contaminated agricultural soil

Toxic metal accumulation in soils of agricultural interest is a serious problem needing more attention, and investigations on soil–plant metal transfer must be pursued to better understand the processes involved in metal uptake. Arbuscular mycorrhizal (AM) fungi are known to influence metal transfer in plants by increasing plant biomass and reducing metal toxicity to plants even if diverging results were reported. The effects of five AM fungi isolated from metal contaminated or non-contaminated soils on metal (Cd, Zn) uptake by plant and transfer to leachates was assessed with Medicago truncatula grown in a multimetallic contaminated agricultural soil. Fungi isolated from metal-contaminated soils were more effective to reduce shoot Cd concentration. Metal uptake capacity differed between AM fungi and depended on the origin of the isolate. Not only fungal tolerance and ability to reduce metal concentrations in plant but also interactions with rhizobacteria affected heavy metal transfer and plant growth. Indeed, thanks to association with nodulating rhizobacteria, one Glomus intraradices inoculum increased particularly plant biomass which allowed exporting twofold more Cd and Zn in shoots as compared to non-mycorrhizal treatment. Cd concentrations in leachates were variable among fungal treatments, but can be significantly influenced by AM inoculation. The differential strategies of AM fungal colonisation in metal stress conditions are also discussed.     

Heavy metals contamination and accumulation in submerged macrophytes in an urban river in China

Deteriorating urban water quality has attracted considerable attention in China. We investigated the contamination levels and distribution of heavy metals (As, Cd, Cu, Ni, Pb, and Zn) in Yuxi River water and sediments, and assessed the heavy metal accumulation capability of five species of submerged macrophytes: Vallisneria natans (Lour.) Hara, Potamogeton pectinatus L., Hydrilla verticillata (L. f.) Royle, Myriophyllum spicatum L., and Potamogeton crispus L. Samples were collected from upstream and downstream locations in different season. The results showed that the levels of heavy metals in the downstream areas were higher than in the upstream areas. Heavy metal concentrations in the river water during the dry seasons were higher than those during the rainy seasons, and the opposite results appeared in sediments and submerged macrophytes. In general, the river was slightly contaminated by heavy metals, and the concentrations of Pb and Ni in this river should serve as a warning, while Cd and Zn pollution in the sediments desperately needs to be removed. Furthermore, Potamogeton pectinatus L. showed a higher accumulation capacity for these metals among the five native submerged macrophytes and could be defined as a hyperaccumulator for Cd. Therefore, the potential use of native aquatic plants in contaminated rivers is worth further exploration.     

Proteomic analysis of Arabidopsis halleri shoots in response to the heavy metals cadmium and zinc and rhizosphere microorganisms

Arabidopsis halleri has the rare ability to colonize heavy metal‐polluted sites and is an emerging model for research on adaptation and metal hyperaccumulation. The aim of this study was to analyze the effect of plant–microbe interaction on the accumulation of cadmium (Cd) and zinc (Zn) in shoots of an ecotype of A. halleri grown in heavy metal‐contaminated soil and to compare the shoot proteome of plants grown solely in the presence of Cd and Zn or in the presence of these two metals and the autochthonous soil rhizosphere‐derived microorganisms. The results of this analysis emphasized the role of plant–microbe interaction in shoot metal accumulation. Differences in protein expression pattern, identified by a proteomic approach involving 2‐DE and MS, indicated a general upregulation of photosynthesis‐related proteins in plants exposed to metals and to metals plus microorganisms, suggesting that metal accumulation in shoots is an energy‐demanding process. The analysis also showed that proteins involved in plant defense mechanisms were downregulated indicating that heavy metals accumulation in leaves supplies a protection system and highlights a cross‐talk between heavy metal signaling and defense signaling.     

Natural variation in cadmium tolerance and its relationship to metal hyperaccumulation for seven populations of Thlaspi caerulescens from western Europe

Thlaspi caerulescens J. & C. Presl is a distinctive metallophyte of central and western Europe that almost invariably hyperaccumulates Zn to> 1.0% of shoot dry biomass in its natural habitats, and can hyperaccumulate Ni to> 0.1% when growing on serpentine soils. Populations from the Ganges region of southern France also have a remarkable ability to accumulate Cd in their shoots to concentrations well in excess of 0.01% without apparent toxicity symptoms. Because hyperaccumulation of Cd appears to be highly variable in this species, the relationship between Cd tolerance and metal accumulation was investigated for seven contrasting populations of T. caerulescens grown under controlled conditions in solution culture. The populations varied considerably in average plant biomass (3.1‐fold), shoot : root ratio (2.2‐fold), Cd hyperaccumulation (3.5‐fold), shoot : root Cd‐concentration ratio (3.1‐fold), and shoot Cd : Zn ratio (2.6‐fold), but the degree of hyperaccumulation of Cd and Zn were strongly correlated. Two populations from the Ganges region were distinct in exhibiting high degrees of both Cd tolerance and hyperaccumulation (one requiring 3 µM Cd for optimal growth), whereas across the other five populations there was an inverse relationship between Cd tolerance and hyperaccumulation, as has been noted previously for Zn. Metal hyperaccumulation was negatively correlated with shoot : root ratio, which could account quantitatively for the differences between populations in shoot Zn (but not Cd) concentrations. On exposure to 30 µM Cd, the two Ganges populations showed marked reductions in shoot Zn and Fe concentrations, although Cd accumulation was not inhibited by elevated Zn; in the other five populations, 30 µM Cd had little or no effect on Zn or Fe accumulation but markedly reduced shoot Ca concentration. These results support a proposal that Cd is taken up predominantly via a high‐affinity uptake system for Fe in the Ganges populations, but via a lower‐affinity pathway for Ca in other populations. Total shoot Cd accumulated per plant was much more closely related to population Cd tolerance than Cd hyperaccumulation, indicating that metal tolerance may be the more important selection criterion in developing lines with greatest phytoremediation potential.     

Pyrene degradation and copper and zinc uptake by Fusarium solani and Hypocrea lixii isolated from petrol station soil

Aims: This study aimed to isolate and identify potential polycyclic aromatic hydrocarbon (PAH)‐degrading and/or metal‐tolerant fungi from PAH‐contaminated and metal‐contaminated soils. Methods and Results: Pyrene‐degrading fungi were isolated from contaminated soil and tested for metal (Cu, Zn and Pb) compound solubilization and metal accumulation. Three strains of Fusarium solani and one of Hypocrea lixii were able to degrade more than 60% of initial supplied pyrene (100 mg l^?1) after 2 weeks. The isolates were grown on toxic metal (Cu, Pb and Zn)‐containing media: all isolates accumulated Cu in their mycelia to values ranging from c. 5·9 to 10·4 mmol per kg dry weight biomass. The isolates were also able to accumulate Zn (c. 3·7–7·2 mmol per kg dry weight biomass) from zinc phosphate‐amended media. None of the isolates accumulated Pb. Conclusions: These fungal isolates appear to show promise for use in bioremediation of pyrene or related xenobiotics and removal of copper and zinc from wastes contaminated singly or in combination with these substances. Significance and Impact of the Study: Microbial responses to mixed organic and inorganic pollution are seldom considered: this research highlights the abilities of certain fungal strains to interact with both xenobiotics and toxic metals and is relevant to other studies on natural attenuation and bioremediation of polluted sites.     

Cadmium biosorption properties of the metal‐resistant Ochrobactrum cytisi Azn6.2

The aim of this work was to establish the conditions for using Ochrobactrum cytisi Azn6.2 as a metal biosorbent. Azn6.2 is a novel strain from the legume symbiont O. cytisi that has been isolated from nodules of Medicago polymorpha plants grown on heavy metal‐polluted soils. Compared with the strain ESC1, Azn6.2 showed some biochemical differences, as well as antibiotic susceptibility, Azn6.2 was multi‐resistant to heavy metals, such as Cu, Cd and Zn, and bacterial pellets were able to biosorb high amounts of Cd and Zn. As shown by scanning electron microscopy coupled to energy dispersive X‐ray, most of Cd was attached to the cell surface. Optimal conditions for Cd biosorption were established, being 1 mM Cd ions in solution and 2 h of contact with the biosorbent at room temperature. At these conditions, maximal Cd loading capacity reached 32–34 mg/g. Cd desorption from bacterial pellets was achieved after washing with EDTA or, at higher efficiency, at pH 1.0. These results indicated that biosorption/desorption on O. cytisi Azn6.2 biomass should be a cost‐effective method for Cd recovery from contaminated solutions.     

Determination of DNA methylation associated with Acer rubrum (red maple) adaptation to metals: analysis of global DNA modifications and methylation‐sensitive amplified polymorphism

Red maple (Acer rubum), a common deciduous tree species in Northern Ontario, has shown resistance to soil metal contamination. Previous reports have indicated that this plant does not accumulate metals in its tissue. However, low level of nickel and copper corresponding to the bioavailable levels in contaminated soils in Northern Ontario causes severe physiological damages. No differentiation between metal‐contaminated and uncontaminated populations has been reported based on genetic analyses. The main objective of this study was to assess whether DNA methylation is involved in A. rubrum adaptation to soil metal contamination. Global cytosine and methylation‐sensitive amplified polymorphism (MSAP) analyses were carried out in A. rubrum populations from metal‐contaminated and uncontaminated sites. The global modified cytosine ratios in genomic DNA revealed a significant decrease in cytosine methylation in genotypes from a metal‐contaminated site compared to uncontaminated populations. Other genotypes from a different metal‐contaminated site within the same region appear to be recalcitrant to metal‐induced DNA alterations even ≥30 years of tree life exposure to nickel and copper . MSAP analysis showed a high level of polymorphisms in both uncontaminated (77%) and metal‐contaminated (72%) populations. Overall, 205 CCGG loci were identified in which 127 were methylated in either outer or inner cytosine. No differentiation among populations was established based on several genetic parameters tested. The variations for nonmethylated and methylated loci were compared by analysis of molecular variance (AMOVA). For methylated loci, molecular variance among and within populations was 1.5% and 13.2%, respectively. These values were low (0.6% for among populations and 5.8% for within populations) for unmethylated loci. Metal contamination is seen to affect methylation of cytosine residues in CCGG motifs in the A. rubrum populations that were analyzed.