Genetic variability parameters of
1. Introduction
Metals such as cobalt (Co), copper (Cu), iron (Fe) and nickel (Ni) are essential for normal plant growth and development since they contribute to the function of many enzymes and proteins. However, metals can potentially become toxic to plants when they are present at high levels in their bioavailable forms (Hall, 2002). Phytotoxic levels of one or more inorganic ions in soil can be found in various parts of the world. These toxic sites occurred through natural processes or by anthropogenic effects. Naturally toxic soils include saline, acidic and serpentine soils, while anthropogenic polluted soils occur through mining activities, aerial fallout, and the run-off from galvanized sources of electricity pylons or motorway verges polluted by vehicle exhaust fumes (Bradshaw, 1984). The biochemical effect of metals on plants varies and the excess metal usually results in oxidative damage which affects their phenotype (Kachout et al., 2009)
Plants colonizing metal-contaminated soils are classified as resistant and have adapted to this stressed environment. Heavy metal resistance can be achieved by avoidance and/or tolerance. Avoiders are plants that are able to protect themselves by preventing metal ions from entering their cellular cytoplasm, while tolerant plants are able to detoxify metal ions that have crossed the plasma membrane or internal organelle biomembranes (Millaleo et al., 2010). Based on strategies used by plants growing on metal-contaminated soils, Baker and Walker (1990) classified them into three categories; metal excluder, indicators and accumulators/hyperaccumulators. The excluder group includes the majority of plant species that limit the translocation of heavy metals and maintain low levels of contaminants in their aerial tissues over an extensive range of soil concentrations. Plants that are metal indicators accumulate metals in their harvestable biomass and these levels generally are reflective of the metal concentration in the soil. Metal accumulators/hyperaccumulators are plants that increase internal sequestration, translocation and accumulation of metals in their harvestable biomass to levels that far exceed those found in the soil (Mganga et al., 2011; Baker and Walker 1990). Plants can accumulate and cope with the effects of high internal metal concentrations by the upregulation of the antioxidant defense system. This system is activated in order to respond to the deleterious effects caused by reactive oxygen species (Solanki and Dhankhar 2011).
Coping strategies allow the establishment of plant communities on metal contaminated soils. This is possible since some plants have adapted to these hostile sites by evolving mechanisms to deal with the toxic effects of metals in soil on plants. There is a need of identifying plants that are able to deal with excess metal in soil. Without these plants, the lands would remain barren and unsustainable.
The importance of plants in the remediation of heavy metal polluted soil is discussed in details in the present chapter. A review of the current knowledge on metal resistance mechanisms, as well as the potential genes and their role in metal homeostasis in plants will be examined. Finally, the coping mechanisms used by plants growing under metal contamination will be discussed.
2. Remediation of heavy metal contaminated sites
Soils that are heavily contaminated by metals may pose health risks to humans and to other living organisms in an ecosystem. Current techniques used to remediate metal contaminated soils include excavation, chemical stabilization, soil washing or soil flushing, but these methods are costly and impractical. There is a need to develop effective, low-cost and sustainable methods for soil bioremediation. The revegetation of these sites appears to be the most suitable method for long term land reclamation since plants can improve nutrient soil conditions. This can lead to the establishment of a self-sustaining vegetative cover, which in turn can prevent soil erosion (Wei et al., 2005). Phytoremediation is an inexpensive and solar-driven approach that is performed
Phytoremediation is composed of five main subgroups: phytoextraction, phytovolatilisation phytostabilization, phytodegradation and rhizofiltration. Phytoextraction is a process by which plants extract metals from soil by accumulating them in their aerial biomass. These plants can be harvested and metals can be extracted from their tissues. Plants that accumulate metals in their aerial tissues have been involved in the phytoextraction of several metals including Cd, Cr, Cu, Hg, Pb, Ni Se and Zn (Yong and Ma 2002).
The accumulation of metals by plants is interesting from an environmental or agronomic point of view. In mining or industrial sites, as well as their surrounding areas, heavy metals are responsible for severe soil contamination. In these cases, accumulator plants could be used for phytoremediation as they are likely able to remove metals from soils (Salt et al., 1998; Salt et al., 1995). Since some heavy metals are also essential minerals that can be deficient in staple food crops, genetic determinants of hyperaccumulation could be utilized in biofortification to improve the nutritional value of these crops (Frérot et al., 2010; Cakmak, 2008; Jeong and Guerinot 2008; Mayer et al., 2008).
These metal accumulator plants could also convert metals and release them in a volatile form. This process is known as phytovolatilization. Phytostabilization on the other hand, is a method that uses plants with a low ability for metal uptake to stabilize the contaminated soil thus preventing erosion. This limits the metals from entering the food chain. Plants can also be utilized for phytodegradation since they can in combination with microorganisms degrade organic pollutants. Finally, rhizofiltration is a process by which plant roots absorb metals from waste streams (Pulford and Watson 2003; Dushenkov et al., 1995).
3. Resistance
Levitt (1980) stated that heavy metal contaminated environments act as stress factors on plants, which causes physiological reaction change that reduces or inhibits plant vigor and growth. A plant showing injury or death due to metal stress is deemed sensitive to its environment. On the other hand, resistant plants can survive and reproduce under metal stress conditions (Ernst et al., 2008). In general, plants can achieve resistance to heavy metals by avoidance or tolerance.
3.1. Avoidance
Avoidance occurs when plants restrict the uptake of metals within root tissue by several strategies. In environments where the soil metal contamination is heterogeneously distributed, plants can prevent metal uptake by exploring less contaminated soil. Another avoidance strategy involves mycorrhizal fungi, where they can extend their hyphae outside the plants rooting zone up to several tens of meters and transfer the necessary elements to the plant (Ernst, 2006; Baker, 1987). Also, these metal tolerant fungi can increase plant metal resistance by changing the metals speciation or by restricting the metal transfer into the plant (Ernst, 2006). Arines et al. (1989) found that mycorrhizal
3.2. Tolerance
In the absence of avoidance strategies, some plants can grow and survive in soil contaminated with toxic levels of heavy metals which are otherwise lethal or detrimental for growth and survival of others genotypes of the same or of different species (Maestri and Marmiroli 2012). Plants exhibiting tolerance are internally protected from the stress of metals that have entered the cell’s cytoplasm (Baker, 1987). Metallophytes (metal tolerant plant) can function normally even in the presence of higher plant-internal metal levels. Plants adapt to their environments by developing heritable tolerance mechanisms. Tolerance to specific metals has evolved independently several times in different species from local non tolerant ancestral plant populations (Schat et al., 2000). Plants can exhibit tolerance to metals that are present in surplus in the soil. Each metal is under control of specific genes.
According to Bradshaw (1991) most species are in a state of genostasis. It is the restriction of genetic variability which limits the evolution of the population/species. In the absence of avoidance pathways, metal contaminated soil acts as a selection force on a population, where only the plants with tolerant genotype can survive and reproduce. This leads to a bottleneck, where few individuals survive and reproduce. In turn, metal tolerant populations can evolve rapidly following a disturbance such as contamination of soil with heavy metals. Plant adaptation to these sites occurs in populations for which tolerance variability already exists prior to the contamination (Maestri and Marmiroli 2012; Baker, 1987). Genes for the tolerance of metals are pre-existing at a low frequency in non tolerant populations of certain plant species (Ernst, 2006; Macnair, 1987).
3.2.1. Variation in tolerance and accumulation characteristics
Variation occurs between species, populations and clones for tolerance and accumulation of metals. Assunçãno et al. (2003) found differences in the degree of chlorosis and concentration of metal for
3.2.2. Genetics of tolerance to metals
Identifying genes involved in a specific adaptation is challenging. Metal tolerance and accumulation in plants are complex genetic systems. Plants have to modify their physiological processes in order to be able to survive in the environment in which they have germinated. In turn, the survival of a population to the contaminated environment is dependent on the inheritance of favourable traits. Tolerance mechanisms are heritable and variable, resulting from genes and gene products (Maestri and Marmiroli 2012). Variation in the evolution of metal tolerance exists over species, populations and clones (Baker, 1987). Some species do not show variation in tolerance and accumulations. In order to determine genes involved in metal tolerance and accumulation, segregating analyses were used, where parents with contrasting phenotypes were crossed to produce progeny. Studies have determined that in many species, metal tolerance and accumulation are genetically independent (Assunçãno et al., 2006). For example, in
Several techniques have been used to isolate and identify genes involved in heavy metal tolerance in plants, one of which is the quantitative trait loci (QTL) mapping. QTL mapping is a powerful tool in examining complex adaptive traits and in determining the number of genes involved in a trait as well as the genes effects and their interactions (Willems et al., 2007). By mapping QTLs, it can be possible to identify or validate candidate genes involved in a complex trait such as metal tolerance and accumulation (Willems et al., 2007). Other techniques used to identify genes for metal tolerance and accumulation are functional complementation in yeast mutants defective in metal homeostasis. These methods use plant cDNA expression libraries, as well as the identification of hypothesized pathways based on sequence similarities with plant cDNA libraries and genomic sequences (Lal, 2010). Transcriptome analyses have also been used to reveal genes involved in hyperaccumulation by analysing the differences in expression profiles or regulation-level of hyperaccumulator and non hyperaccumulator plants (Colzi et al., 2011).
Few specific major genes have been found for Cd, Cu, Ni and Zn tolerance in
Studies aiming at identifying associations between molecular markers and metal tolerance and accumulation trait have been performed using interspecific and intraspecific crosses. When a high Zn accumulating
Courbot et al. (2007), also using progeny from interspecific cross between
Using segregating progeny resulting from intraspecific crosses between a high Cd accumulating parent and a low Cd accumulating parent for
Several QTL studies on rice (
In durum wheat (
In wheat (
3.3. Categories of plants growing on metal contaminated soils
Baker and Walker (1990) categorized plants into three groups according to their strategy for coping with metal toxicity in soil; metal excluders, indicators and accumulators/ hyperaccumulators.
3.3.1. Excluders
The metal excluder strategy consist in limiting the amount of metals translocated from roots to shoots thus maintaining low levels of metal concentration in their aerial parts. Large amounts of metals in the roots of excluder species have been reported (Baker and Walker 1990). Examples of excluder species include
3.3.2. Accumulators/hyperaccumulators
Metal accumulators/hyperaccumulators are plants that can concentrate metals in their above-ground tissues to levels that exceed those in the soil or also to those in the non accumulating species found growing nearby with concentrations up to 100 times more than non hyperaccumulators (Salt et al., 1998). Accumulators/hyperaccumulators growing on metal contaminated environments can naturally accumulate higher levels of heavy metals in their shoots than in their roots (Kachout et al., 2009). Some plants can accumulate only a specific metal while others can accumulate multiple metals ((Mganga et al., 2011; Almås et al., 2009). Presently, at least 45 plant families comprising more than 400 species have been found to accumulate metals in their harvestable tissues, and the majority of them belong to the
3.3.3. Indicators
Like accumulators, metal indicators accumulate metals in their aerial tissue, but the metal levels in the above ground tissue of these plants usually reflect the metal concentration in the surrounding environment (Baker and Walker 1990). If these plants continue to uptake metals, they will eventually die-off. These plants are of biological and ecological importance since they are pollution indicators and also, like accumulators, they absorb pollutants (Mganga et al., 2011).
3.3.4. Determination of excluders, indicators and hyperaccumulators plants
A plant is classified as a hyperaccumulator when it meets four criteria including; a) when the level of heavy metal in the shoot divided by level of heavy metal in the root is greater than 1 (shoot/root quotient > 1); b) when the level of heavy metal in the shoot divided by total level of heavy metal in the soil is greater than 1 (extraction coefficient > 1) (Rotkittikhun et al., 2006; Harrison and Chirgawi 1989); c) when the plant takes up between 10 – 500 times more heavy metals than normal plants (uncontaminated plants - control plants) (Fifield and Haines 2000; Allen,1989); and d) more than 100mg/kg of cadmium, 1000g/kg of copper, lead, nickel, chromium; or more than 10000mg/kg of zinc (Mganga et al., 2011; Ernst, 2006; Brooks, 1998). An excluder is a plant that has high levels of heavy metals in the roots but with shoot/root quotients less than 1 (Boularbah et al., 2006). Finally, Baker and Walker (1990) classified a plant as an indicator when the levels of heavy metals within their tissues reflect those in the surrounding soil.
3.4. Physiological mechanisms of metal resistance
Resistant plants are able to grow on metal contaminated soil due to avoidance and/or tolerance strategies. Plant resistance to high levels of heavy metals in soils can result from either reduced uptake or once taken up, metals have to be transformed into a physiologically tolerable form.
3.4.1. Restriction of metal uptake
The plasma membrane is the first structure of living cells exposed to heavy metals. The membrane functions as a barrier for the movement of heavy metals into cytoplasm. The restriction of metals at the plasma membrane limits the uptake and accumulation of metals by preventing their entry into the cytoplasm. This can be done by changing the ion binding capacity of the cell wall and/or decreasing the uptake of metal ions through modified ion channels, and/or by removing metals from cells with active efflux pumps and/or with root with root exudates (Tong et al., 2004).
3.4.1.1. The cell wall
The cell wall and membrane interface could be a site of metal tolerance since a significant amount of metals has been reported to be accumulated there. Divalent and trivalent metal cations can bind plant cell walls because of the presence of functional groups such as –COOH, -OH and –SH. Pectins are polymers that contain carboxyl groups which enable the binding of divalent and trivalent heavy metals ions. In enriched heavy metal environments, some plants will increase the capacity of their cell wall to bind metals by increasing polysaccharides, such as pectins (Colzi et al., 2011; Pelloux et al., 2007). Konno et al. (2010; 2005) showed that the pectin in root cell walls was important in binding Cu in the fern,
3.4.1.2. Root exudates
Resistant plants can also restrict the entry of metals by immobilizing them in the rhizosphere with root exudates outside the plasma membrane (Colzi et al., 2011). This has been reported in
3.4.2. Chelation
The phytotoxic effect of free metal ions can be eliminated by their chelation by specific high-affinity ligands (Yong and Ma 2002). The chelation of metals allows for the restriction of metal uptake, the uptake of metal ions, sequestration and compartmentation, as well as xylem loading and transport within the plant. Baker et al. (2000) categorized these ligands according to the characteristic electron donor centers, which include sulfur donor ligands, oxygen donor ligands and nitrogen donor ligands.
3.4.2.1. Oxygen donor ligands
Organic acids such as malate, aconitate, malonate, oxalate, tartrate and citrate are involved in metal uptake restriction and dexotification in plants. These carboxylic acid anions form complexes with divalent and trivalent metal ions with high stability. They are involved in the restriction of metal entry into the cell, metal exclusion in the root cells, accumulation and transport within the plants. In wheat (
3.4.2.2. Nitrogen donor ligands
This group consists of amino acids and their derivatives which have relatively high affinity for specific metals. Krämer et al., (1996) revealed histidine to be involved in the Ni tolerance and translocation of the hyperaccumulator plant
3.4.2.3. Sulfur donor ligands
In plants, sulfur donor ligands are composed of two classes of metal chelating ligands which are phytochelatins (PCs) and metallothioneins (MTs). Phytochelatins are small metal binding peptides synthesized from the tripeptide glutathione (γ-Glu-Cys)2-11-Gly) (Solanki and Dhankhar 2011; Hall, 2002). Since there is a γ-carboxamide linkage between glutamate and cysteine, PCs are not synthesized by translation of mRNA, but rather it is a product of an enzymatic reaction involving the enzyme PC synthase (Yong and Ma 2002). The production of PCs is positively correlated with metal accumulation in plant tissues (Pal and Rai 2010). PCs are produced in cells immediately after heavy metal exposure, including Cd, Pb, Zn, Ag, Hg, As and Cu as seen in
Several research groups concurrently and independently cloned and characterized genes encoding PC synthase. These genes were isolated from
The second class of sulfur donor ligands are metallotioneins (MTs). They are low molecular weight (4-14kDa), cysteine-rich, metal-binding proteins found in a wide range of organisms (animals, plants, eukaryotic microorganisms, and prokaryotes) (Huang and Wang 2010). Unlike PCs, they are encoded by structural genes (Yong and Ma 2002). They play essential roles in a variety of organisms including Cu, Cd and Hg detoxification by sequestration (Palmiter, 1998; Ecker et al., 1989), Zn homeostasis (Coyle et al., 2002) and also scavenging of reactive oxygen species (Wong et al., 2004). MTs have been divided into two classes based on their cysteine residue arrangements. Class I MTs are widespread in vertebrates and are composed of 20 highly conserved cysteine residues based on mammalian MTs. Class II MTs have slightly flexible cysteine arrangements and are found in plants, fungi and invertebrates. A third class includes phytochelatins (Chaturvedi et al., 2012). Based on the position and allocation of cysteine residues, class II plant MTs are additionally divided into four types (Cobbett and Goldsbrough 2002). Type 1 plant MT genes have been more highly expressed in roots compared to leaves while the reverse is observed for the expression of type 2 plant MT genes. Type 3 MT genes are highly expressed in ripening fruits or in leaves while the expression of type 4 plant MT gene is restricted to developing seeds (Sekhar et al., 2011; Cobbett and Goldsbrough 2002).
The expression of MT genes in plants subjected to metal stress has been studied.
In general, there are variations between species in the expression of MTs to various metals. The up and down regulation of MTs in response to metal stress is largely unknown in plants. The MT gene expression was shown to be strongly induced by Cu, Cd, Pb and Zn (Huang and Wang 2009; Gonzalez-Mendoza et al., 2007; van Hoof et al., 2001). MT gene expression is also influenced by other abiotic stressors including absciscic acid (ABA), drought, salinity, heat, cold light, wounding and senescence (Sekhar et al., 2011).
3.4.3. Mechanisms involved in internal metal tolerance
3.4.3.1. Metal uptake
The uptake of metal from soil into roots is dependent on the bioavailabilty of the metal, as well as its mobility in the rhizosphere (Maestri et al., 2010). The bioavailability of various metals greatly varies. No correlation exists between the metal content in soils and in plants (Clemens, 2006). The bioavailability of metals in the rhizosphere is affected by the chemical environment. For example, in
3.4.3.2. Metal uptake across the plasma membrane
The uptake of heavy metals in plants is mediated by a group of metal transporter families which consists of iron-responsive transport proteins (ZIP-IRT), the heavy metal-transporting P1B-type subfamily of P-type ATPases, the natural resistance associated macrophage proteins (NRAMP) and the cation diffusion facilitators (CDF) (Baxter et al., 2003). Transporters were originally identified for Fe2+ or Zn2+ homeostasis, but it was demonstrated that most transporters of essential metal ions can also carry non essential metals, such as Cd (Zhou et al., 2012). The uptake of non essential metals may be the result of their close chemical characteristics or metal ion size to essential metals. Some metal transporters, present in the plasma membrane of root cells, exhibit low substrate specificity which can lead to the accumulation of other metals in plants (Schaaf et al., 2006). For example, the non-functional metal Cd can be taken up via a Ca2+ transporter (Perfus-Barbeoch et al., 2002) or also via the Fe2+ transporter IRT1 (Korshunova et al., 1999). Plant tolerance to metal stress can be achieved with the modification of these transporter activities (Zhou et al., 2012). Plants can prevent the uptake of certain metals by down-regulating the expression of such transporters, as observed in
Increased Zn uptake is driven by an overexpression of members of the ZIP family of transporters. Under Zn deficiency conditions, many members of the ZIP transporter family are overexpressed in non hyperaccumulator species, while in hyperaccumulators, they are independently expressed regardless of Zn supply (Verbruggen et al., 2009). Nishida et al. (2011) and Schaaf et al. (2006) showed that
The uptake of Ni of some Ni hyperaccumulator accessions of
Heavy metal-transporting P1B-type transporters are also involved in metal-ion homeostasis and tolerance in plants by transporting essential and non essential heavy metals such as Cu, Zn, Cd, Pb across cell membrane. Transporters located at the plasma membrane function as efflux pumps by removing toxic metals form cytoplasm. They have also been found in membranes of intracellular organelles for compartmentalization of metals for sequestration in vacuoles, golgi or endoplasmic reticulum (Yang et al., 2005b). These ion pumps transport ions across a membrane by hydrolysing ATP (Benitez et al., 2012). Eight P1B-ATPases, AtHMA1–AtHMA8, have been reported in
In addition, the gene
In bacteria and in some eukaryotes, Zn, Co and Cd are transported by the CDF transport proteins. Within the
Plants can make metal ions more available for uptake by acidifying the rhizosphere and pumping protons via plasma membrane-localized proton pumps; and also by exuding low molecular weight (LMW) compounds that act as metal chelators (Clemens, 2006). The secretion of organic acids can render heavy metals mobile and enhance their absorption by plant roots. Krishnamurti et al. (1997) reported that when Cd was complexed with organic acids, it was readily available for transport across the membrane, while free Cd ions were restricted for uptake. Cieśliński et al. (1998) revealed a higher acetic acid and succinate in the rhizosphere of the
3.4.3.3. Sequestration/compartmentation
Some metal tolerant plants can accumulate large amounts of metals within the cell without exhibiting toxicity symptoms (Entry et al., 1999). These plants are able to store the surplus of accumulated metals where no sensitive metabolic activities occur such as organs or subcellular compartments (Ernst, 2006). This avoidance of metal poisoning involves the intracellular sequestration and apoplastic or vacuolar compartmentation of the toxic metal ions (Liu et al., 2007). Compartmentation of metals can also be found in the cells central vacuole. This was observed in the Zn resistant
Schaaf et al. (2006) determined that the transporter AtIREG2, located at the tonoplast, was involved in Ni detoxification in roots. AtREG2, confined to roots, prevents heavy metal translocation to shoots restricting metals to roots. This transporter counterbalances the low substrate specificity of transporter AtIRT1 and other iron transporters in iron deficient root cells
Members of the CDF protein play a role in tolerance to various metals including Cd, Co, Mn, Ni and Zn by their sequestration into vacuoles (Montanini et al., 2007). Increased Zn tolerance and accumulation was reported in non accumulator
Phytochelatins are also thought to be involved in the restriction of metals to the roots (Zenk, 1996). When
3.4.3.4. Root to shoot translocation
The translocation of metals to the aerial biomass can be an important biochemical process used by plants to remediate polluted areas. In some plants, the mobilization of metals from their roots to their above aerial organs can minimize the damage that could be exerted by these heavy metals on the root physiology and biochemistry (Zacchini et al., 2009). Excluders prevent or limit the translocation of toxic metals or essential metals from roots to shoots. On the other hand, accumulators/hyperaccumulators translocate metals from roots to shoots via the xylem with the transpiration stream. This is accomplished by increasing the uptake of metals in roots, and by reducing the sequestration of metals in the root.
The chelation of metals with ligands, such as organic acids, amino acids and thiols facilitates the movements of heavy metals from roots to shoots (Zacchini et al., 2009). The xylem cell wall has a high cation exchange capability, thus the movement of metal cations is severely retarded when the metals are not chelated by ligands. Organic acids are involved in the translocation of Cd in the species
The chelation of Ni to histidine is involved in the long distance translocation of Ni in the hyperaccumulator
The chelation of metals with nicotianamine (NA) also contributes to improved tolerance. Nicotianamine can chelate and transport divalent Ni, Cu and Zn (Takahashi et al., 2003; Pich et al., 2001; Ling et al., 1999). The nicotianamine synthase (NAS) enzyme is responsible for the synthesis of NA by trimerization of S-adenosylmethionine (Shojima et al., 1990). When exposed to high levels of Zn, Cd, and/or Ni, all four
Visioli et al. (2010) also showed that metallothioneins may be involved in the translocation of Ni in
Transporters are not only involved in the uptake of metals from the soil, but also in their transport out of the vacuole. These mobilized metals can then be translocated to aerial tissue. Visioli et
al. (2012) subsequently found for sub-population MP2p, which exhibited the highest level of Ni translocation of the four sub-populations analyzed, significantly higher levels of the ABC27 transporter. This transporter is part of the ABC family of transporters which are involved in removing metals from the cytoplasm by pumping outside the cell wall, metals sequestered in vacuoles and other subcellular compartments (Visioli et al., 2012; Martinoia et al., 2002; Sanchez-Fernandez et al., 2001). Hassinen et al. (2007) showed that the AtMRP10 homolog, also part of the ABC family of transporters, had different expression in roots of two
The passage of metal ions and/or metal ligand complexes from the cytosol of root cells into the vascular tissue requires their transport across the cell membrane. Transporters involved in this activity are the heavy metal transporting P-type ATPases (HMAs) (Clemens, 2006). The AtHMA2 and 4 are involved in translocation of Zn in
3.4.3.5. Metal storage
Metals have to undergo a xylem unloading process prior to their distribution and their detoxification in the shoot and their redistribution via the phloem (Schmidke and Stephan 1995). Once unloaded, the metals are either taken up into surrounding cells and are symplastically transported through the leaf tissues or they are apoplastically distributed over the leaf (Hassan and Aarts 2011; Marschner, 1995). NA is important in the chelation of metals for their symplastic transport through the leaf. This occurs through the Yellow Stripe Like proteins (YLS) (Hassan and Aarts 2011; DiDonato et al., 2004). In the hyperaccumulator
The sequestration of excess essential and non essential metals is localized in various parts of the aerial tissue, such as trichomes, leaf epidermal cell vacuole and mesophyll vacuole. Broadhurst et al. (2004) grew five
3.4.4. Antioxidative defence involved in metal tolerance
In environments, where metals are present in toxic levels, the elevated activities of antioxidant enzymes and non-enzymatic constituents are important in the plant tolerance to stress. Metal tolerance may be enhanced by the plant’s antioxidant resistant mechanisms. There is an indication that the alleviation of oxidative damage and increased resistance to stresses in the environment is often correlated with an effective antioxidative system. The minimization of damage due to oxidative stress is a universal feature of plants defense responses (Kachout et al., 2009). The detrimental effect of heavy metals in plants is due to the production of ROS and induction of oxidative stress. Oxidative stress is expressed by the increase levels of reactive oxygen species such as singlet oxygen (1O2), superoxide radical (O
Oxidative damage could result when the balance between the detoxification of the ROS products and the antioxidative system is altered (Kachout et al., 2009). The tolerance of deleterious environmental stresses, such as heavy metals, is correlated with the increased capacity to scavenge or detoxify activated oxygen species (Kachout et al., 2009). Boominathan and Doran (2003a,b) determined the role of antioxidative metabolism of heavy metal tolerance in
3.5. Problems associated with plant metal tolerance
Soils enriched with metals are demanding on tolerant and accumulator plants. The costs associated with their adaptation to these sites are related to energy and resources allocations. When a metal tolerant or accumulator plant is growing in a metal contaminated soil, there is an increase in cost because the organism has to spend energy to counter the effects of the metals (Maestri et al., 2010). Slow growth and low reproduction are the main characteristics of plants growing on metal enriched soils (Ernst, 2006; Ernst et al., 2000). Haldane (1954) stated that costs are associated with the natural selection of new alleles. More energy and resources are required for the maintenance of the tolerance mechanisms at the cellular level. It has been demonstrated that tolerant plants have increased synthesis of complexing molecules in the cytosol. For example, metallothioneins and phytochelatins for the detoxification of metals such as As, Cd, and Cu. ATP are also needed for the active transport of metals across the plasma membrane and tonoplast. The synthesis of these agents withdraws N, S and energy from the primary metabolism (Ernst, 2006; Verkleij et al., 1998). Energy is also required for the translocation of metals from root to shoot as well as for their allocation to various tissues and cell types. The reduced biomass of metal tolerant plants compared to their non metal tolerant ancestors might also be the result of less than favourable environmental conditions such as low water and nutrient supply. The diminished biomass and seed production might be the result of all costs associated with their survival to these metal contaminated sites, such as adaptation and environmental constraints (Ernst, 2006). Plants have an advantages growing on metal contaminated soil. As previously mentioned, there is a lack of competitive species on these sites. With high metal accumulation of metals in their aerial tissues, the “elemental hypothesis” speculates that hyperaccumulators can deter predators such as herbivores from feeding on them (Maestri et al., 2010; Vesk and Reichman 2009). However, some insects feed on hyperaccumulator plants and in turn accumulate the metals in their tissue which then aid in their defence against predators (Maestri et al., 2010). This contradiction may explain why there is a mix of excluders, accumulator and hyperaccumulators growing on metal contaminated sites. Another advantage of hyperaccumulation is the elimination of competitive plants by further contaminating the surrounding soil by shedding their metal contaminated leaves (Maestri et al., 2010).
3.6. Effects of metals on plant population diversity and structure
Elevated accumulations of metals in soil and vegetation have been documented within short distances of the smelters compared to control sites (Nkongolo et al., 2008; Gratton et al., 2000). Several authors have reported differences in genetic structure of plants growing in contaminated areas (Vandeligt et al., 2011; Nkongolo et al., 2008; Scholz and Bergmann 1984). Enzymatic studies of Norway spruce (
Molecular analyses of several conifer and hardwood species clearly indicated that the exposure to metals for more than 30 years has no effect on genetic structure and diversity of early generations of
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Vale site 1 (metal contaminated) | 31.25 | 0.1120 | 0.1653 | 1.2035 | 1.3125 |
Vale site 2 (metal contaminated | 31.25 | 0.1171 | 0.1727 | 1.2061 | 1.3125 |
Xtrata 2 (metal contaminated) | 27.08 | 0.0995 | 0.1467 | 1.1758 | 1.2708 |
Xtrata 3 (metal contaminated) | 20.83 | 0.0630 | 0.0982 | 1.1004 | 1.2083 |
Vale Tailing (metal contaminated) | 35.42 | 0.0977 | 0.1552 | 1.1514 | 1.3542 |
Temagami site (control) | 29.17 | 0.0818 | 0.1284 | 1.1310 | 1.2917 |
Low Water Lake (control) | 31.25 | 0.0812 | 0.1297 | 1.1256 | 1.3125 |
Mean | 31.63 | 0.1001 | 0.1528 | 1.1679 | 1.3163 |
P represents percentage of polymorphic loci; h, Nei’s gene diversity; I, Shannon’s information index; Ne, effective number of alleles; Na, observed number of alleles.
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Sudbury | Coniston | 72 | 74 (Sudbury) |
(moderately contaminated) | Xtrata | 92 | |
Copper Cliff | 67 | ||
Walden | 65 |
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Cobalt | Cobalt-3 | 48 | 46 (Cobalt) |
(highly contaminated) | Cobalt-4 | 46 | |
Cobalt-5 | 44 |
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Manitoulin | Little Current | 70 | 69 (Manitoulin) |
(control) | Mississagi Lighthouse | 68 |
P represents percentage of polymorphic loci; Sudbury and Cobalt regions were moderately and highly contaminated with metals, respectively. Manitoulin Island region was not contaminated with metals and was used as a control region.
4. Conclusion
Plants play an essential role in the remediation of metal enriched soils. Coping mechanisms developed by some group of plants growing on metal contaminated soil facilitate the establishment of sustainable ecosystems in areas that would otherwise remain barren. A number of studies have been completed to explain the complex mechanisms involved in tolerance genotypes, and also the biological variability in their environmental adaptation. Depending on the circumstances, metal excluders or hyperaccumulators may be used to remediate polluted soil. Excluders may be useful for soil stabilization by preventing wind and water erosion and also by limiting the entry of heavy metals in the food chain. Metals can also be extracted by hyperaccumulators but since majority of these plants have low biomass, the extraction of metals from soil is very slow. The remediation of these sites using this technique may take up to hundreds of years. With genetic engineering, it may be possible to design the ideal plant prototype for the remediation of metal contamination in different environments. Many genes and mechanisms have been identified to have a role in tolerance and hyperaccumulation of metals. However, there is still a need for a better understanding of the mechanisms such as characterization of promoters of genes controlling metal tolerance and hyperaccumulation. This new knowledge would significantly contribute to a better understanding of the regulation and expression of different genes in hyperaccumulators. It is essential to mimic this regulation and expression of genes in high biomass non hyperaccumulators in order to obtain the hyperaccumulator phenotype.
Acknowledgments
We express our appreciation to the Natural Sciences and Engineering Research Council of Canada (NSERC) for a Postdoctoral Fellowship to M. Mehes-Smith.References
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