Keywords

1 Introduction

During the past five decades, the role of fertilizer in enhancing food grain production has been widely recognized in both the developed and developing countries. It has been highlighted that fertilizer is the kingpin of the green revolution and the best hope for meeting the food security challenges in future. Despite considerable progress made in the field of fertilizer use research, the recovery efficiency of applied fertilizer nutrients hardly exceeds 50 % and considerable amount is lost from the soil system. Though the consumption of chemical fertilizers in India increased steadily over the years, the use efficiency of nutrients applied through fertilizers continues to remain low (in the range of 30–50 % for N, 20 % for P, 55 % for K, and 2–5 % for Zn, Fe, and Cu) owing to nutrient losses from soils or conversions of nutrients into slowly cycling/recalcitrant pools within soil. Improvement in fertilizer use efficiency, therefore, is necessary to increase crop productivity and reduce environmental pollution. The nutrient use efficiency may be improved by regulating the supply from the fertilizer material and enhancing the uptake and utilization efficiencies by the plant. Efficient use of nutrients in agriculture may be defined differently when viewed from agronomic, economic, or environmental perspectives. Proper definition for the intended use is essential to understand published values and have meaningful discussion. It is a fallacy that the highest possible nutrient efficiencies should be the ultimate goal of fertilizer users. The highest “efficiency” occurs when small amounts of nutrients are applied in deficient soils. While efficiency may be very high in this condition, crop growth in this region is generally stunted and profitability is low, compared with the situation where balanced and appropriate nutrition is provided. Another example of inadequate understanding of “efficiency” is when an insufficient quantity of nutrients is regularly added to meet crop needs. In this condition, soil productivity will gradually decline as crop production continues to be increasingly reliant on nutrient stocks from soil reserves. Nominal nutrient use efficiency may be very high under these circumstances, but it is clearly a non-sustainable scenario.

2 Nutrient Use Efficiency

Nutrient efficiency can be classified into three groups, viz., agronomic efficiency, physiological efficiency, and apparent recovery efficiency. The agronomic efficiency is defined as this economic production obtained per unit of nutrient applied. It can be calculated with the help of following equation:

$$ \mathrm{Agronomic}\;\mathrm{efficiency}=\frac{\mathrm{Grain}\;\mathrm{yield}\;\mathrm{of}\;\mathrm{fertilized}\;\mathrm{crop}\;\mathrm{in}\;\mathrm{kg}-\mathrm{Grain}\;\mathrm{yield}\;\mathrm{of}\;\mathrm{unfertilized}\;\mathrm{crop}\;\mathrm{in}\;\mathrm{kg}}{\mathrm{Quantity}\;\mathrm{of}\;\mathrm{fertilizer}\;\mathrm{applied}\;\mathrm{in}\;\mathrm{kg}}=\mathrm{kg}\;{\mathrm{kg}}^{-1} $$

Sometime, agronomic efficiency is also called economic efficiency. If the efficiency is determined under greenhouse conditions, the agronomic efficiency may be expressed in g g−1 or mg mg−1.

Physiological efficiency is defined as the biological production obtained per unit of nutrient observed. Sometimes, it is also known as biological efficiency or efficiency ratio. It can be calculated with the help of the following equation:

$$ \mathrm{Physiological}\;\mathrm{efficiency}=\frac{\mathrm{Total}\ \mathrm{dry}\ \mathrm{matter}\ \mathrm{y}\mathrm{ield}\ \mathrm{of}\;\mathrm{fertilized}\ \mathrm{crop}\ \mathrm{in}\ \mathrm{kg}-\mathrm{Total}\ \mathrm{dry}\ \mathrm{matter}\ \mathrm{y}\mathrm{ield}\ \mathrm{of}\;\mathrm{unfertilized}\ \mathrm{crop}\ \mathrm{in}\ \mathrm{kg}}{\mathrm{Nutrient}\ \mathrm{uptake}\ \mathrm{b}\mathrm{y}\;\mathrm{fertilized}\ \mathrm{crop}\ \mathrm{in}\ \mathrm{kg}\kern2em \mathrm{Nutrient}\ \mathrm{uptake}\ \mathrm{b}\mathrm{y}\;\mathrm{unfertilized}\ \mathrm{crop}\ \mathrm{in}\ \mathrm{kg}}=\mathrm{kg}\;{\mathrm{kg}}^{-1} $$

The apparent recovery efficiency is defined as the quantity of nutrient absorbed per unit of nutrient applied. It can be calculated with the help of the following expression:

$$ \mathrm{Apparent}\ \mathrm{recovery}\;\mathrm{efficiency}=\frac{\mathrm{Nutrient}\ \mathrm{uptake}\ \mathrm{b}\mathrm{y}\;\mathrm{fertilized}\ \mathrm{crop}-\mathrm{Nutrient}\ \mathrm{uptake}\ \mathrm{b}\mathrm{y}\;\mathrm{unfertilized}\ \mathrm{crop}}{\mathrm{Quantity}\ \mathrm{of}\ \mathrm{fertilizer}\ \mathrm{applied}}\times 100\% $$

Physiological and recovery efficiency can be combined to obtain the nutrient use efficiency: Nutrient use efficiency = physiological efficiency × recovery efficiency

In addition to agricultural intensification on the best arable land, management practices need to be identified for rational utilization of marginal lands for agriculture for enhancing sustainable crop production in developing countries (Lal 2000). In these context possibilities of harnessing newly emerging concepts, mechanisms, and techniques in cellular and molecular biology need to be explored for better understanding of tolerance mechanisms to stress so that appropriate strategies could be developed for identification of crop genotypes with superior resource use efficiency. Integration of such crops into crop rotations in conjunction with improved management practices will play an increasingly important role in enhancing crop production especially under conditions of low availability of nutrients and other stresses. Significant progress has been made in understanding the physiological traits of several crop species responsible for tolerance to a wide range of nutrient deficiencies in soils. Preliminary studies have shown that inclusion of such promising crop species into crop rotations or to intercropping would be a useful strategy to improve nutrient use efficiency, crop nutrition, and yields (Hocking 2001).

2.1 What Is Phytometallophore?

In response to iron-deficiency stress conditions, roots of cereals (Poaceae family) release non-proteinogenic amino acids, called phytometallophores or phytosiderophores that solubilize and chelate inorganic Fe. In cereal, Fe (III) is taken up in toto as the Fe (III)-phytometallophore complex. Apparently, there are specific recognition sites on the plasma membrane of the root cells which allow the binding and transport of the metal-phytometallophore (PM) complex across the plasma membrane and into the cystosol. Absorption across the plasma membrane is thought to be via an amino acid cotransport system. In addition to their role in Fe acquisition, it has been hypothesized that PM has a universal role for acquisition of other trace metals such as Zn, Mn, and Cu that also have low solubilities in alkaline soils. Recent study reported that Zn (II)-PM complexes (i.e., 2-deoxymugineic acid, epi-hydroxymugineic acid, and mugineic acid) were readily absorbed by corn roots. These results suggest that corn root-cell plasma membrane binding sites are not highly specific for Fe (III)-phytometallophore, allowing the transport of other transition metals into cell. PM release may vary with the physiological status and age of the plant and with severity of the deficiency. PM release rates generally increase at first and afterwards decrease with plant age. Photoperiod and temperature also directly influenced by diurnal release patterns. PM release rates and their recovery may be influenced by nonspecific root-microbial interaction and by the physical environment of the roots such as aeration or root contact that may influence root morphology and exudation rates.

2.2 Biosynthesis of PM

In graminaceous species (strategy II), these iron-deficiency-induced morphological and physiological changes are absent. Instead, roots release phytosiderophores (PS) are chelators for Fe (III). The pathway of PS biosynthesis is understood reasonably well. L-Methionine is the dominant precursor, and three molecules of it are used to form one molecule of nicotianamine which, after deamination and hydroxylation, is converted into 2-deoxymugineic acid and further to PS, depending upon plant species. Nicotianamine (NA) is not only a precursor of PS biosynthesis but is also a strong chelator of Fe (II), but not of Fe (III). It is also essential for the proper functioning of Fe (II)-dependent processes. Nicotianamine might be the link between the two strategies of iron-deficiency-induced root responses, perhaps reflecting differences in codon usage in genes of dicots in comparison with monocots. From DMA, the biosynthesis pathway may diverge in different plant species, resulting in different PS being exuded into the rhizosphore of different species. The PS is synthesized mainly in root tips, even though biosynthesis may occur in the meristematic tissue of the shoot as well. In either case, PS are synthesized continuously and are stored in roots for release into the rhizosphore during a defined period of the day. An increased exudation of PS under Fe deficiency occurs in a distinct diurnal rhythm, with a peak exudation after the onset of illumination, the light ensuring the continuous supply of assimilates from photosynthetically active plant parts. More detailed studies on the diurnal rhythm revealed that it is an increase in temperature during the light period, rather than the onset of light itself, which causes an increase in PS exudation.

2.3 Strategy I and Strategy II Plants

Classical studies of Fe nutrition in plants have resulted in the division of plants into various strategies.

Strategy I, dicotyledonous plants respond to iron stress by inducing a cell surface reduction system and in some instances can be accompanied by the release of protons and reductants that alter chemical conditions in the rhizosphere to increase inorganic Fe solubility. Strategy I is typically for dicots and non-graminaceous monocots and characterized by at least two distinct components of iron-deficiency response, increased reducing capacity, and enhanced net excretion of protons. In many instances also the release is enhanced of reducing and/or chelating compounds, mainly phenolics. These root responses are often related to changes in root morphology and anatomy, particularly in the formation of transfer cell-like structures in rhizodermal cells. The most sensitive and typical response is the increase in activity of a plasma membrane-bound reductase in the rhizodermal cells. The supposed existence of two reductases, a constitutive (basic) reductase with low capacity, and an iron-deficiency-induced high capacity reductase increases the activity Marschner et al. (1989). Although the transmembrane redox pump may contribute to the net excretion of protons, the strongly enhanced net excretion of protons under iron deficiency is most probably the result of higher activity of the plasma membrane proton efflux pump and not of the reductase. The activity of the reductase (R) which is strongly stimulated by low pH, i.e., enhanced proton excretion by the ATPase, is important for the efficiency in Fe (III) reduction. Accordingly, high concentrations of HCO−3 counteract this response system in strategy I plants.

Strategy II, monocotyledonous grasses, responds to Fe stress by production of Ps that are secreted into the rhizosphere and which are subsequently transported by a specific uptake system on the root surface. Strategy II is confined to graminaceous plant species and characterized by an iron-deficiency-induced enhanced release of non-proteinogenic amino acids, so-called phytosiderophores. The release follows a distinct diurnal rhythm and is rapidly depressed by resupply of iron. The diurnal rhythm in release of PS in iron-deficient plants is inversely related with the volume of a particular type of vesicles in the cytoplasm of root cortical cells. PS such as mugineic acid form highly stable complexes with Fe (III); the stability constant in water is in the order of 1023. As a second component of strategy II, a highly specific constitutive transport system translocator is present in the plasma membrane of root cells of grasses which transfers the Fe (III) PS into the cytoplasm. In plant species with strategy I, this transport system is also lacking. Although PS form complexes also with other heavy metals such as Zn, Cu, and Mn, the translocator in the plasma membrane has only a low affinity to the corresponding complexes. Nevertheless, release of PS may indirectly enhance the uptake rate of these other metals by increasing their mobility in the rhizosphere and in the root apoplasm. Under iron deficiency not only the release of phytosiderophores is increased but also the uptake rate of the Fe (III) PS complexes indicating a higher transport capacity due either to an increase in number of the turnover rate of the translocator. Although this PS system resembles features of the siderophore system in microorganism, its affinity to PS is two to three orders of magnitude higher than for siderophores such as ferrioxamine B or for synthetic iron chelates such as FEEDDHA.

3 Methodology

A rapid, simple, and accurate method of determining concentrations of Fe-chelating agents in solution was developed Shenker et al. (1995). The assay employs Cu-CAS (chrome azurol S) complex as a testing agent and measures the equilibrium concentration of this complex in the presence of other Fe chelators. This method is of particular importance for colorless chelates, which are very difficult to determine otherwise.

The Cu-CAS assay—a proposed protocol to quantify unknown chelate concentration.

  1. 1.

    Cu-CAS reagent preparation:

Prepare a stock solution of the Cu-CAS reagent consisting of: 200 μM CuCl2, 210 μM CAS, 40 mM MES, pH adjusted to 5.7 with NaOH. This solution is stable for a long period (months).

  1. 2.

    Sampling of unknown chelate solution:

Prepare a series of six 1.5 mL microtubes for each chelate solution to be assayed. Add an accurate volume containing approximately 140 nmol of the unknown chelate to the first microtube and add distilled water to a final volume of 1.4 mL.

  1. 3.

    Preparation for dilution step:

Add 700 μL of distilled water to each of the other microtubes.

  1. 4.

    Constructing a 1:1 serial dilution of the tested chelate:

Prepare 1:1 serial dilution by transferring 700 μL from each microtube to the next. Ensure good mixing after each dilution step.

  1. 5.

    Ligand exchange color reaction and absorbance reading:

Add 700 μL of Cu-CAS assay solution to each microtube, including reference microtubes containing 700 μL of water, and read absorbance at 582 nm.

  1. 6.

    Plotting of results:

Plot the ratio of Abs(sample)/AbS(ref) vs. the actual chelate solution volume in each microtube (similar to Fig. 7, where x axis is sample volume rather than concentration). Note that one-half of the initial volume added in step 2 is taken for the next microtube in the dilution step (4).

  1. 7.

    Calculations and interpretation:

Calculate chelate concentration according to sample volume at the intercept with the x axis at y = 0, the Cu-known concentration (100 μM), and the Cu/ligand ratio of the complex.

3.1 Diurnal Rhythm of Release of PS

The rate of PS release in the Fe-deficient plants showed a distinct diurnal rhythm with a maximum value about 4 h after the onset of the light period. In contract to the Fe-deficient plants, in the Fe-sufficient plants the release of PS was very low and nearly constant throughout the daytime. Among graminaceous plants examined, there are two patterns for the secretion of phytosiderophores. A distinct diurnal rhythm in secretion has been reported in barley (Takagi et al. 1984), wheat (Zhang et al. 1991), Hordelymus europaeus (Gries and Runge 1992), and Festuca rubra (Ma et al. 2003). In other species, such as maize (Yehuda et al. 1996) and rice (Inoue et al. 2009), there is no diurnal rhythm in the secretion. Species with diurnal secretion patterns have different times of peak secretion rate. For example, in barley and wheat, maximum secretion rates occurs 4 h after the onset of light period (Takagi et al. 1984; Zhang et al. 1991). On the other hand, the secretion peak occurred at 5.5 h in Hordelymus europaeus (Gries and Runge 1992) and between 2 and 5 h in Festuca rubra (Ma et al. 2003). Since the growth conditions are different in these previous studies, it is difficult to conclude whether the differences in the secretion time results from the growth conditions or from species itself. In the present study, the secretion pattern between P. pratensis and L. perenne under the same growth conditions was compared. P. pratensis and L. perenne secrete different kinds of phytosiderophores in response to Fe deficiency; P. pratensis secretes DMA, AVA, and HAVA, while L. perenne secretes DMA, HDMA, and epiHDMA (Ueno et al. 2007). The amount of these phytosiderophore secreted is higher in P. pratensis than in L. perenne, but the secretion amount increased with the progression of Fe deficiency in both species (Ueno et al. 2007). Since the growth rate was different between the two species tested and the experiments were conducted in different seasons, we had to use plants of different ages and with different lengths of Fe-deficiency duration in order to obtain sufficient amounts of phytosiderophores for quantitative determination. However, P. pratensis and L. perenne showed distinct diurnal rhythms in the secretion of phytosiderophores irrespective of plant age and the duration of Fe deficiency, with a difference of 2–3 h in the secretion peak between the two species. Experimental results revealed that comparing ten cultivars of perennial grasses but with fewer collection times, the secretion pattern differed between the species, but not cultivars (Ueno et al. 2007). These results indicated that the secretion time differ consistently between P. pratensis and L. perenne. The diurnal rhythm in the secretion of phytosiderophore was suggested to be affected by both temperature and light (Ma et al. 2003; Reichman and Parker 2007). Earlier secretion was correlated with increased temperature; however, shading experiments revealed that phytosiderophore secretion is not triggered by light in both perennial grass species. Furthermore, the temperature of the rooting zone, but not the air temperature, controls secretion time. These results support the idea that the initiation of phytosiderophore secretion is triggered by the temperature around the roots. By using a square-wave light regime, Reichman and Parker (2007) concluded that the secretion of phytosiderophores in wheat is mainly mediated by changes in light rather than temperature. However, in their study, plants were shaded for a longer time, which may have affected phytosiderophore biosynthesis. Therefore, the lack of secretion of phytosiderophores under darkness may be the result of decreased synthesis. Another possibility is that phytosiderophore secretion time is controlled differently between wheat and perennial grasses. The mechanism responsible for diurnal rhythm of phytosiderophore secretion is still unknown. Recently, diurnal changes were reported in the expression of some genes involved in biosynthesis of phytosiderophores in rice (Nozoye et al. 2004) and uptake of Fe (III)-phytosiderophore complex in rice and barley (Inoue et al. 2009). Some elements associated to the diurnal change have been proposed to be present in the promoter region of these genes. Since the gene responsible for the secretion of phytosiderophore has not been cloned yet, it remains to be examined whether similar elements are involved in the diurnal rhythm of phytosiderophore secretion or the secretion is regulated independent of biosynthesis and uptake. Phytosiderophore secretion by grasses may increase Fe availability for coexisting species. Recently, it was reported that citrus can utilize Fe effectively from Fe(III)-phytosiderophore complex secreted from Poa (Cesco et al. 2006). Moreover, the combination of three perennial grasses, F. rubra, L. perenne and P. pratensis, has been shown to prevent Fe deficiency more effectively compared to single species. The secretion peak time of F. rubra is between those of L. perenne and P. pratensis (Ma et al. 2003). When these perennial grass species are grown together in an orchard, the combined effect of different secretion peak times may maintain Fe availability for orchard trees for longer period of time relative to single grass species.

4 Role of the PM in Acquisition of Different Nutrients in Plants

In addition to their role in Fe acquisition, it has been hypothesized that PS have a universal role for acquisition of other trace metals such as Zn, Mn, and Cu that also have low solubilities in alkaline soils. In support of this hypothesis, it has been shown that PS form stable chelates with Zn, Mn, and Cu and are effective in extracting these elements from calcareous soils. Production of PS has also been shown to be induced by Zn deficiency in wheat. However, the importance of PS as a general response to trace metal deficiencies remains uncertain. Presently there are no data on root chelator exudation for Poaceae species subjected to Cu and Mn deficiencies, and PS release rates under Zn deficiency reported by Zhang and coworkers were considerably lower than those typically observed with Fe-stressed wheat. MA enhanced the solubility of Fe (III) between pH 4 and 9, when added to nutrient solution MA strongly stimulated the uptake of Fe by “Fe-inefficient” rice seedlings. Commonly used chelating agents such as EDTA, EDDHA, and citrate, etc. had no stimulative effects. The MA-mediated Fe uptake proved to be dependent on metabolic energy. These results suggest the possibility of MA as a phytosiderophore for graminaceous plants.

4.1 Iron

Graminaceous species acquire Fe by releasing PS with a high binding affinity for Fe and by taking up ferreted PS through a specific transmembrane uptake system. An increased mobilization of Fe from a calcareous soil, even as far away from the root surface as 4 mm, demonstrated a high capacity of PS to mobilize Fe. The rate of PS exudation from roots (an average exudation rate of 2.9 nmol cm−1 root h−1) is possibility related to tolerance of different species and genotypes to Fe deficiency, Zuo et al. (2000) which formed a basis for screening plants for their relative Fe uptake efficiencies. In general, the broad trend seems to be that plants which release low levels of PS in response to Fe deficiency are adapted poorly to Fe-limiting soils. It has been well established that the undissociated Fe (III)-PS complex is taken up by corn and rice roots. The uptake of Fe (III)-PS was inhibited by metabolic inhibitors (DCCD or CCCP) and chilling, indicating that the transport of the Fe (III)-PS complex across the plasma membrane is an energy-dependent process. Aciksoz (2011) reported that root release of phytosiderophores (PSs) is an important step in iron (Fe) acquisition of grasses, and this adaptive reaction of plants is affected by various plant and environmental factors. The results show that the root release of PS, mobilization of Fe from 59Fe(OH)3, and root uptake and shoot translocation of Fe(III)-PS by durum wheat are markedly affected by N nutritional status of plants. The complex formation properties of mugineic acid, which is a biologically important molecule for iron uptake, were studied using the density-functional methods combined with the IEF-PCM continuum solvation model. In particular, it has been found that the inclusion of explicit water molecules interacting with mugineic acid is a key factor for obtaining reliable computational results. The present computational results show that the metal coordination structure is somewhat different between the Fe II-mugineic acid and Fe III-mugineic acid complexes; the former has a five-coordinated structure while the latter has a nearly octahedral binding structure. Sugiura et al. have suggested that the reduction of the Fe III complex into the Fe II complex is an important first process in the iron release mechanism in organism’s cell since the iron ion can be easily released from the weakly bonded ferrous complex [Fe II(HMA)] formed in the reduction process. The structural difference theoretically predicted in this work may play a role in this iron release mechanism although the detailed mechanism has not yet been understood at a molecular level. The characterization of phytosiderophore secretion patterns of perennial grasses provides important information for designing an optimal combination of species that can be used for effective correction of Fe-induced chlorosis of crops grown on calcareous soils.

4.2 Zinc

Exudation of PM from roots increases under Zn deficiency in a range of plant species. However, an equivocal experimental proof that PM play a role in mobilization and uptake of Zn from Zn-deficient soils has yet to be reported. This is especially important because PM has a greater affinity for Fe than for Zn (e.g., DMA has a twofold higher stability constant for Fe than for Zn). Either Zn or Fe deficiency may stimulate production of PM in wheat. However, an increased release of PM under Zn deficiency might be due to an indirect effect, for example, as a response to impaired translocation of Fe from roots to shoots under Zn deficiency. Such an imbalance in Fe circulation in plants might cause hidden physiological Fe deficiency, resulting in the increased PM release. For wheat, recent cause study suggested a model in which PM, released across plasma membrane, mobilize Zn in the apoplasm of root cells, But dissociation of the Zn (II)-PM complex occurs at the plasma membrane, and only Zn is taken up into the cytoplasm. However, more recent research has indicated that not only splitting of Zn (II)-PM complex as the plasma membrane and uptake of ionic Zn (II) occur but that the Zn (II)-PM complex can also be taken up undissociated, at least by corn roots. In addition to exudation of PM, Zn deficiency increases root exudation of amino acids, sugars, and phenolics in a range of plant species, including wheat. The importance of this exudation has yet not been assessed in terms of increasing plant capacity to acquire Zn from soils with low Zn availability. Sorghum and wheat plants increased the release of phytosiderophore in response to Zn deficiency but corn did not. The total amount of phytosiderophore released by the roots was in the order wheat > sorghum > corn. The total Zn uptake by the species in this study decreased in the order corn > sorghum > wheat, which is inversely related to their tolerance to Zn deficiency. The absence of a “phytosiderophore” response to Zn deficiency in corn, coupled with the evidence that this species accumulates more Zn than wheat or sorghum, provides an explanation for why Zn deficiencies are more prevalent for corn than wheat or sorghum. Soils in many agricultural areas are alkaline and have high amounts of calcium carbonate, resulting in low availability of Zn (Welch et al. 1991). Wheat grown on such soils suffers from Zn deficiency, although tolerance to Zn deficiency largely varies among wheat genotypes (Khoshgoftar et al. 2006). Wheat genotypes differ in their mechanisms for improved root Zn uptake under Zn-deficient conditions (Hacisalihoglu and Kochian 2003). These differences in the ability of genotypes for root uptake means the available pools of Zn in soil may vary for different genotypes (Marschner 1995). Plant Zn uptake is influenced by root architecture, the presence of mycorrhizal fungi, proton exudation from roots, and release of phytosiderophores (Hacisalihoglu and Kochian 2003). Some genotypes are able to release phytosiderophores from roots to the surrounding rhizosphere soil, which increases the solubility of Zn and, as a result, its availability for plant uptake (Hacisalihoglu and Kochian 2003). To date, nine mugineic acids have been identified in different graminaceous plants (Ueno et al. 2007). Under Zn-deficiency stress, the rate of phytosiderophore release differs among and within cereal species (Cakmak et al. 1996). The well-known differences among durum and bread wheat genotypes in Zn efficiency are closely related to the differences in the rate of phytosiderophore release from roots (Cakmak et al. 1996). Soil salinity is frequently associated with alkaline soils, which are commonly deficient in available Zn (Khoshgoftar et al. 2006). Salinity may reduce Zn uptake by plant roots due to competition of other cations, e.g., Ca and Na, at the root surface (Marschner 1995). For all three of the wheat genotypes studied, salinity stress resulted in greater amounts of phytosiderophores exuded by the roots. In general, for Kavir, the greatest amount of phytosiderophores was exuded from the roots at the highest salinity level (120 mM NaCl). Greater phytosiderophore exudation under Zn-deficiency conditions was accompanied by greater Fe transport from root to shoot. The relationship between Fe transport to shoots and differential exudation of phytosiderophores by wheat genotypes has been proposed as a physiological mechanism behind differential genotypic tolerance to zinc deficiency (Rengel and Graham 1995). Under such circumstances, decreased transport of Fe toward leaves under Zn deficiency would result in physiological Fe deficiency in leaves that would trigger increased exudation of phytosiderophores into the rooting medium by genotypes tolerant to Zn deficiency. In contrast, genotypes sensitive to Zn deficiency would transport relatively large amounts of Fe to leaves, thus avoiding physiological Fe deficiency and lacking a trigger for increasing root exudation of phytosiderophores. Many vascular plant species are unable to colonize calcareous sites. Thus, the floristic composition of adjacent limestone and acid silicate soils varies greatly. The inability of calcifuge plants to establish in limestone sites could be related to a low capacity of such plants to solubilize and absorb Fe from these soils. Under Fe deficiency, species of Poaceae enhance their Fe uptake by releasing non-proteinogenic amino acids, phytosiderophores (PS), from their roots which mobilize Fe from the soil by forming a chelate that is then taken up by the root (Römheld and Marschner 1986). In previous research it was shown that calcicole grasses are better adapted to low Fe availability on calcareous sites, as a consequence of higher PS release rates and lower tissue Fe demand. Phytosiderophore release and uptake is thought to be specific for Fe deficiency. However, a universal role of phytosiderophores in the acquisition of micronutrient metals has been proposed (Crowley et al. 1987), since PS form stable chelates with Zn, Mn, and Cu (Nomoto et al. 1987; Murakami et al. 1989); they extract considerable amounts of Zn, Mn, and Cu from calcareous soils (e.g., Treeby et al. 1989), and deficiencies of Zn, Mn, and Cu are quite common on calcareous and non-calcareous soils. Enhanced PS release in Zn deficient wheat has been reported by Zhang et al. (1989, 1991). The effect of zinc nutritional status of the plant on the release of zinc mobilizing root exudates was studied in various dicotyledonous (apple, bean, cotton, sunflower, tomato) and graminaceous (barley, wheat) plant species grown in nutrient solutions. In all species, zinc deficiency increased root exudation of amino acids, sugars, and phenolics. However, the root exudates of zinc-deficient dicotyledonous species did not enhance zinc mobilization from a synthetic resin (Zn chelate), or a calcareous soil, although mobilization of iron from FeIII hydroxide was increased. By contrast in the graminaceous species, root exudates from zinc-deficient plants greatly increased mobilization of both zinc and iron from the various sources. These differences in capability of mobilization of zinc and iron between the plant species are the result of an enhanced release of phytosiderophores with zinc deficiency in the graminaceous species.

Ptashnyk et al. (2011) reported that rice (Oryza sativa L.) secretes far smaller amounts of metals complexing phytosiderophores (PS) than other grasses. But there is increasing evidence that it relies on PS secretion for its zinc (Zn) uptake. After nitrogen, Zn deficiency is the most common nutrient disorder in rice, affecting up to 50 % of lowland rice soils globally. A mathematical model was developed of PS secretion from roots and resulting solubilization and uptake of Zn, allowing for root growth, diurnal variation in secretion, decomposition of the PS in the soil, and the transport and interaction of the PS and Zn in the soil.

4.3 Manganese

Environmentally controlled changes in redox potential occur when oxygen is depleted; NO3−, Mn, and Fe then serve as alternative electron acceptors for microbial respiration and are transformed into reduced ionic species. This process greatly increases the solubility and availability of Mn and Fe but is not under direct control of the plant. In some circumstances, such as in poorly aerated soils, this results in Mn and Fe toxicities to plants. Manganese availability may be further influenced by the activity of Mn-oxidizing and Mn-reducing bacteria that colonize plant roots. Since differential Mn efficiency can only be demonstrated for plants growing in soil, but not for those growing in the nutrient solution, it appears obvious that a change in the biology and/or chemistry of the rhizosphere precedes an increase in Mn availability to plants. However, the nature and activity of root exudates components that might be involved in mobilization on Mn is still unclear. Further research on root exudates effective in mobilizing Mn from the high-pH substrates for uptake by plants roots is warranted.

4.4 Copper

Phytosiderophore (PS) release in H. europaeus was rapidly induced in response to both Fe and Cu deficiencies. This is the first reported case of Cu-deficiency-induced PS release in grasses. Fe- and Cu-deficient plants were able to maintain release rates well above background levels even when growth was reduced to 6 or 31 % of the control, respectively. However, the plants in the metal-deficiency treatments progressed from normal growth to severely deficient in 30 day. For the induction of Cu deficiency, this is in agreement with Gries et al. (1995) but contrary to the findings of Bell et al. (1991), who suggested that Cu deficiency could only be obtained using a combined BPDS-HEDTA chelator-buffered system. Theoretically, other nonspecific chelators such as citric acid, which has some affinity for Cu and is present in root exudates of Fe- and Cu-deficient H. europaeus, could contribute to the Fe-mobilizing capacity of root exudates under Cu deficiency. The fact that almost the same constant ratio between the two assay methods was found for root exudates from both Fe-deficient and Cu-deficient plants suggests that the same chelators are produced under Cu deficiency as under Fe deficiency. Several studies using wheat (Triticum aestivum and Triticum durum) have demonstrated that PS release is enhanced under Zn deficiency (Zhang et al. 1989, 1991) and can reach levels comparable to those of PS release by Fe-deficient barley (Cakmak et al. 1994). Theoretically, the same could be true for the response to Cu deficiency that was observed. The hypothesis that PS release is a physiological response to Cu deficiency is further supported by the fact that Cu-deficient plants were able to maintain high rates of PS exudation throughout the experiment even when growth was reduced to less than one-third of the control. Also, the diurnal pattern of PS release under Cu deficiency was identical to that known from Fe-deficient H. europaeus plants (Gries and Runge 1992). In combination, these findings suggest that PS release in response to Cu deficiency is a well-regulated mechanism. In barley, uptake rates of PS-complexed Cu are tenfold lower than those of the PS-Fe complex (Ma et al. 1993), suggesting that the PS system functions primarily for Fe transport. This preferential recognition of Fe-PS complexes remains to be examined for H. europaeus. Nonetheless, even tenfold lower uptake rates of Cu-PS could still be sufficient to meet plant Cu demand. Based on calculations of plant yield and tissue Cu concentration, the quantities of chelators released under Cu deficiency greatly exceed the Cu uptake rate required for normal growth. Studying metal extraction by PS from a wide range of calcareous and non-calcareous soils revealed that PS preferentially mobilized Fe but also significant quantities of Zn and Cu from soils. Considering that plant Cu demand is much lower than Fe demand, the amounts of Cu mobilized appeared sufficient to meet plant requirements for this metal. This suggests that PS release would be an advantage for plants growing on soils low in available Cu. Release of PS under Cu deficiency could have an ecological significance, regardless of whether it is indirectly caused by impaired metabolism or as a specific response mechanism. This question needs further study. Also, it remains to be examined whether Cu-deficiency-induced PS release is a general phenomenon in native grass species.

5 Intercropping and Phytosiderophore Release

Micronutrient deficiency in plants is becoming an increasingly important global problem. Proper metal transport and homeostasis are critical for the growth and development of plants and in order to potentially fortify plants preharvest. Also, improvement in Fe and Zn content in the edible portions of the plant will be helpful for alleviating human nutritional disorders worldwide (Welch and Graham 2002; Grotz and Guerinot 2006). reported that peanut intercropping with different gramineous species not only improved the iron nutrition of the peanut but also enhanced the Cu and Zinc content in the peanut shoot in the greenhouse experiment. Although, this was not statistically significant difference in the field experiment, the Cu and Zinc content in peanut shoot of intercropping showed a general increasing trend, which means that agronomic intercropping helps mobilize and uptake limiting nutrient elements as well as provides benefits through effects on plant growth, development, and adaptability to adverse environments. The possible reason for such differential effects on Cu and Zn concentrations of peanut plants caused by intercropping could be root exudates from gramineous species. Specifically, production and release from phytosiderophores of gramineous species may improve solubility of Fe, Zn, and Cu by chelation, which helps plants obtain those essential elements from the soil (Inal et al. 2007). The release of phytosiderophore by strategy II plants also improves Zn nutrition (Khalil et al. 1997). In the current study, Zn and Cu nutrition of peanut was improved by the associated maize, barley, oats, and wheat. Enhanced production of phytosiderophore by maize might be responsible for the increases in Zn and Cu concentration of peanut. On the other hand, one metal deficiency might cause an excess of another metal to be absorbed. We have learned from studies to date that the transporters involved in Fe uptake can transport a variety of divalent cations such as Zn and Cu. However, Mn concentration of the peanut shoot is significantly enhanced by serious iron deficiency of peanut in monocropping. The higher Mn concentration may be caused by the enhanced Mn uptake by the peanut roots due to increased reducing capacity of peanut roots in monoculture in conjunction with the appearance of Fe-deficiency chlorosis symptoms in young leaves. Peanut plants have similar uptake mechanisms of Fe and Mn that require the reducing capacity of the root to accumulate Mn2+ and Fe2+ ions in the rhizosphere soil. The peanut intercropped with strategy II plants could not only improve iron nutrition of peanut but also enhance other critical micronutrients, such as Zn and Cu, which are critical metals for the growth and development of plants. However, systemic mechanisms might be involved in adaptation to these nutrient stresses at the whole plant level. Reasonable intercropping system of nutrient-efficient species should be considered to prevent or mitigate iron and zinc deficiency of plants in agricultural practice.

6 Ecological and Soil Chemical Factors Affecting the Efficacy of PS

The mechanism by which plants acquire Fe from siderophores in soils is not yet known. Many indirect processes such as extracellular reduction, chelate degradation, or passive diffusion may also contribute to root uptake of Fe mobilized by PS. Soil chemical and ecological factors can also affect the efficacy of PS in soils. Among the most obvious soil factors are Fe mineral dissolution rates, exchange kinetics for Fe complexed by organic matter, and solution pH of Fe redox potential. Physical structure and soil moisture content influence the diffusion path of the Fe and are important in soil aeration. Soil clay and organic matter strongly adsorb PS. Important biological factors include plant growth rates which affect Fe demand and the consequent induction of responses to Fe stress.

7 Summary

The availability of nutrients in plants is determined by the type of soil, climate conditions, and crop species, and the cultivars within the species determine the availability of nutrients in plants. Those crop species or cultivars that have the ability to absorb large amounts of nutrients and convert them into useful dry matter on highly enriched soils, in which a less-efficient species of cultivars reaches a yield plateau, have been described as the nutrient-efficient species or cultivars. However, with recent economic developments and the large potential of infertile soils that are located in developing regions of the world, it has been realized that the most significant contribution to world food production must come from crops grown on soils with relatively low fertility. More emphasis is now being given to that plant species or cultivars that should produce more on soils having a low fertility. Plant species or cultivars that produce higher yields under low nutrient supply have evolved one or more of the characteristics like an efficient internal economy, which may result from efficient redistribution within the plant, or lower requirements at functional sites. Due to escalating cost of chemical fertilizers, the nutrient uptake and utilization in crop plants should be most efficient to cause reduction in the cost of production and in achieving a higher profit for the farmers,. To arrive at these objectives, it is important to understand nutrient use efficiency, the factor effecting it, and ways of enhancing it in modern crop production system without reducing the crop/yields. To improve the nutrient use efficiency by crop plants, one of the strategies is to screen out the plants’ (high nutrient efficiency) group which can secret phytometallophores. Phytometallophores help the plants in greater absorption of the nutrients under sub-optional supply conditions. Phytometallophores (PM) are released in graminaceous species (Gramineae) under iron (Fe)- and zinc (Zn)-deficiency stress and are of great ecological significance for acquisition of Fe and presumably also of Zn. The potential for release of PS is much higher than reported up to now. Rapid microbial degradation during PM collection from nutrient solution-grown plants is the main cause of this underestimation. Due to spatial separation of PM release and microbial activity in the rhizosphere, a much slower degradation of PM can be assumed in soil-grown plants. Concentrations of PM up to molar levels have been calculated under non-sterile conditions in the rhizosphere of Fe-deficient barley plants. Besides Fe, PM mobilize also Zn, Mn, and Cu. Despite this unspecific mobilization, PS mobilizes appreciable amounts of Fe in calcareous soils and arc of significance for chlorosis resistance of graminaceous species. In most species the rate of PS release is high enough to satisfy the Fe demand for optimal growth on calcareous soils. In contrast to the chelates, ZnPM and MnPM, FePM are preferentially taken up in comparison with other soluble Fe compounds. In addition, the specific uptake system for FePM (translocator) is regulated exclusively by the Fe nutritional status.