Characterization of cadmium uptake, translocation and storage in near-isogenic lines of durum wheat that differ in grain cadmium concentration

Here we examined several physiological properties of two near-isogenic lines of durum wheat (Triticum turgidum var. durum) that differ in grain cadmium accumulation, to identify the function of a gene locus that confers differential grain Cd concentrations. Time- and concentration-dependent uptake and translocation studies using 109Cd were conducted on nutrient solution-grown seedlings. Root extracts were analysed by inductively coupled plasma emission spectrometry, gel filtration and capillary electrophoresis to determine the interaction between Cd and phytochelatins (PCs) in storage of Cd in roots. The two isolines did not differ in time- or concentration-dependent root Cd uptake, but the low grain-Cd-accumulating isoline showed decreased movement of Cd from roots to shoots. All buffer-soluble Cd extracted from roots of both isolines was in the form of a low-molecular-weight PC-containing complex. The data suggest that PC synthesis is not a limiting factor in the differential storage of Cd in roots, and that movement of Cd through the root and into the transpiration stream may be the cause of differential Cd partitioning in the two isolines.     

Zinc effects on cadmium accumulation and partitioning in near-isogenic lines of durum wheat that differ in grain cadmium concentration

Here, we examined the effectiveness of two approaches for reducing cadmium (Cd) accumulation in durum wheat (Triticum turgidum L. var durum) grain: the application of supplemental zinc (Zn), and the use of cultivars exhibiting reduced grain Cd concentrations. Two durum wheat near-isogenic lines (NIL) that differ in grain Cd accumulation were grown to maturity in solution culture containing a chelating agent to buffer the free activities of Zn and Cd at levels approximating those of field conditions. The low Cd accumulating (L-Cd) isoline had Cd concentrations, in grains and shoot parts, which were 60-70% lower than those of the high Cd accumulating (H-Cd) isoline. Increasing the Zn activities in the nutrient solution from deficient to sufficient levels reduced the concentration of Cd in grains and vegetative shoot parts of both isolines. The results suggest that supplemental Zn reduces Cd tissue concentrations by inhibiting Cd uptake into roots. Cd partitioning patterns between roots and shoots and between spike components suggest that the physiological basis for the low Cd trait is related to the compartmentation or symplasmic translocation of Cd.     

Differences in developmental plasticity and growth rate among drought-resistant and susceptible cultivars of durum wheat (Triticum turgidum L. var. durum)

Understanding how growth and development of durum wheat cultivars respond to drought could provide a basis to develop crop improvement programmes in drought-affected tropical and subtropical countries. A greenhouse experiment was conducted to study the responses of five durum wheat cultivars to moisture stress at different developmental phases. Phenology, total dry matter (TDM), relative growth rate (RGR), leaf area ratio (LAR), net assimilation rate (NAR), leaf weight ratio (LWR), specific leaf area (SLA) and shoot:root ratio were compared. Pre-anthesis moisture stress delayed phenological development, whereas post-anthesis moisture stress accelerated it. TDM accumulation rate was different between drought-resistant and susceptible cultivars. RGR and its components changed with age and moisture availability. Drought-resistant cultivars had a high RGR in favourable periods of the growing season and a low RGR during moisture stress. In contrast, the drought-susceptible cultivar (Po) showed an opposite trend. LAR explained the differences in RGR (r=0.788) best, whereas the relationship between NAR and RGR was not significant. Even though both LWR and SLA were important factors determining the potential growth rate, LWR was of major importance to describe cultivar differences in LAR, and consequently in RGR. The drought-resistant cultivars Omrabi-5 and Boohai showed vigorous root development and/or a low shoot:root ratio. It is concluded that biomass allocation is the major factor explaining variation in RGR among the investigated durum wheat cultivars.     

Transformation of pasta wheat (Triticum turgidum L. var. durum) with high-molecular-weight glutenin subunit genes and modification of dough functionality

Particle bombardment has been used to transform three cultivars (L35, Ofanto, Svevo) and one breeding line (Latino × Lira) of durum wheat (Triticum turgidum L. var. durum). These varieties were co-transformed with plasmids containing selectable and scorable marker genes (bar and uidA) and plasmids containing one of two high-molecular-weight (HMW) glutenin subunit genes (encoding subunits 1Ax1 or 1Dx5). Ten independent transgenic lines were recovered from 1683 bombarded scutella (transformation efficiency thus 0.6%). Five lines expressed either subunit 1Dx5 or 1Ax1 at levels similar to those of endogenous subunits encoded on chromosome 1B. To identify the effects of the transgenes on the functional properties of grain, three lines showing segregation for transgene expression were used to isolate sibling T₂ plants which were null or positive for the transgene product. Analysis of these plants using a small-scale mixograph showed that expression of the additional subunits resulted in increased dough strength and stability, demonstrating that transformation can be used to modify the quality of durum wheat for bread and pasta making.     

Influence of varied zinc supply on re-translocation of cadmium (109Cd) and rubidium (86Rb) applied on mature leaf of durum wheat seedlings

Effect of varied zinc (Zn) supply (0, 0.1, 1, 5 M) on re-translocation of radio-labeled cadmium (¹⁰⁹Cd) and rubidium (⁸⁶Rb) from mature leaf to root and other parts of shoot was studied in 11-day-old durum wheat (Triticum durum cv. C-1252) plants grown in nutrient solution under controlled environmental conditions. Application of ¹⁰⁹Cd and ⁸⁶Rb was carried out by immersing the tips (3 cm) of mature leaf in radio-labeled solutions for 10 s at three different times over a 42 h period. Differences in Zn supply for 11 days did not affect plant growth nor did it cause visual leaf symptoms, such as necrosis and chlorosis, at either the lowest or the highest Zn supply. Only at the nil Zn supply (0 M), shoot and root dry weights tended to decrease and increase, respectively, causing a lower shoot/root dry weight ratio. Partitioning of more dry matter to roots rather than shoots, a typical phenomena for Zn-deficient plants in nutrient solution experiments, indicated existence of a mild Zn deficiency stress at the nil-Zn treatment. Irrespective of Zn supply, plants could, on average, retranslocate 3.8% and 38% of the total absorbed ¹⁰⁹Cd and ⁸⁶Rb from the treated leaf to roots and other parts of shoots within 42 h, respectively. At nil-Zn treatment, 2.8% of the total absorbed ¹⁰⁹Cd was re-translocated from the treated leaf, particularly into roots. The highest re-translocation of ¹⁰⁹Cd (6.5%) was found in plants supplied with 0.1 M Zn. Increases in Zn supply from 0.1 M reduced ¹⁰⁹Cd re-translocation from 6.5% to 4.3% at 1 M Zn and 1.3% at 5 M Zn. With the exception of the nil-Zn treatment, the proportion of re-translocated ¹⁰⁹Cd was greater in the remainder of the shoot than in the roots. Contrary to the ¹⁰⁹Cd results, re-translocation of ⁸⁶Rb was not (at 0, 0.1 and 1 M Zn), or only slightly (at 5 M), affected by changing Zn supply. The results indicate an inhibitory action of increased concentrations of Zn in shoot tissues on phloem-mediated Cd transport. This effect is discussed in relation to competitive inhibition of Cd loading into phloem sap by Zn.     

Effects of suboptimal root zone temperatures and shoot demand on net translocation of micronutrients from the roots to the shoot of maize

The effects of suboptimal root zone temperatures (RZTs) on net translocation rates from the roots to the shoots and the concentrations of Fe, Mn, Zn, and Cu were examined in maize grown in nutrient solution or soil. Plants were grown at 12 °C, 18 °C and 24 °C RZT. At each RZT, the growth-related shoot demand for nutrients was varied by independently modifying the temperature of the shoot base (SBT) including the apical shoot meristem. The net translocation rates of Mn and Zn from the roots to the shoots were reduced at low RZTs, irrespective of the SBT and of the substrate (soil or nutrient solution). Obviously, the net translocation rates of Mn and Zn at low RZT were mainly regulated by temperature effects on the roots and not by the chemical nutrient availability in the rhizosphere or by shoot growth rate as controlled by SBTs. When both RZT and SBT were reduced, the decrease in net translocation rates of Mn and Zn was similar to the decline in the shoot growth rate and concentrations of Mn and Zn in the shoot fresh matter were not greatly affected or were even increased by low RZT. However, at high SBT and low RZT in nutrient solution, the depressed net translocation rates of Mn and Zn combined with the increased shoot growth resulted in significantly decreased concentrations of Mn and Zn in the shoot, indicating that Mn and Zn may become deficient even at high chemical availability. By contrast to Mn and Zn, the net translocation rates of Fe and Cu at all RZTs were markedly enhanced by increased SBTs. Accordingly, the concentrations of Fe and Cu in the shoot fresh matter were not greatly affected by RZTs, irrespective of the SBTs. These results indicate that the ability of roots to supply Fe and Cu to the shoot was internally regulated by the growth related shoot demand per unit of roots. Deceased 21 September 1996 Deceased 21 September 1996     

Micronutrient inflow rates and mobilisation into soil solution in the root zone of winter wheat (Triticum aestivum L.)

Concentrations of Cu, Mn and Zn in root-zone soil solutions were determined for a field-grown winter wheat crop, and their inflow rates were calculated. This latter parameter was highest in mid-July. Depletion rates of the root-zone soil solution were calculated for Cu, Mn and Zn. Although calculated depletion rates indicated adequate early season levels of micronutrients, these reserves were depleted more rapidly than could be explained by plant uptake. The size of the soil solution reservoir of micronutrients appears to depend on the balance between mobilisation and immobilisation which in turn depends on the flux of biologically produced chelating ligands. Commonwealth Aircraft Corporation Ltd, 304 Lorimar Street, Port Melbourne, Victoria 3207, Australia.     

Carbon isotope discrimination at vegetative stage as an indicator of yield and water use efficiency of spring wheat (Triticum turgidum L. var. durum)

A field experiment was conducted to investigate if carbon isotope (¹³C) discrimination () measured at the vegetative stage of spring wheat (Triticum turgidum L. var. durum) is related with the yield and water use efficiency (WUE) at ripening. A line source sprinkler irrigation system exposed the wheat genotypes to different watering regimes, from rainfed to full irrigation and thereby increased the range in yield and WUE attainable in the four genotypes studied. The results indicated that values measured at the late stem elongation stage 60 days after planting (DAP), showed strong positive correlation with total dry matter yield (r=0.732^***), and a highly significant negative correlation with WUE (r=–0.755^***) measured at ripening 105 DAP. The data suggest that the imprints of measured at vegetative growth stage persists throughout the entire growth period, until maturity. Subject to confirmation from additional studies in other crops and locations, early measurements of may prove a useful tool for rapid and early screening of cultivars, for high yield and high WUE.     

Differences between maize and wheat in growth-related nutrient demand and uptake of potassium and phosphorus at suboptimal root zone temperatures

The effects of low root zone temperatures (RZT) on nutrient demand for growth and the capacity for nutrient acquisition were compared in maize and wheat growing in nutrient solution. To differentiate between direct temperature effects on nutrient uptake and indirect effects via an altered ratio of shoot to root growth, the plants were grown with their shoot base including apical shoot meristem either within the root zone (low SB), i.e. at RZT (12°, 16°, or 20°C) or, above the root zone (high SB), i.e. at uniformly high air temperature (20°/16° day/night).At low SB, suboptimal RZT reduced shoot growth more than root growth in wheat, whereas the opposite was true in maize. However, in both species the shoot growth rate per unit weight of roots, which was taken as parameter for the shoot demand for mineral nutrients per unit of roots, decreased at low RZT. Accordingly, the concentrations of potassium (K) and phosphorus (P) remained constant or even increased at low RZT despite reduced uptake rates.At high SB, shoot growth at low RZT in both species was higher than at low SB, whereas root growth was not increased. At high SB, the shoot demand per unit of roots was similar for all RZT in wheat, but increased with decreasing RZT in maize. Uptake rates of K at high SB and low RZT adapted to shoot demand within four days, and were even higher in maize than in wheat. Uptake rates of P adapted more slowly to shoot demand in both species, resulting in reduced concentrations of P in the shoot, particularly in maize.In conclusion, the two species did not markedly differ in their physiological capacity for uptake of K and P at low RZT. However, maize had a lower ability than wheat to adapt morphologically to suboptimal RZT by increasing biomass allocation towards the roots. This may cause a greater susceptibility of maize to nutrient deficiency, particularly if the temperatures around the shoot base are high and uptake is limited by nutrient transport processes in the soil towards the roots.     

Antagonistic effect of calcium, zinc and selenium against cadmium induced chromosomal aberrations and micronuclei in root cells of Hordeum vulgare

The antagonistic effect of calcium (Ca²⁺), zinc (Zn²⁺) and selenium (Se⁴⁺) at different concentrations (10⁻²–10⁻⁶ M) against cadmium (Cd²⁺) induced genotoxic effects in root cells of Hordeum vulgare were studied. The results showed that 10⁻³–10⁻⁵ M could induce chromosomal aberrations and micronuclei formation. But in the treatment with 10⁻²–10⁻⁶ M of Ca²⁺, Zn²⁺ and Se⁴⁺ together with Cd²⁺ (10⁻³–10⁻⁵ M), respectively, the frequencies of chromosomal aberrations and micronuclei effectively decreased after 48 h of treatment. The treatment with 10⁻⁴–10⁻⁶ M of Ca²⁺ together with 10⁻⁴–10⁻⁵ M Cd²⁺, 10⁻⁶ M of Zn²⁺ together with 10⁻⁵ M Cd²⁺ and 10⁻⁶ M of Se⁴⁺ together with 10⁻⁵ M Cd²⁺ suggested rather obvious antagonistic effects. The order of the antagonisms of Ca²⁺, Se⁴⁺ and Zn²⁺ against Cd²⁺ toxicity was Ca²⁺>Se⁴⁺>Zn²⁺. The degree of antagonisms of Ca²⁺, Se⁴⁺ and Zn²⁺ against Cd²⁺ related to their concentration ratio.     

The effect of rhizoctonia root disease and applied nitrogen on growth,nitrogen uptake and nutrient concentrations in spring wheat

Root disease caused by Rhizoctonia solani is a common problem of spring wheat in South Australia. There are reports that nitrogen applications can reduce the incidence and severity of the disease. A glasshouse trail in pots examined the effects of disease and of applied nitrogen on wheat growth, and evaluated the utility of the basal stem nitrate concentration in diagnosing deficiency in plants with and without root disease. Plants were harvested at the mid-tillering stage. Shoot growth was increased by applied nitrogen until a maximum yield was attained, after which additional N had no effect on shoot yield. Root growth, however, responded positively only to low levels of applied N, after which it declined, and in the highest N treatment root mass was less than in the plants without applied N. Root disease caused severe reductions in plant growth, and both root and shoot mass were affected similarly. Even though growth of diseased plants responded positively to applied nitrogen the response was less than that of disease-free plants. The critical concentration of basal stem nitrate-N did not appear to be affected by root disease, and was estimated at 1200 mg kg^-1, consistent with other glasshouse data. The basal stem nitrate-N concentration, either in fresh or dried tissue, appeared a better diagnostic tool of N stress than did total shoot N concentration or content, because of sharper definition of critical concentrations. Concentrations of other nutrients in shoot tissue were affected differentially by both applied nitrogen and root disease, but generally did not reach critical levels, although phosphorus and magnesium appeared deficient in very disease-stressed plants.     

Effect of genome, induction medium and temperature pretreatment on green plant production in durum ( Triticum turgidum var. durum ) x bread wheat ( Triticum aestivum L. em Thell) crosses

The recalcitrancy of durum wheat (Triticum turgidum var. durum) to anther culture, was attempted to be overcome by transferring the responsible genes form bread wheat B-genome to the respective on durum wheat, determining an appropriate induction medium and clarifying the necessity of cold pretreatment. For this, three durum wheat cultivars were crossed to two bread wheat (Triticum aestivum L. em Thell) cultivars. The resulting F₁ plants and their original cultivars were grown in the field and anthers at the appropriate microspore stage were cultured on potato-2 and W₁₄ media with and without low temperature pretreatment. No green plants were produced from the parental durum wheat cultivars. In contrast, green plants were produced from the F₁ plants. The best results in three of the four F₁ hybrids were recorded when potato-2 was used as induction medium. A more variable response of the examined genotypes was noticed with respect to temperature pretreatment. Regarding green plant production, a negative effect of cold pretreatment was observed in two of the F₁ hybrids when they were cultured on potato-2. Chromosome counts on root tips from the resulting green plants revealed that they all carried D-genome chromosomes. The last observation could suggest that D-genome chromosomes are necessary for anther culture response in wheat. Yet, the production of one green plant with 15 chromosomes may indicate that the development of extracted durum genotypes from bread wheat genotypes with good response to in vitro anther culture might be possible. Further work however, is needed for this to be verified.     

Effects of the chemical form of inorganic nitrogen fertilizers on the dynamics of the soil solution composition and on nutrient uptake by wheat

Adding nitrogen (N) fertilizers to soil affects not only the concentration in the soil solution of the added ions, but also those of other ions already present in the soil. This secondary effect is caused by ion exchange and electrochemical equilibrium processes. We studied how different N fertilizers affected the chemical composition of the soil solution over time, and how this related to nutrient uptake by wheat. Soil was fertilized either with (NH4)2SO4 or Ca(NO3)2, or no N was added. Each of these N treatments was either planted or not with spring wheat (Triticum aestivum L.). Soil solutions were collected repeatedly with looped hollow fiber samplers from the root zone in situ, six times during a 50-day pot experiment. Plants were harvested five times, and their nutrient contents determined. In the soil solution, NO3- was significantly less concentrated if (NH4)2SO4, rather than Ca(NO3)2 was applied, until after net nitrification had ended on day 20. In contrast, Ca2+, Mg2+ and K+ were significantly more concentrated in the former treatment. This was probably caused by the greater concentration of anions that resulted from nitrification. P was always very dilute and unaffected by the form of N fertilizer. The form of N fertilizer had no significant effect on plant growth and nutrient uptake. The likely contribution of mass flow of the soil solution in supplying Ca, Mg and N to the plants was greatest when (NH4)2SO4 was supplied. The supply of K and P was unaffected by N fertilizer. The potential for N leaching loss was lower with (NH4)2SO4 than with Ca(NO3)2, especially up to day 20. However, the potential for cations leaching loss was greater in the (NH4)2SO4 treatment. This suggests that there is only a limited advantage in fertilizing with (NH4)2SO4 to reduce the total loss of nutrients from soil.     

Influence of pH on Cd and Cu uptake, distribution and their effect on nitrate reductase activity in cucumber (Cucumis sativus L.) seedling roots

Influence of different pH solutions (5.0 and 7.0) on Cu²⁺ and Cd²⁺ absorption and distribution in root cells as well as effects of these metals on nitrate reductase activity (NR) in roots of cucumber seedlings were estimated. The absorption of Cu and Cd by roots measured as metal depletion in uptake solution was similar, both metal absorption was independent of the pH of solution. However, after rinsing of roots in distilled water (30 minutes), more Cu than Cd was found in protoplasts of root cells. More Cu was measured in all cell fractions when Cu was uptaken from pH 5.0 than from 7.0. The nitrate reductase activity after one hour of metal treatments was drastically decreased by Cu. The strongest reduction of enzyme activity was observed in roots treated with Cu in buffer with pH 5.0. Influence of Cd on the enzyme activity was weaker and was independent of the pH of solution. Lower concentration of Cd in solution (20 μM) increased NR activity. The data obtained prove the higher mobility of Cu than Cd into the cells of root. The mobility of Cu depends on pH of solution. Cu ions, but not Cd, influenced membrane permeability (K leakage). Cu acted more drasticly than Cd on NR activity.     

Efficient and rapid <Emphasis Type="Italic">Agrobacterium-</Emphasis>mediated genetic transformation of durum wheat (<Emphasis Type="Italic">Triticum turgidum L. var. durum)</Emphasis> using additional virulence genes

Genetic transformation of wheat, using biolistics or Agrobacterium, underpins a range of specific research methods for identifying genes and studying their function in planta. Transgenic approaches to study and modify traits in durum wheat have lagged behind those for bread wheat. Here we report the use of Agrobacterium strain AGL1, with additional vir genes housed in a helper plasmid, to transform and regenerate the durum wheat variety Ofanto. The use of the basic pSoup helper plasmid with no additional vir genes failed to generate transformants, whereas the presence of either virG⁵⁴² or the 15 kb Komari fragment containing virB, virC and virG⁵⁴² produced transformation efficiencies of between 0.6 and 9.7%. Of the 42 transgenic plants made, all but one (which set very few seeds) appeared morphologically normal and produced between 100 and 300 viable seeds. The transgene copy number and the segregation ratios were found to be very similar to those previously reported for bread wheat. We believe that this is the first report describing successful genetic transformation of tetraploid durum wheat (Triticum turgidum L. var. durum) mediated by Agrobacterium tumefaciens using immature embryos as the explant.