Iron-induced Nickel Deficiency in Pecan

Economic loss resulting from nickel (Ni) deficiency can occur in horticultural and agronomic crops. This study assesses whether excessive iron (Fe) can induce Ni deficiency. Both chelated Fe and diethylenetriaminepentaacetic acid (DPTA; a commonly used Fe-chelant) induces Ni deficiency in pecan [Carya illinoinensis (Wangenh.) K. Koch]. Foliar sprays of Fe [Fe-DPTA (1.1995 g·L)] during early post-budbreak shoot growth can trigger, or increase in severity, Ni deficiency symptoms in the emerging pecan canopy. Deficiency is also inducible in greenhouse-grown ‘Desirable’ seedlings at budbreak by Fe-DPTA application to soil and to a much lesser extent by DPTA alone. Endogenous Fe, just after budbreak, triggers Ni deficiency-associated distortions in pecan seedling leaf growth and morphology when the Fe:Ni is ’150 or greater with subsequent severity being proportional to the Fe:Ni ratio and Fe:Ni ’1200 or greater triggering extreme dwarfing of canopy organs. Timely treatment of symptomatic organs with foliar-applied Ni-sulfate restores normal growth, whereas foliar treatment with salts of other transition metals (titanium, vanadium, chromium, cobalt, copper, zinc, and molybdenum) of possible metabolic significance is ineffective. Results indicate that excessive endogenous Fe, and DPTA to a lesser extent, in organs and tissues during early post-budbreak growth can trigger Ni deficiency. A similar Fe on Ni antagonism may also occur with the Ni-associated nutritional physiology of other crops; thus, excessive exposure to chelated Fe not only triggers Ni deficiency in pecan, but may also occur in other horticultural and agronomic crops. Nickel is an often-overlooked plant (Brown et al., 1987, 1990) and animal (Welch and Graham, 2005) essential micronutrient. Although Ni deficiency in plants severe enough to trigger visual symptoms is relatively rare, compared with other essential micronutrients, both visual and non-visual deficiencies may be more common than generally supposed. This is partially because of antagonistic interactions between Ni and certain first-period transition metals (Wood, 2010). There is a dearth of information regarding the physiology of Ni’s interaction with other essential and beneficial micronutrients; however, excessive tissue zinc (Zn) or copper (Cu)—i.e., a high Zn:Ni or Cu:Ni ratio—can trigger symptoms of Ni deficiency (Wood, 2010). Because of relatively great physiochemical similarity between Fe and Ni, it is likely that excessive endogenous Fe can disrupt Ni-dependent physiology enough to trigger economic crop loss. A natural consequence of insufficient understanding is accidental induction of Ni deficiency in crops resulting from either excessive supplemental fertilization with certain trace metals and/or cropping on mineralized soils relatively rich in these metals. Pecan trees growing in commercial orchards, yards, gardens, and nurseries often exhibit Ni deficiency during early spring when canopies are rapidly expanding. Ni deficiency often manifests itself as a potentially fatal orchard replant malady when young transplants replace missing trees in mature orchards or are planted in secondgeneration orchard sites (Wood et al., 2003a, 2003b, 2004, 2006a). Incidence and severity of deficiency vary with tree age, or size, and on the nature of the action and interaction of several biotic and abiotic soil factors (Wood et al., 2006b). Severe Ni deficiency can kill young pecan trees (Wood et al., 2004), supporting conclusions by Brown et al. (1987, 1990) that Ni is an essential nutrient element for higher plants. The fundamental cause(s) of Ni deficiency in soils containing sufficient Ni to meet plant needs vary but include nematode damage to feeder roots (Nyczepir et al., 2006), excessively cool and/or dry soils during early spring (Wood et al., 2006b), excessive Zn and/or Cu (Wood, 2010), and possibly excessive long-term use of glyphosate (Yamada et al., 2009). Iron fertilizers are typically ‘‘chelates’’ that bind Fe (ferric, or oxidized Fe). A common form is Fe-DPTA. Iron (Fe) chelates bind to the cytoplasmic plasmalemma, where, in dicots, sequestered Fe is chemically reduced to Fe before release from the chelant molecule and subsequent transport across the plasma membrane into the cytoplasm (Chaney et al., 1972; Romheld and Marschner, 1986). Roots can also absorb small amounts of chelants (Tiffin and Brown, 1961; Tiffin et al., 1960; Weinstein et al., 1951), which in turn can disrupt plant processes by sequestering divalent or trivalent metal ions needed for physiologically active complexes such as metalloenzymes. Pecan orchards, especially those established on relatively high pH soils, occasionally receive Fe-DPTA sprays for correction of Fe deficiency. Other field, vineyard, nursery, and hydroponic crops also receive Fe-DPTA on occasion. Excessive orchard fertilization can trigger Ni deficiency, especially if excessively high soil/tissue Zn and/or Cu reduces the physiological availability of Ni within the plant (Wood, 2010; Wood et al., 2003a). Pecan foliage often exhibits visible Ni deficiency symptoms although the absolute foliar Ni concentration exceeds the apparent ‘‘lower critical’’ concentration (Nyczepir et al., 2006) of 0.85 mg·g dry weight, thus indicating that other nutrient elements can affect endogenous Ni bioavailability/use. Such micronutrient interactions are common in plants, especially in situations of extreme soil pH or metal composition (Kabata-Pendias, 2001). These interactions can trigger chemical stress linked to either antagonistic or synergistic effects on root uptake and/or cellular/enzymatic bioavailability/use. There are reports of Ni on Fe antagonism in which high Ni reduces endogenous Fe concentration and/or bioavailability (Chen et al., 2009; Ghasemi et al., 2009; Hewett, 1953; Khalid and Tinsley, 1980; Koch, 1956; Kovacik et al., 2009; Misra and Dwivedi, 1977; Nicholas and Thomas, 1954; Nishida et al., 2012). However, there is little information regarding the reverse Fe on Ni antagonism, especially in woody perennials. Cataldo et al. (1978) found that Fe suppresses Ni absorption and translocation in soybean (Glycine max), whereas Wallace et al. (1977a) found that Fe (as Fe-EDDHA) did not suppress Ni concentration in foliage of bush bean (Phaseolus vulgaris). Khalid and Tinsley (1980) concluded that in annual rye grass (Lolium multiflorum), it is the Ni:Fe ratio, rather than absolute concentration of either, in plant tissues and organs that is most tightly associated with reduced Fe bioavailability/use under high Ni conditions. The reverse Fe on Ni antagonism merits investigation. This study reports the effect of Fe-DPTA and DPTA on induction of Ni deficiency in pecan and documents a Fe on Ni antagonism in a long-lived woody perennial crop. Materials and Methods The following experiments test whether Fe-DPTA or DPTA induces or increases severity of Ni deficiency in pecan. Received for publication 24 June 2013. Accepted for publication 1 Aug. 2013. I gratefully acknowledge field and laboratory assistance by James Stuckey and Kirby Moncrief. Use of trade names does not imply endorsement of the products named or criticism of similar ones not named. To whom reprint requests should be addressed; e-mail [email protected]. HORTSCIENCE VOL. 48(9) SEPTEMBER 2013 1145 Effect of foliar applied Fe on Ni deficient shoots of ‘Wichita’ trees: Expt. 1 A single 30-year-old ‘Wichita’ tree growing in a commercial orchard near Cordele, GA, almost annually exhibits substantial Ni deficiency on certain major limbs 7 d after budbreak (i.e., parachute stage) with certain limbs failing to exhibit deficiency symptoms. This unique situation enabled testing of Fe’s ability to influence expression of Ni deficiency by treating individual small (4 cm or less) branches with Fe-DPTA, a common Fe-chelate (Sequestrene 330 Fe; a 10% Fe DTPA; Novartis Crop Protection, Inc., Greensboro, NC). Treatments were: 1) nontreated Ni-deficient control; 2) nontreated Ni sufficient (i.e., no visible symptoms) control; 3) Ni treatment of Ni-deficient branch; 4) Fe-DPTA treatment of Ni-deficient branch; and 5) Fe-DPTA treatment of Ni-sufficient branch. Ni was applied as Ni-sulfate (Ni at 100 mg·L) and Fe was applied as Fe-DPTA (at 1.1995 g·L). Treatments were applied at the parachute stage ( 5 to 7 d post-budbreak) of shoot development, immediately before onset of rapid organ growth and at a time when it became evident which branches were going to exhibit Ni deficiency sufficient to affect subsequent shoot and foliage morphology. Treatments were applied by foliar spray to runoff with treated limbs selected to avoid cross-contamination by other treatments. The experimental design consisted of five treatments replicated eight times throughout the tree’s canopy (n = 40). Statistical analysis was by analysis of variance (ANOVA) at P # 0.05 and mean separation by Tukey’s honestly significant difference (HSD) test at the same level after parameters were demonstrated to fit a normal distribution through the Shapiro-Wilk-W test for goodness of fit. Treatment effects were assessed in mid-May, 4 weeks after application of foliar sprays. Severity of Ni deficiency was noted using a scale representing a typical progression in degree of visible Ni deficiency symptoms: 1 = normal growth, no Ni-associated morphological distortions of shoots, compound leaves, or leaflets (i.e., normal appearance); 2 = 25% or less of leaflets on shoot exhibiting morphological distortions (i.e., slightly blunted leaflet apex); 3 = 26% to 50% of leaflets exhibiting some degree of morphological distortion; 4 = greater than 50% of leaflets exhibiting morphological distortion; 5 = #4, plus leaflet cupping; 6 = #5, plus necrosis of leaflet tips; 7 = #6, plus necrosis of leaflet margins, plus crinkled and dwarfed leaflets; 8 = #7, plus dwarfed shoots (i.e., short internodes); 9 = #8, plus rosetting; and 10 = #9, plus shoot death (Nyczepir et al., 2006; Wood, 2010; Wood et al., 2003a, 2003b). Effect of soil-applied Fe-DPTA on Ni deficiency in ‘Desirable’ trees: Expt. 2 Potted ‘Desirable’ trees on open-pollinated ‘Elliott’ rootstocks were grown in 15-L plastic pots filled with an artificial potting mix (SunGro Metro Perennial Mix; 62% to 72% composted bark, Canadian sphagnum peat, perlite, dolomite lime, and gypsum; Sun Gro, Bellevue, WA). Trees were treated at bud swell, going into their third leaf, with treatments being: 1) ‘‘control’’ (i.e., putative Ni-sufficient control); 2) ‘‘Fe’’ (3.2 g Fe-DPTA dissolved in 500 mL of water and applied as a soil drench); 3) ‘‘Ni’’ (Ni-sulfate, with Ni at 100 mg·L) sprayed onto buds at the parachute stage of budbreak; and 4) ‘‘Fe + Ni’’ (i.e., Fe applied as a soil drench as noted previously and with Ni applied as a foliar spray at the parachute stage of budbreak, as described previously). The experimental design consisted of four treatments and 15 replicates per treatment with single trees serving as replicates (n = 60). Trees were watered as needed and received 5 g of urea per potted tree (i.e., urea dissolved in 500 mL of water and then applied to each pot at bud swell and again at budbreak). Trees were rated for severity of Ni deficiency as noted in Expt. 1 4 weeks post-budbreak (i.e., rating the most severely affected leaflet based on the previously described rating scale). Statistical analysis was by ANOVA at P # 0.05 and mean separation by Tukey’s HSD at the same level after parameters were demonstrated to exhibit fit a normal distribution through the Shapiro-Wilk-W test for goodness of fit. Effect of Fe:Ni ratio on expression of Ni deficiency: Expt. 3 This study assessed the relationship between degree of Ni deficiency and tissue concentrations of Ni and Fe and the Fe:Ni ratio of affected foliage. Open-pollinated ‘Desirable’ seedlings were grown in plastic pots (15 · 15 · 15 cm) containing the previously described artificial potting mix. Third-leaf seedling trees, 30 to 40 cm tall, were defoliated in June to force new growth from dormant spring buds. Trees were fertilized with 1 g urea per pot at the time of defoliation and another gram at budbreak. Urea was applied in 100 mL deionized water flooded onto the surface of each pot. To achieve a gradient in Ni deficiency symptoms, the study consisted of seedlings fertilized with Fe-DTPA at different amounts per pot with there being 10 trees per rate. FeDPTA treatments were 0X, 1X, 2X, 4X, 8X, 16X, 32X, 64X, and 128X; with X = FeDPTA at 0.40 g/pot. The study used 90 seedlings. Fe-DPTA was applied once, at defoliation, by flooding the pot with the Fechelate dissolved in 300 mL of deionized water. Subsequent watering was such that there was no mass flow of water through the pot’s soil to avoid leaching of Fe. Buds broke at several nodes 10 to 14 d after defoliation. Seedlings were rated for severity of Ni deficiency 2 weeks after budbreak and leaflet lamina tissue (excluding the midrib) of the new canopy sampled for micronutrients. At the same time, all foliage and shoot tissue was sampled from the largest shoot mass of the three to four nodes that broke bud. These were subsequently measured for fresh weight, dry weight, and nutrient element concentration. Nutrients analyzed were first period transition elements [i.e., titanium (Ti), valadium (V), chromium (Cr), manganese (Mn), Fe, cobalt (Co), Ni, copper (Cu), Zn, plus molybdenum (Mo)]. Foliage collection used zirconium oxide ceramic scissors to avoid metal contamination of samples. Nickel deficiency of seedlings was verified by restoration of normal growth in the youngest emerging shoots after treating a subpopulation of symptomatic seedlings with analyticalgrade Ni-sulfate (i.e., Ni at 100 mg·mL) sprayed on the leaf lamina 7 d post-budbreak. Sample processing. Leaflet samples were air-dried to a constant weight at room temperature, diced using zirconium oxide ceramic scissors with small pieces thoroughly mixed in an acid-rinsed plastic container, and then 100 to 500 mg of bulked tissue placed in nylon tubes for processing. Sample digestion used 10 mL of 70% ultra-low trace elementgrade nitric acid (Sigma-Aldrich, Atlanta, GA) in a MarsXpress carousel placed within a Mars-5 (CEM Corporation, Matthews, NC) microwave digester. Cooled samples were filtered and brought to 20 mL (by weight; using 2% nitric acid) in 50-mL polypropylene centrifuge tubes and 0.5 mL (or 0.100 mL, for certain elements) added to a 15-mL plastic tube and brought up to 14.5 mL using a 2% nitric acid solution in preparation for metal analysis. Inductively coupled plasma mass spectrophotometry analysis. The concentration of Ti, V, Cr, Mn, Co, Cu, Zn, Mo, Fe, and Ni in leaf tissue samples was determined using an inductively coupled plasma mass spectrophotometer; PerkinElmer SCIEX ELAN-9000; Concord, Ontario, CA). Quantitative analysis was facilitated by a similar mass internal standard (Ge) and external standards using multielement standard solutions (PerkinElmer Multielement Calibration Standard Sets) diluted to cover three to four orders of magnitude. Each sample was analyzed in triplicate. Statistical analysis. Differential application of Fe produced a population of seedlings exhibiting Ni deficiency symptoms covering most of the entire rating range reflecting symptom severity. Measured parameters were therefore analyzed by curvilinear regression for the seedling population. Efficacy of transition metals for correcting Fe induced Ni deficiency symptoms: Expt. 4 Because of an association between concentrations of certain other transition metals and severity of Fe-induced Ni deficiencylike symptoms, these other transition metals were also tested for their ability to correct Ni deficiency. These were: Ti (TiH2), V (VCl2), Cr [Cr2(SO4)3·xH2O]; Mn (MnCl2·4H2O); Fe (FeSO4·7H2O); Co (CoSO4·7H2O); Ni (NiSO4·7H2O); Cu (CuSO4·7H2O); Zn (ZnSO4·7H2O); and Mo (H2MoO4). All were applied as a foliar spray 7 d post-budbreak with metal concentration at 25 mg·L for each transition metal. The study consisted of Fe-induced Ni deficiency in ‘Desirable’ seedling trees as described in the previous experiment (Expt. 3). The experiment design 1146 HORTSCIENCE VOL. 48(9) SEPTEMBER 2013 consisted of two treatments for each transition metal being tested—i.e., emerging shoots of symptomatic seedling were divided into either ‘‘symptomatic nontreated control’’ or ‘‘symptomatic + transition metal’’ treatments and treated accordingly with the relevant transition metal solution. Seedling shoot treatments were subsequently rated as described previously for severity of morphologically based Ni-like deficiency symptoms exhibited 10 d posttreatment. The study used 14 replications for each transition metal tested. The study was analyzed by ANOVA to determine treatment effects. Influence of DPTA and FeDPTA on Ni deficiency symptoms: Expt. 5 Chelating agents are potentially absorbed by roots and then xylem transported to the canopy where they can traverse the cellular plasma membranes and sequester metals within the cellular cytoplasm of developing canopy tissues. Two experiments were conducted to assess the possibility that the DPTA chelant might be the causal factor for triggering Ni deficiency rather than Fe. Both used 2-year-old open-pollinated seedling ‘Desirable’ trees grown in a potting mix as described in Expt. 2. In Study 1, seedlings were defoliated, fertilized with urea, and then treated through a soil drench with 1) deionized water control; 2) DPTA (at 500 mmoles/pot); 3) Fe-DPTA (at 500 mmoles/pot); 4) control plus Ni; 5) DPTA plus Ni; or 6) FeDPTA plus Ni. Nickel was applied as a foliar spray of Ni-sulfate (Ni-sulfate, with Ni at 2 mM) 7 d postbudbreak, whereas the DPTA and FeDPTA treatments were by soil application. Seedlings were rated for Ni deficiency symptoms as described previously. In Study 2, seedlings were defoliated, fertilized with urea, and then treated by a foliar spray 6 d post-budbreak with 1) deionized water control; 2) DPTA (at 3 mM) or 3) Fe-DPTA (at 3 mM); 4) control plus Ni; 5) DPTA plus Ni; or 6) FeDPTA plus Ni. Nickel was applied as a foliar spray of Ni-sulfate (Ni-sulfate, with Ni at 2 mM) 7 d postbudbreak and again 10 d post-budbreak. The seedlings were foliarly sprayed in a manner that ensured no DPTA and Fe-DPTA was deposited onto the potting soil mix in which trees were growing, thus ensuring that DPTA or Fe-DPTA had effects on expanding foliage from canopy absorption rather than from root absorption. Seedlings were rated for Ni deficiency symptoms as described previously, 4 weeks after treatment. The experimental design consisted of six treatments replicated eight times (n = 48). Statistical analysis was by ANOVA at P # 0.05 and mean separation by Tukey’s HSD test at the same level after parameters were demonstrated to fit a normal distribution through the Shapiro-Wilk-W test for goodness of fit.