Does high bicarbonate supply to roots change availability of iron in the leaf apoplast?

The role of the leaf apoplast in iron (Fe) uptake into the leaf symplast is insufficiently understood, particularly in relation to the supposed inactivation of Fe in leaves caused by elevated bicarbonate in calcareous soils. It has been supposed that high bicarbonate supply to roots increases the pH of the leaf apoplast which decreases the physiological availability of Fe in leaf tissues. The study reported here has been carried out with sunflower plants grown in nutrient solution and with grapevine plants grown on calcareous soil under field conditions. The data obtained clearly show that the pH of the leaf apoplastic fluid was not affected by high bicarbonate supply in the root medium (nutrient solution and field experiments). The concentrations of total, symplastic and apoplastic Fe were decreased in chlorotic leaves of both sunflower (nutrient solution experiment) and grapevine plants in which leaf expansion was slightly inhibited (field experiment). However, in grapevine showing severe inhibition of leaf growth, total Fe concentration in chlorotic leaves was the same or even higher than in green ones, indicative to the so-called `chlorosis paradox'. The findings do not support the hypothesis of Fe inactivation in the leaf apoplast as the cause of Fe deficiency chlorosis since no increase was found in the relative amount of apoplastic Fe (% of total leaf Fe) either in the leaves of sunflower or grapevine plants. It is concluded that high bicarbonate concentration in the soil solution does not decrease Fe availability in the leaf apoplast.     

Relationship between iron chlorosis and alkalinity in Zea mays

Mengel, K. and Geurtzen, G. 1988. Relationship between iron chlorosis and alkalinity in Zea mays . - Physiol. Plant. 72: 460–465.
Maize ( Zea mays L. cv. Anjou 21) grown in nutrient solution with Fe-EDTA and with nitrate as the sole nitrogen source showed typical Fe-chlorosis symptoms after a growth period of 14–21 days. Alkalinity in roots, stems and leaves of the chlorotic plants was high. Transferring the chlorotic plants from the nitrate-containing nutrient solution to a solution of (NH₄)₂SO₄ resulted in a regreening of leaves within 2–3 days which was associated with a decrease in solution pH, a decrease in alkalinity of plant parts, a translocation of Fe from roots to tops and a release of Fe into the outer solution. Similar effects were obtained when Fe chlorotic plants were transferred to a dilute HO solution with pH 3.5.
Spraying chlorotic leaves with indoleacetic acid or with fusicoccin led also to a regreening of leaves without having a major effect on leaf alkalinity.
Interpretation of the experimental results is based on the assumption that nitrate as sole N source leads to a high pH level in the apoplast resulting in the precipitation of Fe compounds, probably Fe oxide hydrate. Ammonium nutrition has the reverse effect since it lowers the apoplast pH and this can result in the dissolution of Fe compounds. Application of indoleacetic acid as well as fusicoccin supposedly stimulates the proton pumps in the plasmalemma of the leaf tissue. The resulting decrease in apoplast leaf pH in the microenvironment also leads to a dissolution of Fe compounds in the apoplast and thus promotes the uptake of Fe by the symplasm.     

Mechanism of Fe uptake by the leaf symplast: Is Fe inactivation in leaf a cause of Fe deficiency chlorosis?

The mechanism of iron (Fe) uptake from the leaf apoplast into leaf mesophyll cells was studied to evaluate the putative Fe inactivation as a possible cause of Fe deficiency chlorosis. For this purpose, sunflower (Helianthus annuus L.) and faba bean plants (Vicia faba L.) were precultured with varied Fe and bicarbonate (HCO ₃ ^- ) supply in nutrient solution. After 2–3 weeks preculture, Fe^III reduction and ⁵⁹Fe uptake by leaf discs were measured in solutions with Fe supplied as citrate or synthetic chelates in darkness. The data clearly indicate that Fe^III reduction is a prerequisite for Fe uptake into leaf cells and that the Fe nutritional status of plants does not affect either process. In addition, varied supply of Fe and HCO ₃ ^- to the root medium during preculture had no effect on pH of the xylem sap and leaf apoplastic fluid. A varied pH of the incubation solution had no significant effect on Fe^III reduction and Fe uptake by leaf discs in the physiologically relevant pH range of 5.0–6.0 as measured in the apoplastic leaf fluid. It is concluded that Fe inactivation in the leaf apoplast is not a primary cause of Fe deficiency chlorosis induced by bicarbonate. This revised version was published online in June 2006 with corrections to the Cover Date.     

Bicarbonate,the most important factor inducing iron chlorosis in vine grapes on calcareous soil

Summary In pot experiments grape vine was grown on a calcareous and on a non calcareous soil with a low and with a high water saturation. During the growing period soil solution samples were collected and analyzed for their pH and for HCO ₃ ⁻ , phosphate, Fe, and Ca. High water saturation resulted in a pH increase and in an increase of the HCO ₃ ⁻ concentration in both soils. The level in pH and HCO ₃ ⁻ , however, was much higher in the calcareous soil than in the non calcareous soil. The Fe concentration varied much throughout the experimental period, but there was no major differences between soils and water saturation treatments. The Ca concentration of the soil solution increased with time in the calcareous soil; for the non calcareous soil rather the reverse was true. The phosphate level in the soil solution of the non calcareous soil was about 10 times higher than in the calcareous soil. After 3 weeks growth all plants of the calcareous soil with the high water saturation showed first symptoms of Fe deficiency. These became more intense from day to day. Plants of the other treatments did not show any chlorotic symptoms. In the treatment with the chlorotic plants the HCO ₃ ⁻ concentration of the soil solution was the highest, the phosphate concentration the lowest from all treatments. It is therefore concluded that HCO ₃ ⁻ and not phosphate is the primary cause for lime induced Fe chlorosis. Despite the low phosphate concentration in the soil solution, the P concentration in the chlorotic leaves was more than twice as high as the P concentration in green leaves grown on the same soil. It is thus assumed that the high P content frequently found in chlorotic leaves is the result and not the cause for Fe chlorosis.     

Manganese Toxicity in Sugarcane Plantlets Grown on Acidic Soils of Southern China

Ratoon sugarcane plantlets in southern China have suffered a serious chlorosis problem in recent years. To reveal the causes of chlorosis, plant nutrition in chlorotic sugarcane plantlets and the role of manganese (Mn) in this condition were investigated. The study results showed that the pH of soils growing chlorotic plantlets ranged from 3.74 to 4.84. The symptoms of chlorosis were similar to those of iron (Fe) deficiency while the chlorotic and non-chlorotic plantlets contained similar amount of Fe. Chlorotic plantlets had 6.4-times more Mn in their leaf tissues compared to the control plants. There was a significantly positive correlation between Mn concentration in the leaves and the exchangeable Mn concentration in the soils. Moreover, leaf Mn concentration was related to both seasonal changes in leaf chlorophyll concentration and to the occurrence of chlorosis. Basal stalks of mature sugarcanes contained up to 564.36 mg·kg^-1 DW Mn. Excess Mn in the parent stalks resulted in a depress of chlorophyll concentration in the leaves of sugarcanes as indicated by lower chlorophyll concentration in the leaves of plantlets emerged from basal stalks. Ratoon sugarcane plantlets were susceptible to chlorosis due to high Mn accumulation in their leaves (456.90–1626.95 mg·kg^-1 DW), while in planted canes chlorosis did not occur because of low Mn accumulation (94.64–313.41mg·kg^-1 DW). On the other hand, active Fe content in chlorotic plantlets (3.39 mg kg^-1 FW) was only equivalent to 28.2% of the concentration found in the control. These results indicate that chlorosis in ratoon sugarcane plantlets results from excessive Mn accumulated in parent stalks of planted cane sugarcanes grown on excessive Mn acidic soils, while active Fe deficiency in plantlets may play a secondary role in the chlorosis.     

Effects of iron chlorosis and iron resupply on leaf xylem architecture, water relations, gas exchange and stomatal performance of field-grown peach (Prunus persica)

There is increasing evidence suggesting that iron (Fe) deficiency induces not only leaf chlorosis and a decline of photosynthesis, but also structural changes in leaf morphology, which might affect the functionality of leaves. In this study, we investigated the effects of Fe deficiency on the water relations of peach ( Prunus persica (L.) Batsch.) leaves and the responses of previously chlorotic leaves to Fe resupply via the root or the leaf. Iron deficiency induced a decline of maximum potential photosystem II (PSII) efficiency (F _V/F _M), of rates of net photosynthesis and transpiration and of water use efficiency. Iron chlorosis was associated with a reduction of leaf xylem vessel size and of leaf hydraulic conductance. In the course of the day, water potentials in chlorotic leaves remained higher (less negative) than in green leaves. In chlorotic leaves, normal stomatal functioning was disturbed, as evidenced by the lack of opening upon withdrawal of external CO₂ and stomatal closure after sudden illumination of previously darkened leaves. We conclude that the Fe deficiency induced limitations of xylem conductivity elicited a water saving strategy, which poses an additional challenge to plant growth on high pH, calcareous soils. Fertilisation with Fe improved photosynthetic performance but the proper xylem structure and water relations of leaves were not fully restored, indicating that Fe must be available at the first stages of leaf growth and development.     

Apoplastic pH and Fe3+ Reduction in Intact Sunflower Leaves

It has been hypothesized that under NO₃⁻ nutrition a high apoplastic pH in leaves depresses Fe³⁺ reductase activity and thus the subsequent Fe²⁺ transport across the plasmalemma, inducing Fe chlorosis. The apoplastic pH in young green leaves of sunflower (Helianthus annuus L.) was measured by fluorescence ratio after xylem sap infiltration. It was shown that NO₃⁻ nutrition significantly increased apoplastic pH at distinct interveinal sites (pH ≥ 6.3) and was confined to about 10% of the whole interveinal leaf apoplast. These apoplastic pH increases presumably derive from NO₃⁻/proton cotransport and are supposed to be related to growing cells of a young leaf; they were not found in the case of sole NH₄⁺ or NH₄NO₃ nutrition. Complementary to pH measurements, the formation of Fe²⁺-ferrozine from Fe³⁺-citrate was monitored in the xylem apoplast of intact leaves in the presence of buffers at different xylem apoplastic pH by means of image analysis. This analysis revealed that Fe³⁺ reduction increased with decreasing apoplastic pH, with the highest rates at around pH 5.0. In analogy to the monitoring of Fe³⁺ reduction in the leaf xylem, we suggest that under alkaline nutritional conditions at interveinal microsites of increased apoplastic pH, Fe³⁺ reduction is depressed, inducing leaf chlorosis. The apoplastic pH in the xylem vessels remained low in the still-green veins of leaves with intercostal chlorosis.     

The effect of soil moisture on the tolerance of Lupinus pilosus genotypes to a calcareous soil

Commercial narrow-leafed lupins (Lupinus angustifolius L.) grown on calcareous soils commonly display chlorotic symptoms resembling Fe deficiency. The severity of chlorosis increases with concurrent increases in soil moisture content. Our research has indicated that the rough-seeded lupin species, Lupinus pilosus Murr., has a range of adaptation to calcareous soils, from tolerant to intolerant. A pot experiment was conducted comparing a tolerant, a moderately tolerant and a moderately intolerant genotype of L. pilosus. Plants were grown for 35 days in a calcareous soil (50% CaCO₃) at three moisture contents (80%, 100% and 120% of field capacity); the growth was compared with that on a fertile black cracking clay control soil at 70% of field capacity. Visual chlorosis score, chlorophyll meter readings, number of leaves and shoot dry weights were recorded at 14, 21, 28 and 35 days after sowing. Concentrations of chlorophyll, active Fe and nutrients in the youngest fully expanded leaves were also measured. Results showed that increased soil moisture increased the severity of chlorotic symptoms (increased chlorosis score) in all genotypes. The tolerant genotype showed significantly less symptoms than other genotypes at all moisture contents. All genotypes were able to recover from chlorosis symptoms at 80% moisture in the calcareous soil. Chlorosis score negatively correlated with chlorophyll meter readings, chlorophyll concentration and foliar active and total Fe, and Mn concentrations. Visual chlorosis score appeared to be a cost effective, accurate and efficient method enabling classification of the tolerance of genotypes. The chlorotic symptoms were likely to be due to HCO₃ ^- induced nutrient deficiencies or a direct effect of HCO₃ ^- on chlorophyll synthesis. This study indicates that the most probable mechanism of tolerance is related to an ability to prevent uptake of HCO₃ ^- or efficiently sequester it once inside the root which prevents increases in internal pH and transport to the shoots.     

Differential susceptibility of two populations of Eucalyptus viminalis labill. to iron chlorosis

Summary In comparing two populations of E. viminalis observations indicated that plants of a calcareous population (i) showed a greater yield at high pH, and when subjected to Fe-stress, (ii) took longer to develop chlorosis, (iii) more quickly developed new roots, and (iv) were capable of removing more Fe from solution than were plants of an acid population. Some Fe-stressed plants also appeared to be able to reduce Fe³⁺ to Fe²⁺, but population differences have not yet been clearly established. Plants from an acidic population accumulated very high levels of P in leaves when grown in alkaline solutions and, consequently, exhibited high P/Fe ratios, chlorosis, and symptoms of P toxicity.     

Nitrate does not result in iron inactivation in the apoplast of sunflower leaves

It has been hypothesized that nitrate (NO(3)(-)) nutrition might induce iron (Fe) deficiency chlorosis by inactivation of Fe in the leaf apoplast (H.U. Kosegarten, B. Hoffmann, K. Mengel [1999] Plant Physiol 121: 1069-1079). To test this hypothesis, sunflower (Helianthus annuus L. cv Farnkasol) plants were grown in nutrient solutions supplied with various nitrogen (N) forms (NO(3)(-), NH(4)(+) and NH(4)NO(3)), with or without pH control by using pH buffers [2-(N-morpholino)ethanesulfonic acid or 4-(2-hydroxyethyl)-1-piperazineethanesulfonic acid]. It was shown that high pH in the nutrient solution restricted uptake and shoot translocation of Fe independently of N form and, therefore, induced Fe deficiency chlorosis at low Fe supply [1 micro M ferric ethylenediaminedi(O-hydroxyphenylacetic acid)]. Root NO(3)(-) supply (up to 40 mM) did not affect the relative distribution of Fe between leaf apoplast and symplast at constant low external pH of the root medium. Although perfusion of high pH-buffered solution (7.0) into the leaf apoplast restricted (59)Fe uptake rate as compared with low apoplastic solution pH (5.0 and 6.0, respectively), loading of NO(3)(-) (6 mM) showed no effect on (59)Fe uptake by the symplast of leaf cells. However, high light intensity strongly increased (59)Fe uptake, independently of apoplastic pH or of the presence of NO(3)(-) in the apoplastic solution. Finally, there are no indications in the present study that NO(3)(-) supply to roots results in the postulated inactivation of Fe in the leaf apoplast. It is concluded that NO(3)(-) nutrition results in Fe deficiency chlorosis exclusively by inhibited Fe acquisition by roots due to high pH at the root surface.     

The effect of high pH and P on the development of lime-chlorosis in two seedling populations ofEucalyptus obliqua L'Hérit

Summary Glasshouse experiments have shown that the application of an acidulating agent to a calcareous soil can increase growth and alleviate severe chlorosis in an acidic population ofE. obliqua. In contrast, a calcareous population showed only a slight response to this treatment and maintained adequate growth and a low frequency of chlorosis on both control and treated calcareous soils. Foliar analyses of seedlings of the acidic population showed that alleviation of chlorosis was concomitant with a reduction in the levels of P, Ca and K, and an increase in uptake of Fe. However, the total Fe content of foliage was poorly correlated with the occurrence of severe chlorosis. Although this evidence suggested that the differential susceptibility ofE. obliqua to lime-chlorosis can be reduced by increasing the availability of Fe, the greater concentration of Fe in chlorotic seedlings indicated that lime-chlorosis may also be due to an inactivation of Fe within the plant (i.e. by P). This hypothesis was partly confirmed by a water culture experiment which showed that a combination of relatively high pH and high external levels of P could induce severe chlorosis in seedlings of the acidic population. In contrast, it appears that the calcareous population has a more efficient mechanism for absorbing Fe and holding it in an available form, even when external concentrations of P are high. It is suggested that plants which have an efficient mechanism for the uptake of Fe at relatively high pH and are less susceptible to the detrimental effects of P have been selected for on these alkaline calcareous soils.     

Apoplastic pH and FeIII reduction in young sunflower (Helianthus annuus) roots

The relationship between the apoplastic pH in young sunflower roots ( Helianthus annuus L.) and the plasmalemma ferric chelate reductase (FC-R; EC 1.16.1.7) activity in roots was investigated. The hypothesis was tested that a high apoplastic pH depresses FC-R activity, thereby restricting the uptake of Fe²⁺ into the cytosol. Until recently, little has been known about this relationship, because pH and redox reaction measurements are difficult to perform within the confines of the root apoplast. We recorded the apoplastic pH by means of the fluorescence ratio in conjunction with video microscopy by covalently tagging fluorescein boronic acid to OH groups of the root cell wall. Fe^III reduction was measured using a similar approach by tagging ferrozine diboronic acid with OH groups of the cell wall. Ferrozine forms an Fe²⁺ complex, thus indicating the reduction of ferric iron. In roots bathing in buffered outer solutions of different pH, a high pH sensitivity of apoplastic Fe^III reduction was found, with the highest ferric iron reduction rates at an apoplastic pH of 4.9; above an apoplastic pH of 5.3, no reduction was observed. Nitrate in the bathing solution increased the apoplastic pH and hence depressed the Fe^III reduction; ammonium had the reverse effect. Nitrate together with HCO₃^–, a combination which is typical of calcareous soils, had the strongest depressing effect. From the results, it can be concluded that the main reason for the frequently occurring iron deficiency chlorosis of plants grown on calcareous soils is the inhibition of Fe^III reduction in the apoplast, and hence Fe²⁺ uptake into the cytosol.     

Physiological root responses of iron deficiency susceptible and tolerant tomato genotypes and their reciprocal F1 hybrids

By using two tomato genotypes line 227/1 (Fe chlorosis susceptible) and Roza (Fe chlorosis tolerant) and their reciprocal F₁hybrid, some root morphological changes, pH changes of nutrient solution, reduction capacity of Fe^III and uptake and root-to-shoot translocation of ⁵⁹Fe were studied under controlled environmental conditions in nutrient solution with 3 different Fe supplies as Fe EDDHA (i.e., 10^–7 M, severe Fe deficiency; 10^–6 M, intermediate Fe deficiency; 10^–4 M, adequate Fe supply). Tolerant parent `Roza' was less affected by low Fe supply than susceptible parent `line 227/1' as judged from the severity of leaf chlorosis. Under both Fe deficient conditions there were no differences between the reciprocal hybrids concerning the appearance of chlorosis. Under intermediate Fe deficiency, reciprocal F₁ hybrids (`line 227/1 × Roza' and `Roza × line' 227/1) showed an intermediate chlorosis between tolerant and susceptible parents. However, under severe Fe deficiency the reciprocal hybrids were more chlorotic than the tolerant parent irrespective of which parent was the cytoplasm contributor. A decreased Fe supply during preculture enhanced Fe^III reduction capacities of the parents and reciprocal hybrids. Differences in the tolerance to Fe deficiency always were better correlated with Fe^III reduction capacity of the genotypes than the Fe deficiency-induced release of H⁺ ions. Under both Fe deficient conditions the tolerant parent Roza had a much higher Fe^III reduction capacity than the susceptible parent line 227/1. The reduction capacity of the hybrids `Roza × line 227/1' was very similar to the capacity of the parent Roza, but higher than the capacity of the hybrids `line 227/1×Roza' at both Fe-deficient conditions. Under both Fe deficient conditions tolerant parent had higher number of lateral roots than the susceptible parent. Among the reciprocal hybrids `Roza × line 227/1' possessed more lateral roots than the `line 227/1 × Roza' under both Fe deficient conditions. Low Fe nutritional status resulted in marked increase in root uptake of ⁵⁹Fe. At adequate Fe supply, reciprocal hybrids and their parents did not differ in uptake and root-to-shoot translocation of Fe. However, under Fe-deficient conditions uptake and root-to-shoot translocation of ⁵⁹Fe were significantly higher in the Fe chlorosis tolerant than the susceptible parent. Based on the reduction capacity of Fe^III and uptake and root-to-shoot translocation of Fe, the F₁ hybrids obtained from the cross in which the maternal genotype was Roza appeared to be more tolerant than when the maternal genotype was the susceptible line 227/1. Uptake and translocation ratio of the F₁ hybrids obtained from `Roza × line 227/1' were similar to those of the parent Roza, but higher than the F<sub>1</sub> hybrids obtained from `line 227/1 × Roza', particularly under intermediate Fe deficiency. The results indicate that Fe^III reduction show a better relationship to Fe efficiency than Fe deficiency induced release of H⁺ ions. The inheritance of Fe deficiency tolerance of Roza seems not to be simple monogenic. It might be characterised by both, nuclear and extranuclear heredity. The intermediate responses of the reciprocal hybrids of the `line 227/1 × Roza' indicates that the Fe deficiency tolerance character of Roza is transferable by nuclear heredity. The better responses of the hybrids of `Roza × line 227/1' than the hybrids of `line 227/1 × Roza' may be due to maternal transmission from the parent Roza besides the nuclear transmission.     

Apoplastic pH and Fe(3+) reduction in intact sunflower leaves

It has been hypothesized that under NO(3)(-) nutrition a high apoplastic pH in leaves depresses Fe(3+) reductase activity and thus the subsequent Fe(2+) transport across the plasmalemma, inducing Fe chlorosis. The apoplastic pH in young green leaves of sunflower (Helianthus annuus L.) was measured by fluorescence ratio after xylem sap infiltration. It was shown that NO(3)(-) nutrition significantly increased apoplastic pH at distinct interveinal sites (pH >/= 6.3) and was confined to about 10% of the whole interveinal leaf apoplast. These apoplastic pH increases presumably derive from NO(3)(-)/proton cotransport and are supposed to be related to growing cells of a young leaf; they were not found in the case of sole NH(4)(+) or NH(4)NO(3) nutrition. Complementary to pH measurements, the formation of Fe(2+)-ferrozine from Fe(3+)-citrate was monitored in the xylem apoplast of intact leaves in the presence of buffers at different xylem apoplastic pH by means of image analysis. This analysis revealed that Fe(3+) reduction increased with decreasing apoplastic pH, with the highest rates at around pH 5. 0. In analogy to the monitoring of Fe(3+) reduction in the leaf xylem, we suggest that under alkaline nutritional conditions at interveinal microsites of increased apoplastic pH, Fe(3+) reduction is depressed, inducing leaf chlorosis. The apoplastic pH in the xylem vessels remained low in the still-green veins of leaves with intercostal chlorosis.     

Leaf structural changes associated with iron deficiency chlorosis in field-grown pear and peach: physiological implications

Plants grown in calcareous, high pH soils develop Fe deficiency chlorosis. While the physiological parameters of Fe-deficient leaves have been often investigated, there is a lack of information regarding structural leaf changes associated with such abiotic stress. Iron-sufficient and Fe-deficient pear and peach leaves have been studied, and differences concerning leaf epidermal and internal structure were found. Iron deficiency caused differences in the aspect of the leaf surface, which appeared less smooth in Fe-deficient than in Fe-sufficient leaves. Iron deficiency reduced the amount of soluble cuticular lipids in peach leaves, whereas it reduced the weight of the abaxial cuticle in pear leaves. In both plant species, epidermal cells were enlarged as compared to healthy leaves, whereas the size of guard cells was reduced. In chlorotic leaves, bundle sheaths were enlarged and appeared disorganized, while the mesophyll was more compacted and less porous than in green leaves. In contrast to healthy leaves, chlorotic leaves of both species showed a significant transient opening of stomata after leaf abscission (Iwanoff effect), which can be ascribed to changes found in epidermal and guard cells. Results indicate that Fe-deficiency may alter the barrier properties of the leaf surface, which can significantly affect leaf water relations, solute permeability and pest and disease resistance.     

Response of lupins (Lupinus angustifolius L.) and peas (Pisum sativum L.) to Fe deficiency induced by low concentrations of Fe in solution or by addition of HCO3 -

Lupins appear to be more sensitive than peas to Fe deficiency. However, when grown in nutrient solutions between pH 5–6, little difference existed between them in their ability to acidify the solution or to release Fe^III reducing compounds. This experiment was aimed at determining whether differences between species which occurred when Fe deficiency was induced by withholding Fe from an acid solution, are maintained when Fe deficiency is induced by addition of HCO₃ ^-. Lupins and peas were grown in nutrient solutions at 0, 2 and 6 μM of Fe^III EDDHA and either with or without HCO₃ ^- (6 mM). Bicarbonate induced symptoms of Fe deficiency (chlorosis) in both lupins and peas, and markedly decreased the growth of shoots. Symptoms appeared sooner and were more severe in lupins than in peas. Growing plants without HCO₃ ^-, but at the lowest Fe level, decreased the growth and Fe concentration of shoots of lupins but did not induce chlorosis. Growing peas in this treatment, decreased Fe concentrations, but to a lesser extent than in lupins, and did not decrease growth. H⁺-ion extrusion and release of Fe^III reducing compounds was greater in lupins than in peas. Bicarbonate also decreased the growth of roots of lupins but increased the growth of roots of peas. Results indicate that when Fe deficiency is induced by HCO₃ ^-, then the response of lupins and peas are similar to their response in acid solution culture. Differences between species therefore could not be explained by their relative abilities to acidify or release Fe^III reducing compounds. Greater control of the distribution of Fe within the shoots, the presence of a pool of Fe within the roots, a lower threshold for Fe uptake, or a higher content of seed-Fe, may therefore be the reason for the lower sensitivity of peas than lupins to Fe deficiency.     

Rhizosphere acidification and Fe3+ reduction in lupins and peas: Iron deficiency in lupins is not due to a poor ability to reduce Fe3+

While lupins suffer severely from Fe deficiency when grown on calcareous soils, field peas under the same conditions grow normally. This paper aimed to identify whether these differences were related to differences in either the pattern or capacity for rhizosphere acidification or Fe³⁺ reduction between these species. Two lupin species (Lupinus angustifolius, L. cosentinii) and field peas (Pisum sativum) were grown in solution culture for 5 weeks with both an adequate and a low supply of Fe. Plants were reliant on symbiotically fixed N. The extent of iron reduction was determined using the chelates TPTZ and BPDS. The pattern of reactions around roots was determined by placing roots in agar containing either bromocresol purple or TPTZ. The low supply of Fe decreased the growth of lupins by over 30% and induced severe chlorosis and necrosis. Growth of the peas was reduced by less than 15% and no symptoms appeared. All species acidified the solutions by about 1 pH unit regardless of the Fe treatment. The level of Fe³⁺ reduction was higher for all species grown with low Fe than with adequate Fe. Capacity for Fe³⁺ reduction was higher for all species grown with low Fe than with adequate Fe. Capacity for Fe³⁺ reduction was similar for all species. The pattern of acidification and reduction around roots was also similar between species. Thus it appears that the capacity of lupins to reduce Fe³⁺ in the rhizosphere is not the primary cause of Fe deficiency in lupins.     

Evaluation of different iron compounds in chlorotic Italian lemon trees (Citrus lemon).

The severe deficiency of iron or ferric chlorosis is a serious problem of most citrus trees established in calcareous soils, as a result of the low availability of iron in these soils and the poor uptake and limited transport of this nutrient in trees. The objective of this study was to evaluate the response of chlorotic Italian lemon trees (Citrus lemon) to the application of iron compounds to roots and stems. On comparing the effects of aqueous solutions of ferric citrate, ferrous sulphate and FeEDDHA chelate, applied to 20% of the roots grown in soil and sand, of trees that were planted in pots containing calcareous soil, it was observed that the chelate fully corrected ferric chlorosis, while citrate and sulphate did not solve the problem. EDDHA induced the root uptake of iron as well as the movement of the nutrient up to the leaves. With the use of injections of ferric solutions into the secondary stem of adult trees, ferric citrate corrected chlorosis but ferrous sulphate did not. The citrate ion expanded the mobility of iron within the plant, from the injection points up to the leaves, whereas the sulphate ion did not sufficiently improve the movement of iron towards the leaf mesophyll.     

H2O2 mediates nitrate‐induced iron chlorosis by regulating iron homeostasis in rice

The uptake of nitrate by plant roots causes a pH increment in rhizosphere and leads to iron (Fe) deficiency in rice. However, little is known about the mechanism how the nitrate uptake‐induced high rhizosphere pH causes Fe deficiency. Here, we found that rice showed severe leaf chlorosis and large amounts of Fe plaque were aggregated on the root surface and intercellular space outside the exodermis in a form of ferrihydrite under alkaline conditions. In this case, there was significantly decreased Fe concentration in shoots, and the Fe deficiency responsive genes were strongly induced in the roots. The high rhizosphere pH induced excess hydrogen peroxide (H₂O₂) production in the epidermis due to the increasing expression of NADPH‐oxidase respiratory burst oxidase homolog 1, which enhanced root oxidation ability and improved the Fe plaque formation in rhizosphere. Further, the concentrated H₂O₂ regulated the phenylpropanoid metabolism with increased lignin biosynthesis and decreased phenolics secretion, which blocked apoplast Fe mobilization efficiency. These factors coordinately repressed the Fe utilization in rhizosphere and led to Fe deficiency in rice under high pH. In conclusion, our results demonstrate that nitrate uptake‐induced rhizosphere alkalization led to Fe deficiency in rice, through H₂O₂‐dependent manners of root oxidation ability and phenylpropanoid metabolism.     

Immobilization of tissue iron on calcareous soil: differences between calcicole and calcifuge plants

Deficiency of P and sometimes of micronutrients, especially Fe, is of importance to the calcicole–calcifuge behaviour of plants. Calcifuge species are unable to solubilize these elements or keep them metabolically active in sufficient amounts on calcareous soils. To demonstrate if calcicole, calcifuge and ‘soil indifferent’ species differ in Fe nutrition dynamics, samples of such species were transplanted on a slightly acid silicate soil (pH BaCl₂ ca 4.0) and on a calcareous soil (pH BaCl₂ ca 7.2). Plants were grown in a computer‐controlled greenhouse at a soil moisture content of 50–60% water holding capacity and with additional light (ca 160 μE s^?1 m^?2, 12 h d^?1) if ambient light was <120 μE s^?1 m^?2.
The calcifuge species developed chlorosis when grown on the calcareous soil, whereas the other species did not. Calcareous‐soil grown plants had less 1,10‐phenanthroline extractable Fe in their leaf tissues than the silicate‐grown plants whereas total leaf Fe showed more species specific properties. The ratio of 1,10‐phenanthroline extractable to total Fe in the leaves was significantly lower in the calcifuges than in the calcicoles when grown on the calcareous soil. ‘Soil indifferent’ species did not differ much from the calcicoles. Root Fe, fractioned as DCB extractable ‘plaque’ on the root surface and Fe remaining in the root after DCB extraction, showed no distinct pattern of DCB‐Fe related to the different categories, but remaining root Fe tended to be lower in the calcifuges compared to the two other categories. Leaf colour estimated by a colour scale correlated well with chlorophyll a+b content measured in the leaves of two calcifuges. Leaf P concentrations did not differ between the different categories but were more species dependent.
We conclude that chlorosis in calcifuge species is related to an immobilization of Fe in physiologically less active forms in the tissue, if plants are forced to grow on a calcareous soil, whereas calcicole and ‘soil indifferent’ species are able to retain a much higher share of their leaf Fe in metabolically active form. This probably decreases the vitality and may exclude calcifuge plants from calcareous soil. We consider this property, previously almost unconsidered in an ecological context, as important to the calcifuge–calcicole behaviour of plants.