Fe deficiency-induced chlorosis is a frequent problem in calcareous soils and represents a severe limitation to the productivity of many crops in the Mediterranean regions.Activation of plant; response mechanisms and the presence of humic substances may in part minimize or prevent the effects of this micronutritional disorder. These aspects were investigated by growing cucumber plants in nutrient solutions in the presence or absence of Fe-EDTA at pH 6.0 (unbuffered) or pH 7.2 (buffered with 10 mM MES-NaOH or 1 g.L(-1) CaCO(3)). Fe-deficient plants developed a physiological response typical of strategy I species (increased Fe(III)-chelate reductase activity and acidifying capacity of roots) which proved to be less efficient at high pH values and in the presence of CaCO(3). To evaluate if a water soluble humic fraction (WEHS) was a suitable source of Fe(III), 10-day-oId Fe-deficient plants were supplied daily with Fe-WEHS (0.1 mu M Fe and 2.5 mu g organic C.mL(-1)) or 0.1 mu M Fe-EDTA for a period of 5 days. Fe-WEHS was more effective than Fe-EDTA in inducing the recovery of chlorophyll content and visible re-greening of the leaves. Plants supplied with Fe-complexes had a higher Fe(III)-chelate reductase activity than Fe starved plants, in particular when the latter were treated with CaCO(3) or MES-NaOH (pH 7.2). Results show that the presence of CaCO(3) and WEHS can be important factors in determining Fe acquisition from the soil and should be taken into account when studying Fe deficiency under natural conditions.

The ability of Fe-deficient cucumber plants to use iron complexed to a water-extractable humic substances fraction (WEHS), was investigated. Seven-day-old Fe-deficient plants were transferred to a nutrient solution supplemented daily for 5 days with 0.2 μM Fe as Fe-WEHS (5 μg org. C mL-1), Fe-EDTA, Fe-citrate or FeCl3. These treatments all allowed re-greening of the leaf tissue, and partial recovery of dry matter accumulation, chlorophyll and iron contents. However, the recovery was faster in plants supplied with Fe-WEHS and was already evident 48 h after Fe supply. The addition of 0.2 μM Fe to the nutrient solution caused also a partial recovery of the dry matter and iron accumulation in roots of Fe-deficient cucumber plants, particularly in those supplied with Fe-WEHS. The addition of WEHS alone (5 μg org. C mL-1, 0.04 μM Fe) to the nutrient solution slightly but significantly increased iron and chlorophyll contents in leaves of Fe-deficient plants; in these plants, dry matter accumulation in leaves and roots was comparable or even higher than that measured in plants treated with Fe-citrate or FeCl3. After addition of the different iron sources for 5 days to Fe-deficient roots, morphological modifications (proliferation of lateral roots, increase in the diameter of the sub-apical zones and amplified root-hair formation) and physiological responses (enhanced Fe(III)-chelate reductase and acidification of the nutrient solution) induced by Fe deficiency, were still evident, particularly in plants treated with the humic molecules. The presence of WEHS caused also a further acidification of the nutrient medium by Fe-deficient plants. The Fe-WEHS complex (1 μM Fe) could be reduced by intact cucumber roots, at rates of reduction higher than those measured for Fe-EDTA at equimolar iron concentration. Plasma membrane vesicles, purified by two-phase partition from root microsomes of Fe-deficient plants, were also able to reduce Fe-WEHS. Results show that Fe-deficient cucumber plants can use iron complexed to water soluble humic substances, at least in part via reduction of complexed Fe(III) by the plasma membrane Fe(III)-chelate reductase of root cells. In addition, the stimulating effect of humic substances on H+ release might be of relevance for the overall response of the plants to iron shortage.