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- Effects of foliar application of organic acids on strawberry plantsPublication . Saavedra, Teresa; Gama, Florinda; Rodrigues, Maria Joao; Abadía, Javier; de Varennes, Amarilis; Pestana, Maribela; Da Silva, José Paulo; Correia, Pedro JoséThe large economic costs and environmental impacts of iron-chelate treatments has led to the search for alter-native methods and compounds to control iron (Fe) deficiency chlorosis. Strawberry plants (Fragaria x ananassa) were grown in Hoagland's nutrient solution in a greenhouse with two levels of Fe: 0 and 10 mu M Fe(III)-EDDHA. After 20 days, plants growing without Fe showed typical symptoms of Fe deficiency chlorosis in young leaves. Then, the adaxial and abaxial sides of one mature or one young leaf in each plant were brushed with 10 mM malic (MA), citric (CA) or succinic (SA) acids. Eight applications were done over a two-week period. At the end of the experiment, the newly emerged (therefore untreated), young and mature leaves were sampled for nutritional and metabolomic analysis, to assess the effectiveness of treatments. Leaf regreening was monitored using a SPAD-502 apparatus, and the activity of the ferric chelate-reductase activity (FCR) was measured using root tips. Iron deficiency negatively affected biomass and leaf chlorophyll but did not increase FCR activity. Application of succinic acid alleviated the decrease in chlorophyll observed in other treatments, and the overall nutritional balance in the plant was also changed. The concentrations of two quinic acid derivatives increased under Fe deficiency and decreased in plants treated with succinic acid, and thus they are proposed as Fe stress markers. Data suggest that foliage treatments with carboxylates may be, in some cases, environmentally friendly alter-natives to Fe(III)-chelates. The importance of Fe mobilization pathways in the formulation of new fertilizers is also discussed.
- Can bicarbonate enhance the performance of carob seedlings grown in nutrient solutions with different Fe concentrations?Publication . Gama, Florinda; Correia, Pedro José; Saavedra, T.; Dandlen, Susana; de Varennes, Amarilis; Nolasco, Gustavo; Pestana, MaribelaThe aim of this work was to assess the effect of bicarbonate (Bic) ion on the nutritional status and performance of carob-tree seedlings, a species that normally grows in calcareous soil without exhibiting iron chlorosis symptoms. Seedlings were previously grown in nutrient solution with a small concentration of Fe (0.5-1 mu M) to induce a moderate chlorosis. Afterwards, two experiments were established: in experiment 1, plants were grown for 21 days in the following treatments: Fe deficiency (Fe0), 0.5 mu M Fe, 5 mu M Fe, and 5 mu M Fe plus calcium carbonate (CaCO3). After assessing these results, a second experiment was conducted for 91 days, with the following treatments: Fe0, 1 mu M Fe, 40 mu M Fe and 40 mu M Fe plus CaCO3 and sodium bicarbonate (NaHCO3). Chlorophyll of young leaves, biomass and mineral composition of leaves, stems and roots were assessed in both experiments. The ferric chelate reductase root activity (FC-R) and the genetic expression of calmodulin-regulated Ca2+-ATPase pump (ACA gene) were evaluated in experiment 2. Fe-deficient plants exhibited reduced growth and enhanced macronutrients in leaves. Root micronutrient homeostasis changed as an adaptive mechanism in carob. The addition of bicarbonate did not aggravate Fe chlorosis, as leaf chlorophyll increased significantly. Root FC-R activity and ACA gene expression was not enhanced under Fe deficiency induced by bicarbonate (Fe40 + BicNa) which suggest a positive effect of bicarbonate in the metabolism of this crop. Nevertheless, small Fe concentrations (Fe1) induced a higher ACA gene expression thus indicating some stress response signalling.
- Silencing of FRO1 gene affects iron homeostasis and nutrient balance in tomato plantsPublication . Gama, Florinda; Saavedra, Teresa; Dandlen, Susana; García‐Caparrós, Pedro; de Varennes, Amarilis; Nolasco, Gustavo; Correia, Pedro José; Pestana, MaribelaBackgroundIron chlorosis is an abiotic stress of worldwide importance affecting several agronomic crops. It is important to understand how plants maintain nutrient homeostasis under Fe deficiency and recovery. AimsWe used the virus-induced gene silencing (VIGS) method to elucidate the role of the FRO1 gene in tomato plants and identify the impact on regulation of the root ferric-chelate reductase (FCR) activity and nutritional homeostasis. MethodsTomato plantlets cv. "Cherry" were transferred into half-strength Hoagland's nutrient solution containing 0.5 & mu;M of Fe (Fe0.5). In phase I, two treatments were established: control (Fe0.5) plants and VIGS-0.5 plants corresponding to plants with the FRO1 gene silenced. In phase II, plants from Fe0.5 and VIGS-0.5 were transferred to new nutrient solution and then grown for a further 14 days under 0 and 10 & mu;M of Fe (as 0.5 & mu;M would not be enough for the larger plants during phase II). Therefore, four treatments were imposed: Fe0, Fe10, VIGS-0, and VIGS-10. ResultsVIGS-0.5 plants had significantly lower chlorophyll (Chl) and root FCR activity compared to the respective non-silenced plants and retained more Cu and Zn in the roots at the expense of stems (Cu) or young leaves (Zn). Iron concentration in roots and stems decreased in FRO1 gene-silenced plants, compared to control plants, but the allocation to different organs was similar in both treatments. ConclusionsThere was a partial recovery of leaf Chl in the VIGS-10 plants and a higher concentration of Fe in all organs. In contrast, the allocation of Cu to roots decreased in the VIGS-10 plants.
- Is there a relationship between ferric-chelate reductase activity in roots of poncirus trifoliata and leaf chlorophyll contents?Publication . Correia, Pedro José; Lopes, Dora; Duarte, Amilcar; Gama, Florinda; Saavedra, Teresa; Pestana, MaribelaPoncirus trifoliata is a citrus rootstock very sensitive to Fe deficiency. This deficiency is very common in crops grown in calcareous soils due to the detrimental effect of bicarbonate ion. Higher plants have distinct behaviours when faced with Fe chlorosis, and several mechanisms may be activated under Fe shortage. The aim of this study was to investigate the activity of ferric-chelate reductase (FC-R), a key enzyme in Fe uptake, and to verify whether relationships with leaf chlorophyll contents could be established. Plants were grown in nutrient solutions without Fe (0 μM Fe), with 1 μM Fe, with 120 μM Fe and with 120 μM Fe plus CaCO3 (1 g L-1). Total leaf chlorophyll in young and mature leaves was determined using a calibration model based on a relationship between SPAD readings and concentration of chlorophyll (r2=0.95; P<0.01). The activity of FC-R was determined in roots apexes and several biomass parameters in shoots (number of leaves, height, dry and fresh weight) and roots (dry and fresh weight) were evaluated at the end of the experiment. The activity of FC-R increased in plants grown without iron (0 μM Fe). The results about the relations between root FC-R and leaf chlorophyll are discussed.
- Nutritional performance of Five Citrus rootstocks under different Fe levelsPublication . Pestana, Maribela; García-Caparrós, Pedro; Saavedra, Teresa; Gama, Florinda; Abadía, Javier; Varennes, Amarilis de; Correia, Pedro JoséIron is an essential micronutrient for citrus, playing an important role in photosynthesis and yield. The aim of this paper was to evaluate the tolerance to Fe deficiency of five citrus rootstocks: sour orange (S), Carrizo citrange (C), Citrus macrophylla (M), Troyer citrange (T), and Volkamer lemon (V). Plants were grown for 5 weeks in nutrient solution that contained the following Fe concentrations (in µM): 0, 5, 10, 15, and 20. At the end of the experiment, biomass (dry weight— DW), leaf area, total leaf chlorophyll (CHL), and the activity of root chelate reductase (FCR) were recorded. Additionally, the mineral composition of roots (R) and shoots (S) was evaluated. Principal component analysis was used to study the relationships between all parameters and, subsequently, the relations between rootstocks. In the first component, N-S, P-S, Ca-S, Cu-S, Zn-S, Mn-S, Zn-R, and Mn-R concentrations were related to leaf CHL and FCR. Increases in leaf CHL, Mg-R, and DW (shoots and roots) were inversely related to Cu-R, which was shown in the second component. The values obtained were consistent for V10, C15, and C20, but in contrast for S0 and S5. In conclusion, micronutrient homeostasis in roots and shoots of all rootstocks were affected by Fe stress conditions. The Fe/Cu ratio was significantly related to CHL, which may be used to assist rootstock performance.
- Silencing of the FRO1 gene and its effects on iron partition in Nicotiana benthamianaPublication . Gama, Florinda; Saavedra, Teresa; Dandlen, Susana; de Varennes, Amarilis; Correia, Pedro José; Pestana, Maribela; Nolasco, GustavoTo evaluate the dynamic role of the ferric-chelate reductase enzyme (FCR) and to identify possible pathways of regulation of its activity in different plant organs an investigation was conducted by virus induced gene silencing (VIGS) using tobacco rattle virus (TRV) to silence the ferric reductase oxidase gene (FROI) that encodes the FCR enzyme. Half of Nicotiana benthamiana plants received the VIGS vector and the rest remained as control. Four treatments were imposed: two levels of Fe in the nutrient solution (0 or 2.5 mu M of Fe), each one with silenced or non-silenced (VIGS-0; VIGS-2.5) plants. Plants grown without iron (0; VIGS-0) developed typical symptoms of iron deficiency in the youngest leaves. To prove that FROI silencing had occurred, resupply of Fe (R) was done by adding 2.5 mu M of Fe to the nutrient solution in a subset of chlorotic plants (0-R; VIGS-R). Twelve days after resupply, 0-R plants had recovered from Fe deficiency while plants containing the VIGS vector (VIGS-R) remained chlorotic and both FRO1 gene expression and FCR activity were considerably reduced, consequently preventing Fe uptake. With the VIGS technique we were able to silence the FRO1 gene in N. benthamiana and point out its importance in chlorophyll synthesis and Fe partition. (C) 2017 Elsevier Masson SAS. All rights reserved.
- Responses of tomato (Solanum lycopersicum L.) plants to iron deficiency in the root zonePublication . Jiménez, María Rocío; Casanova, Laura; Saavedra, Teresa; Gama, Florinda; Suárez, María Paz; Correia, Pedro José; Pestana, MaribelaIron deficiency induces a yellowing in the aerial part of plants, known as iron chlorosis, and reduces the growth, yield, and quality of the fruits. Understanding plant response to iron deficiency is essential for agronomic management. This study decoded the temporal response of tomato plants (Solanum lycopersicum L.) to iron deficiency by quantifying different vegetative parameters. Subapical root swelling in the first 2.0 mm and several shoot and root growth parameters were measured in plants grown in a nutrient solution with and without Fe, on different dates designated as days after transplantation (DAT). Correlations between the total chlorophyll concentration in young leaves and 22 morphological and physiological parameters were also calculated. The plants grown in the absence of Fe had a higher number of secondary roots at 3 DAT, compared to control plants. On the same date, subapical root swelling was also observed, particularly at 1.5 and 2.0 mm from the root tip. Those plants also had a lower chlorophyll content in young leaves and a higher ferric-chelate reductase activity (FCR; EC 1.16.1.17) in the roots. At 9 DAT, the overall vegetative performance (plant height, fresh weight of stems and leaves) was negatively affected. At the end of the experiment (14 DAT), significant correlations were found between chlorophyll and the studied parameters. In conclusion, tomato plants experienced a cascade of responses to Fe deficiency throughout nine days: firstly, root lateralization increased; later, root swelling was observed, and a decrease in leaf chlorophyll content was registered associated with an increase in root FCR. At the end, the biomass of tomato plants decreased.
- Changes in nutritional homeostasis of Poncirus trifoliata and Ceratonia siliquaas a response to different iron levels in nutrient solutionPublication . Correia, Pedro José; de Varennes, Amarilis; Gama, Florinda; Saavedra, Teresa; Pestana, MaribelaIron (Fe) deficiency is a nutritional disorder in plants. Poncirus trifoliata is susceptible to Fe deficiency, but symptoms of Fe deficiency are rare in Ceratonia siliqua, a slow-growing species. Specimens of the two species were grown in nutrient solutions containing three Fe concentrations: without Fe (0 mM), 1 mM Fe, and either 10 mM Fe (for Ceratonia) or 40 mM Fe (for P. trifoliata). Growth, the degree of chlorosis, the plant mineral composition, and the activity of the root ferric chelate-reductase (FCR) were assessed. Ceratonia plants exposed to 1 mM Fe were efficient at using Fe in the synthesis of chlorophyll. The activity of FCR was enhanced in the total absence of Fe. In Poncirus a low activity of the FCR was observed in plants with no Fe. The balance between micronutrients in the Ceratonia roots was not affected with 1 mM Fe compared with the higher Fe concentration treatments.
- Brief characterisation of Fe chlorosis in chia (Salvia hispanica L.) plants grown in nutrient solutionPublication . Vieira, João; Džidić Uzelac, Luna; Thais Fidelis Alves, Izabela; Patrício, Marco; Gama, Florinda; Saavedra, Teresa; Correia, Pedro José; Pestana, MaribelaChia (Salvia hispanica L.) plant is a well-known plant due to the nutraceutical value of its seeds. The aim of this preliminary study was to assess the response of Chia plants to Fe deficiency. Chia plants were grown for 12 days in Hoagland's nutrient solutions without Fe (Fe0) and with Fe (Fe10-10 mu M Fe). Biomass parameters and root ferric chelate-reductase activity (FC-R; EC 1.16.1.17) were determined at the end of the experiment. Chlorophyll estimations (expressed as SPAD readings) decreased progressively, showing the typical symptoms of iron chlorosis. In addition, iron-deficient chia plants exhibit smaller biomass (number of leaves, root, and shoot growth reduction) compared to control plants. These plants also showed morphological changes in roots. Furthermore, root FC-R activity was significantly lower in Fe0 plants.