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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.
- Relationship between tipburn and leaf mineral composition in strawberryPublication . Palencia, P.; Martinez, F.; Ribeiro, E.; Correia, Maribela Pestana; Gama, Florinda; Saavedra, Teresa; de Varennes, A.; Correia, P. J.Malformation of emerging leaves with distortion of leaf tips, a condition known as tipburn, is frequently observed in strawberry. Calcium (Ca) deficiency has been considered the main cause of tipburn. The aim of the present study was to analyse the relationship between leaf mineral composition and the incidence of tipburn in three short-day strawberry (Fragaria x ananassa Duch.) cultivars ('Ventana', 'Camarosa' and 'Candonga') submitted to different concentrations of Ca. The studies were conducted in a hydroponic system in a greenhouse. Calcium was added as Ca(NO(3))(2) at 2 mM, 3 mM, 4 mM and 5 mM. A completely randomized block design (4 Ca concentrations x 3 cultivars) with three replications was used. Each replicate consisted of 12 plants grown in a polyethylene bag (100 cm x 18 cm x 3 cm) filled with coconut peat. Crown diameter and tipburn incidence were evaluated throughout the experimental period, and at the end of the experiment leaf mineral composition was assessed. In general, plants with larger crown diameters had a greater incidence of tipburn. The 'Candonga' cultivar had the smallest incidence of tipburn, while the 'Camarosa' and 'Ventana' cultivars were more susceptible. There was no correlation between level of Ca applied and incidence of tipburn. The incidence of tipburn was associated with foliar K:Ca and K:Mg ratios. Ratios above 3.40 for K:Mg and 1.77 for K:Ca represented a risk of more than 50% of tipburn incidence, when overall means for all cultivars and levels of Ca were used. (C) 2010 Elsevier B.V. All rights reserved.
- Contributos para o estudo da clorose férricaPublication . Saavedra, Teresa; Correia, Maribela PestanaO Relatório de Atividade Profissional agora apresentado reflete o meu percurso profissional desde 2007, no qual exerci sempre funções relacionadas com a minha área de formação, Hortofruticultura e noutras áreas complementares face às técnicas laboratoriais que aprendi. Neste âmbito, o tema de trabalho escolhido ‐Contributos para o estudo da clorose férricadestaca a importância do ferro (Fe), que apesar de ser necessário em pequenas quantidades pelas plantas, a incidência de clorose férrica (deficiência de Fe) é comum em muitas espécies agrícolas sendo necessário recorrer à aplicação ao solo de quelatos de Fe sintéticos. Neste capítulo, faço um breve enquadramento teórico sobre o que se investiga nesta área e toda a problemática da clorose férrica, indico os ensaios e respetivas metodologias em que estive envolvida e apresento, de forma resumida, os resultados obtidos em diversos ensaios de caraterização deste desequilíbrio nutricional e de estudo de novas alternativas para a correção da clorose férrica. Em todos os ensaios, conduzidos em sistema hidropónico em que participei, os sintomas foram induzidos pela ausência do Fe na solução nutritiva e os resultados comparados com um tratamento controlo com Fe. O grau de clorose e a recuperação dos sintomas foram estimados através dos valores de SPAD. A atividade da quelato de Fe(III)‐redutase (QF‐R), enzima responsável pela redução do Fe nas raízes, foi determinada nos ápices radiculares pela quantificação colorimétrica do complexo Fe (II) ‐BPDS. Executei diversos ensaios onde determinei a qualidade, interna e externa, dos frutos. Ainda colaborei na determinação da composição mineral de diverso material vegetal. Participei na validação de um extrato vegetal preparado a partir de aparas de relva (conforme descrição detalhada na patente PT/103584‐2009 da UALG e na patente internacional PCT/PT2007/000041‐2008; em copropriedade entre a UALG e a empresa ADP‐Adubos de Portugal S.A.) e que foi eficaz na recuperação dos sintomas de clorose férrica e que poderá vir a ser alternativa ao uso dos quelatos férricos sintéticos. Os trabalhos que desenvolvi no âmbito da minha atividade profissional abriram novas perspetivas de estudo da clorose férrica e introduziram melhorias nas técnicas de fertilização de fruteiras.
- 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.
- Relationships between strawberry fruit quality attributes and crop loadPublication . Correia, P. J.; Correia, Maribela Pestana; Martinez, F.; Ribeiro, E.; Gama, Florinda; Saavedra, Teresa; Palencia, P.Crop load can influence fruit quality in several horticultural species. The aim of the present study was to determine the effect of different concentrations of calcium on crop quality traits in three short-day strawberry (Fragaria x ananassa Duch.) cultivars ('Ventana','Camarosa' and 'Candonga') and to assess the relationships between crop load and quality parameters. The studies were conducted using a hydroponic system in a greenhouse. Calcium was added as Ca(NO3)(2) at 2 mM, 3 mM, 4 mM and 5 mM. A completely randomized block design (4 Ca concentrations x 3 cultivars) with three replicates was used. Each replicate consisted of 12 plants grown in polyethylene bags (100 cm x 18 cm x 3 cm) filled with coconut peat. Titratable acidity, total soluble solids and firmness were measured throughout the experimental period. Calcium application had no effect on fruit quality attributes but the genotype effect was clear. At the end of the experiment (28th May, 2008), titratable acidity was positively related to the fresh weight of above-ground biomass and number of leaves respectively in the 'Ventana' and 'Camarosa' cultivars. Higher values of total soluble solids were found at low crop load in 'Ventana' but in 'Camarosa' this relation was not found. In 'Candonga', higher total soluble solids were linked to crop load. In 'Ventana', titratable acidity significantly decreased as crop load increased, and in 'Camarosa' high values of titratable acidity were found at different values of crop load. 'Ventana' seemed to be more sensitive to the effects of crop load patterns. Genotype was an important factor in determining fruit quality parameters. (C) 2011 Elsevier B.V. All rights reserved.
- Evaluation of Fe deficiency effects on strawberry fruit qualityPublication . Gama, Florinda; Saavedra, T.; Miguel, Maria Graça; Dandlen, S. A.The effects of Fe deficiency on the antioxidant properties of strawberry juice was carried out with a day-neutral cultivar 'Selva'. Bare root transplants (without leaves) with approximately 18 cm, were transferred to Hoagland's nutrient solution with (Fe2.5) and without Fe (Fe0), using Fe-EDDHMA as the Fe source: 0 and 2.5 5 μM Fe. Plants were grown in 20 L containers in a glasshouse for 6 weeks (from April 27 to June 5) under natural light and air temperature ≤ 25°C. Twelve transplants were used per treatment, distributed in a complete randomized design. Plants grown in absence of Fe revealed chlorotic symptoms approximately after three weeks, based on SPAD values measured in young leaves (<20). The other treatments did not show any symptoms during the experiment. Fruits were harvested from each treatment, and juice was analysed for antioxidant activity by using the free radical α-α-diphenyl-β-picrylhydrazyl capacity (DPPH•), the Trolox equivalent antioxidant capacity (TEAC) and oxygen radical absorbance capacity (ORAC) assays. In addition, fruits were analysed for total phenols and some organic acids. The phenolic content varied between 1251 and 1514 mg gallic acid equivalents (GAE) L-1 juice, respectively, in Fe0 and Fe2.5 treatments, but with no significant differences. Despite the same total soluble solids values in both treatments, it was found that Fe depletion reduced significantly the anthocyanins and total phenols of the fruits. However, ascorbic acid increased as well as antioxidant activity expressed by both DPPH and TEAC methods.
- Root ferric chelate reductase is regulated by iron and copper in straberry plantsPublication . Pestana, M.; Correia, P. J.; Saavedra, Teresa; Gama, Florinda; Dandlen, S.; Nolasco, Gustavo; Varennes, Amarilis deIn the present experiment, we studied the interaction between copper (Cu) and iron (Fe) in strawberry plants grown in nutrient solutions containing different concentrations of Fe. Plants grown in the absence of iron (Fe0) had the characteristic symptoms of Fe deficiency, with smaller chlorotic leaves, less biomass, acidification of the nutrient solution, and roots that were smaller and less ramified, while no symptoms of Fe deficiency were observed in plants grown with Fe. A greater amount of Cu was found in roots of chlorotic plants than in those grown with Fe, while plants grown with 20M of Fe (Fe20) in the nutrient solution had a greater amount of Fe compared with plants from the other treatments. Chlorotic plants (Fe0) and plants grown with the greatest level of Fe (Fe20) had a greater root ferric chelate reductase (FC-R; EC 1.16.1.17) activity compared with the other treatments with 5 or 10M Fe in the nutrient solution. The same pattern was obtained for relative FC-R mRNA concentration and for the sum of Fe and Cu contents in shoots (leaves plus crowns). The DNA obtained from amplification of the FC-R mRNA was cloned and several of the inserts analysed by single strand confirmation polymorphism (SSCP). Although there were different SSCP patterns in the Fe20 treatment, all the inserts that were sequenced were very similar, excluding the hypothesis of more than one FC-R mRNA species being present. The results suggest that Cu as well as Fe is involved in FC-R expression and activity, although the mechanism involved in this regulation is unknown so far. Both small contents of Fe and Cu in plants led to an over-expression of the FC-R gene and enhanced FC-R activity in strawberry roots.
- A novel plant extract as a biostimulant to recover strawberry plants from iron chlorosisPublication . Saavedra, Teresa; Gama, Florinda; Correia, P.J.; Da Silva, José Paulo; Miguel, Maria da Graça; de Varennes, Amarilis; Pestana, MaribelaThe traditional agricultural production systems are evolving more towardorganic, sustainable or environmental friendly systems and products; how-ever, it is important to keep the yield and the quality of crops. The aim ofthis work was to evaluate the recovery of iron deficient strawberry plantsby foliar spray, using a gramineous plants extract (GE). Bare-root trans-plants of strawberry (Fragaria ananassaDuch. cv“Diamond”) were grownin a Hoagland’s nutrient solution with iron (Fe10) or without (Fe0). Forty-two days after Fe deprivation, plants grown without iron became chloroticand GE was applied. Three foliar applications were done, once per weekand the regreening was monitored using a SPAD apparatus. After twoapplications, a rapid but transient regreening was detected in youngleaves. Nevertheless, biomass and root/shoot ratio of recovered plants (GEtreatment) were similar to chlorotic plants (Fe0 treatment). Phosphorous,Mn and Fe contents were higher in crowns of GE plants at the end of theexperiment (67 days). The concentration of organic acids and ferric chela-te–reductase (FC-R) activity was also significantly higher in GE treatment. Itis possible that the high chelating capacity of the extract was responsible or the Fe internal transport and use.
- 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.
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