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- Field performance of seedlings and microproppagated plants of carob treePublication . Osório, Maria Leonor; Osório, Júlio; David, Maria Manuela; Romano, AnabelaMicropropagated plantlets of cv. Mulata and cv. Galhosa growing in pots, after acclimatization in the glasshouse and growth for several months under natural conditions, were transferred to a field trial. Carob seedlings of ‘Mulata’ were also transferred under the same conditions. The field trial has been established with 100% of success in micropropagated plants and 97% in seedlings. Three months after transfer to the field, plants showed good growth but micropropagated plants of ‘Mulata’ exhibited more vigour than ‘Galhosa’ and than seedlings, with the greatest number of branches and a larger length increase in the main stem. Before transference to the field, net photosynthetic rates (PN), water use efficiency (PN/gs) and quantum yield of PSII (fPSII) of the potted plants were higher in seedlings than in micropropagated plants. Three months after transplantation all gas exchange parameters were improved for all types of plants, particularly net photosynthetic rate, and no significant differences were observed between plants either micropropagated or seedlings. On the other hand, fPSII decreased significantly in seedlings. The results suggest that the pre-acclimation, in pots, to external environmental conditions might have improved field survival, plant vigour and enhanced the endurance to adverse conditions of micropropagated carob trees.
- The effects of drought on net CO2 uptake, chlorophyll fluorescence and photosynthetic pigments in mature and young leaves of sunflowerPublication . Osório, Maria Leonor; Barrote, Isabel; Osório, Júlio; David, Maria Manuela; Correia, Maria JoãoPhotosynthesis, photosystem II activity and photosynthetic pigments were analysed in mature and Young leaves of sunflower (Helianthus annuus L.) during dehydration and rehydration, in order to determine the influence of leaf age on the sensitivity to drought. Dehydration caused a more pronounced inhibition in photosynthetic rates in mature leaves compared to younger leaves, but 24 h after rewatering the recovery was complete in spite of leaf age. After exposure to drought, mature leaves showed a larger reduction in the actual PSII photochemical efficiency and a higher increase in nonradiative energy dissipation than young leaves. In well-watered plants the de-epoxidation level represented by the ratio A+Z/V+A+Z was higher in young leaves. However drought had opposite effects on both type of leaves: in young leaves A+Z/V+A+Z decreased with water deficit, while in mature leaves increased with soil drying. The results obtained suggest that leaf age influence the “strategy” to avoid irreversible damage of PSII activity under drought and that older leaves are more drought-sensitive than younger leaves of sunflower.
- Effects of water deficit on the activity of nitrate reductase and content of sugars, nitrate and free amino acids in the leaves and roots of sunflower and white lupin plants growing under two nutrient supply regimesPublication . Correia, Maria João; Fonseca, Filomena; Azedo-Silva, João; Dias, Celeste; David, Maria Manuela; Barrote, Isabel; Osório, Maria Leonor; Osório, JúlioThe effects of a two weeks soil drying period on the activity of nitrate reductase (NR; EC 1.6.6.6) were studied on Helianthus annuus L. and non-nodulated Lupinus albus L. plants, growing under two nutrient supply regimes. NR activity was assessed in leaf and root extracts, by measuring the activity of the unphosphorylated active form (NRact), the maximal extractable activity (NRmax) and the activation state. To get insight into potential signalling compounds, nitrate, amino acids and soluble sugars concentrations were also quantified. On both species, foliar NRact and NRmax were negatively affected by soil drying and reduced supply of nutrients, the observed changes in NR activity being linearly-correlated with the depletion of nitrate. Similar results were obtained in the roots of sunflower. Conversely, in white lupin roots NRmax was found to be independent of tissue nitrate concentration. Regardless of the species and organ, the activation state of the enzyme was unaffected by the nutrient supply regime. In well-watered sunflower roots only about 50% of the existing NR was unphosphorylated, but the activation state increased significantly in response to drought. In contrast, lupin roots always exhibited NR activation state values close to 80% or even higher. At the leaf level, NR activation state was hardly changed in response to soil drying. The contribution of changes in the concentrations of soluble 5 sugars and amino acids to explain the observed variations in NR activity are discussed.
- Photosynthetic responses of Lupinus albus to soil water fluctuationsPublication . Barrote, Isabel; Osório, Maria Leonor; Osório, Júlio; David, Maria Manuela; Correia, Maria JoãoLupinus albus plants were grown in 3 dm3 pots in a semi-controled greenhouse. Two water regimes were imposed: water deficit (S, 47% of soil capacity), and control (T, 86% of soil capacity). Plants water status was monitored through foliar relative water content (TRA) and pre-dawn (ypd) and midday water potential (ymd). Gas exchanges, chlorophyll a fluorescence and photosynthetic capacity (Amax) were measured. Pigments and soluble protein were quantified and antioxidant system enzymes activity was determined. Plants under water deficit were rehydrated and the same measurements and sampling were done again after 48 h (R1) and 72 h (R2). Albeit water availability has diminished from 86% to 47% of soil capacity, there weren’t any remarkable changes on S plants. R1 plants which haven’t shown any alterations neither in Amax nor in stomatal conductance (gH2O), exhibited a significant decrease on net photosynthesis (A), reaching negative values, along with a raise in non-photochemical quenching (NPQ). Presumably this raise should be related with an increase in the xanthophyll cycle deepoxidation state. The significantly increase in Superoxide dismutase (SOD) specific activity in R1 and R2 plants might be related with the activation of Mehler-peroxidase reaction which provokes lumen DpH raising. This can provide photosynthesis protection through VAZ cycle and may justify the NPQ increase.
- Influence of transient shade periods on the effects of drought on photosynthesis, carbohydrate accumulation and lipid peroxidation in sunflower leavesPublication . Correia, Maria João; Osório, Maria Leonor; Osório, Júlio; Barrote, Isabel; Martins, Mónica; David, Maria ManuelaThe effects of a slow-imposing two-weeks soil drying period, and subsequent re-watering, on leaf water potential (Ψ), gas exchange rates, chlorophyll fluorescence and on the concentrations of malondialdehyde (MDA) and non-structural carbohydrates (starch, hexose and sucrose) were determined in mature leaves of sunflower plants growing under controlled environmental conditions. To assess how transient shade periods, associated with increased cloud cover, may influence drought-induced effects on carbon assimilation, measurements were carried out both in plants kept under the photosynthetic photon flux density (PPFD) prevailing during the growth period and stress imposition (750 μmol m-2 s-1), and in plants subjected to a 5 hours-long period under a lower PPFD (200 μmol m-2 s-1). In plants kept under high PPFD, Ψ, stomatal conductance (g), net CO2 uptake rate (A), the quantum yield of photosystem II electron transport (Φe), the photochemical efficiency of open PSII reaction centres (Fv’/Fm’) and * Corresponding author. Tel: +351 289800932; fax: +351 289818419. E-mail address: jcorreia@ualg.pt (M.J. Correia) 1 the diurnal accumulation of total non-structural carbohydrates (TNC) were significantly depressed at the end of the soil drying period, whereas non-photochemical quenching (NPQ), the concentrations of MDA and the predawn pools of soluble sugars were found to increase. Under high-light level, drought-induced effects on lipid peroxidation, chlorophyll fluorescence parameters and gas exchange rates were fully reversed upon re-watering. However, the inhibition of diurnal accumulation of TNC still persisted two days following re-watering, suggesting that carbohydrate export rates were enhanced following stress relief. An overall positive effect was found upon transferring water-stressed plants to low light level, as indicated by the increases in Ψ, intrinsic water use efficiency (A/g), Φe and Fv’/Fm’, as well as the reversal of drought-induced enhancement of both NPQ and MDA concentration. Despite g being similar in shaded well-watered and re-watered plants, the latter exhibited net CO2 uptake rates below those found in well-watered leaves under the same light conditions, together with a diurnal decrease in the concentration of TNC that was mainly attributable to the depletion of starch and sucrose. These results indicate that contrasting with the positive effects of shading on water-stressed plants, low PPFD may negatively affect the recovery of net photosynthesis following stress relief.
- Carob trees (Ceratonia siliqua L.) regenerated in vitro can acclimatize successfully to match the field performance of seed-derived plantsPublication . Osório, Maria Leonor; Osório, Júlio; Gonçalves, Sandra; David, Maria Manuela; Romano, Anabela; Correia, Maria JoãoThe use of in vitro regenerated plants in forestry and orchard depends ultimately on the development of efficient transplantation protocols, ensuring high survival rates and successful establishment under field conditions. We tested the performance of micropropagated carob trees (Ceratonia siliqua L.) throughout the acclimatization process in terms of survival, growth and physiological traits, including field comparisons with seed-derived and mother plants. The field trial was 100 %successful, i.e. we found no major differences between micropropagated, seed-derived and mother plants in terms of growth rate, height, number of leaves, photosynthetic efficiency, chlorophyll fluorescence, chlorophyll content and soluble protein content, although these parameters changed significantly during acclimatization. Stomatal conductance (gs) was reduced by fourfold when plants were transferred from in vitro culture to the growth chamber, thus preventing uncontrolled wilting. The photosynthetic rate (PN) was relatively low in vitro, in the growth chamber and the greenhouse, but increased to match seed-derived and mother plants in the field. The chlorophyll a/b ratio in leaves from in vitro and growth chamber plants was typical of shade plants (2.1) but became more characteristic of sun plants in the subsequent acclimatization stages (3.1–3.5). The maximum efficiency of photosystem II (Fv/Fm) remained mostly constant at ~0.80 throughout acclimatization, as is typical for healthy, non-stressed plants. We conclude that our micropropagation and acclimatization protocols provide a suitable alternative to traditional mass propagation techniques for the carob tree.
- Seasonal changes in CO2 assimilation in leaves of seedlings and micropropagated plants of Carob tree established in the fieldPublication . Osório, Maria Leonor; Osório, Júlio; David, Maria Manuela; Correia, Maria João; Romano, AnabelaIn this communication we evaluate the field performance of two micropropagated Portuguese carob cultivars (Galhosa and Mulata) throughout the season, particularly at extreme conditions of light and temperature. Two irrigated plots were established in the field: 1) micropropagated plants, vs 2) seedlings. During the first year following transplantation to the field, we followed net photosynthetic rate, stomatal conductance, transpiration rate, chlorophyll a fluorescence and leaf contents in chlorophyll, carotenoids and protein. No significant differences were detected between seedlings and micropropagated plants along the year. However, at the end of summer, despite irrigation, the photosynthetic rate (NP), the quantum yield of PSII (fPSII) and the intrinsic efficiency of open PSII reaction centers (F’v/F’m) declined, concomitantly with the increase of the thermal energy dissipation at the PSII (NPQ). As the maximal photochemical efficiency of PSII (Fv/Fm) was maintained high (0.82), these results indicate that regulated thermal dissipation in light harvesting complexes was promoted in order to avoid photoinhibition. After the first growth period in the field, data from micropropagated plants did not differ from seedlings, and those plants showed the characteristic behaviour of plants well adapted to Mediterranean climates. So, in vitro propagation could be use as a promising alternative to traditional propagation and establishment of carob orchards.