The specificity factor (S) of ribulose-1,5-bisphosphate. carboxylase/oxygenase (Rubisco) defines ... more The specificity factor (S) of ribulose-1,5-bisphosphate. carboxylase/oxygenase (Rubisco) defines the relative rates of carboxylation and oxygenation of Ribulose-1,5-bisphosphate (RuBP) catalysed by the enzyme. The determination of S for Rubisco purified from the Vitis vinifera L. cvs Tempranillo and Manto Negro is described here for the first time. Rubisco extraction was made in Bicine buffer with the inclusion of polyethylene glycol (PEG), beta-mereaptoethanol, diethyldithio-carbamic acid (DIECA) and several protease inhibitors. Furthermore, in the same cultivars, the apparent in vivo specificity factor for Rubisco (S*), was obtained from gas exchange and chlorophyll fluorescence measurements. For both cultivars the values of S were close to 100 at 25 degreesC. However, in mature leaves, S* was about 67 for Manto Negro and 55 or 77 for Tempranillo, depending on leaf age. Leaves of plants under drought showed even lower values. These discrepancies between S and S* are ascribed to eq...
The effects of seasonal drought on the distribution of leaf area, photosynthesis and transpiratio... more The effects of seasonal drought on the distribution of leaf area, photosynthesis and transpiration within the canopy were evaluated for two Spanish grapevine cultivars. Both varieties were cultivated according to their typical training system. At early stages of drought, reduction of photosynthesis and transpiration was only observed in sun-exposed leaves. As drought intensified, even less sun-exposed leaves were affected. Severe drought reduced photosynthesis and transpiration in all locations of the canopy except for most shaded leaves in the inner part. However, those leaves were almost unproductive, and seemed to be insensitive to variation of both light intensity and drought. Leaf area was also reduced by drought, but the distribution of these reductions within the canopy differed between cultivars, possibly reflecting differences in the training system. Leaves from all locations of the canopy except those in the central part showed a similar radiation use efficiency, suggestin...
Climate change forecasts suggest temperature increases and lower rainfall rates, both of which ch... more Climate change forecasts suggest temperature increases and lower rainfall rates, both of which challenge viticulture, particularly in semi-arid areas where water availability is critical. In this scenario, the use of the genetic variability in grapevine varieties reported around the world represents an important strategy for the selection of climate-resilient cultivars. In this work, physiological and metabolomics analyses were conducted to compare the water deficit stress (WDS) responses of red and white, local and widespread grapevines cultivars. Leaf gas exchange, water use efficiency (WUE) and water relation parameters were determined in plants under well-watered and WDS conditions alongside assessment of the levels of foliar primary metabolites using gas-chromatography coupled to mass-spectrometry. Results denote that red and white local cultivars displayed more adapted physiological performance under WDS as compared to the widely-distributed ones. Multivariate analyses and spe...
In the Mediterranean region, grapevines usually deal with drought during their summer growth seas... more In the Mediterranean region, grapevines usually deal with drought during their summer growth season. Concurrently, grapevines are hosts to a large number of viruses from which grapevine leafroll associated virus-3 is one of the most widespread and provokes considerable economic losses in many vineyards. However, information concerning grapevine metabolic responses to the combination of drought and viral infection is scarce. Gas-chromatography coupled to massspectrometry based metabolite profiling was used in combination with growth analysis, viral loads and gas exchange data to perform an integrative study of the effects of individual and combined stress in two Majorcan grapevine varieties at two experimental years. Metabolic responses of both varieties to the combination of water stress and virus infection were specific and not predicted from the sum of single stress responses. Correlations between respiration, biomass and key metabolites highlight specific adjustments of respiratory and amino acid 2 metabolism possibly underlying the maintenance of carbon balance and growth in grapevines under stress combination.
The present work aims to study the interactive effect of drought stress and high vapour pressure ... more The present work aims to study the interactive effect of drought stress and high vapour pressure deficit (VPD) on leaf gas exchange, and especially on mesophyll conductance to CO 2 (g m), in two woody species of great agronomical importance in the Mediterranean basin: Vitis vinifera L. cv. Tempranillo and Olea europaea L. cv. Manzanilla. Plants were grown in specially designed outdoor chambers with ambient and below ambient VPD, under both well-irrigated and drought conditions. g m was estimated by the variable J method from simultaneous measurements of gas exchange and fluorescence. In both species, the response to soil water deficit was larger in g s than in g m , and more important than the response to VPD. Olea europaea was apparently more sensitive to VPD, so that plants growing in more humid chambers showed higher g s and g m. In V. vinifera, in contrast, soil water deficit dominated the response of g s and g m. Consequently, changes in g m /g s were more related to VPD in O. europaea and to soil water deficit in V. vinifera. Most of the limitations of photosynthesis were diffusional and especially due to stomatal closure. No biochemical limitation was detected. The results showed that structural parameters played an important role in determining g m during the acclimation process. Although the relationship between leaf mass per unit area (M A) with g m was scattered, it imposed a limitation to the maximum g m achievable, with higher values of M A in O. europaea at lower g m values. M A decreased under water stress in O. europaea but it increased in V. vinifera. This resulted in a negative relationship between M A and the CO 2 draw-down between substomatal cavities and chloroplasts in O. europaea, while being positive in V. vinifera.
... mesophyll and stomatal conductance to CO2 Jaume Flexas* , Josefina Bota , Antonio Diaz-Espejo... more ... mesophyll and stomatal conductance to CO2 Jaume Flexas* , Josefina Bota , Antonio Diaz-Espejo , Alex Gallé , Jeroni Galmés , Miquel Ribas-Carbo , Francesc Rosselló§, Magdalena Tomàs and Hipólito Medrano Research ...
The application of computer vision algorithms and chemometric fingerprinting using near-infrared ... more The application of computer vision algorithms and chemometric fingerprinting using near-infrared spectrometry (NIR) of plant leaves, offers enhanced capabilities for ampelography by providing more accurate methods to discriminate leaves based on morphological parameters, and chemometrics, respectively. This paper showed that machine learning algorithms based on morpho-colorimetric parameters and NIR analysis separately, were able to automatically classify leaves of 16 grapevine cultivars. The artificial neural network (ANN) model developed with morpho-colorimetric parameters as inputs (Model 1), and 16 cultivars as targets, rendered an accuracy of 94% to classify leaves for all cultivars studied. The ANN model obtained with the NIR spectra per leaf as inputs (Model 2), and the real classification as targets, rendered 92% accuracy. The automatic extraction of morphocolorimetric data, NIR chemical fingerprinting and machine learning modelling rendered rapid, accurate and non-destructive methods for cultivar classification, which can aid management practices.
Dark respiration measurements with open-flow gas exchange analyzers are often questioned for thei... more Dark respiration measurements with open-flow gas exchange analyzers are often questioned for their low accuracy as their low values often reach the precision limit of the instrument. Respiration was measured in five species, two hypostomatous (Vitis Vinifera L. and Acanthus mollis) and three amphistomatous, one with similar amount of stomata in both sides (Eucalyptus citriodora) and two with different stomata density (Brassica oleracea and Vicia faba). CO2 differential (ΔCO2) increased twofold with no change in apparent Rd, when the two leaves with higher stomatal density faced outside. These results showed a clear effect of the position of stomata on ΔCO2. Therefore, it can be concluded that leaf position is important to guarantee the improvement of respiration measurements increasing ΔCO2 without affecting the respiration results by leaf or mass units. This method will help to increase the accuracy of leaf respiration measurements using gas exchange analyzers.
Agraim a l'Institut Nacional d'Investigacio i 'Tecnologia Agraria i Alimentaria (INIA... more Agraim a l'Institut Nacional d'Investigacio i 'Tecnologia Agraria i Alimentaria (INIA) i als fons FEDER el financament d'aquest treballa traves del projecte RF2012- 00027-C5-00.
We review the photosynthetic responses to drought in field-grown grapevines and other species. As... more We review the photosynthetic responses to drought in field-grown grapevines and other species. As in other plant species, the relationship between photosynthesis and leaf water potential and/or relative water content in field-grown grapevines depends on conditions during plant growth and measurements. However, when light-saturated stomatal conductance was used as the reference parameter to reflect drought intensity, a common response pattern was observed that was much less dependent on the species and conditions. Many photosynthetic parameters (e.g. electron transport rate, carboxylation efficiency, intrinsic water-use efficiency, respiration rate in the light, etc.) were also more strongly correlated with stomatal conductance than with water status itself. Moreover, steady-state chlorophyll fluorescence also showed a high dependency on stomatal conductance. This is discussed in terms of an integrated down-regulation of the whole photosynthetic process by CO2 availability in the mes...
Leaf mesophyll conductance to CO 2 (g m ) has been recognized to be finite and variable, rapidly ... more Leaf mesophyll conductance to CO 2 (g m ) has been recognized to be finite and variable, rapidly adapting to environmental conditions. The physiological basis for fast changes in g m is poorly understood, but current reports suggest the involvement of protein-facilitated CO 2 diffusion across cell membranes. A good candidate for this could be the Nicotiana tabacum L. aquaporin NtAQP1, which was shown to increase membrane permeability to CO 2 in Xenopus oocytes. The objective of the present work was to evaluate its effect on the in vivo mesophyll conductance to CO 2 , using plants either deficient in or overexpressing NtAQP1. Antisense plants deficient in NtAQP1 (AS) and NtAQP1 overexpressing tobacco plants (O) were compared with their respective wild-type (WT) genotypes (CAS and CO). Plants grown under optimum conditions showed different photosynthetic rates at saturating light, with a decrease of 13% in AS and an increase of 20% in O, compared with their respective controls. CO 2 response curves of photosynthesis also showed significant differences among genotypes. However, in vitro analysis demonstrated that these differences could not be attributed to alterations in Rubisco activity or ribulose-1,5-bisphosphate content. Analyses of chlorophyll fluorescence and on-line 13 C discrimination indicated that the observed differences in net photosynthesis (A N ) among genotypes were due to different leaf mesophyll conductances to CO 2 , which was estimated to be 30% lower in AS and 20% higher in O compared with their respective WT. These results provide evidence for the in vivo involvement of aquaporin NtAQP1 in mesophyll conductance to CO 2 .
The specificity factor (S) of ribulose-1,5-bisphosphate. carboxylase/oxygenase (Rubisco) defines ... more The specificity factor (S) of ribulose-1,5-bisphosphate. carboxylase/oxygenase (Rubisco) defines the relative rates of carboxylation and oxygenation of Ribulose-1,5-bisphosphate (RuBP) catalysed by the enzyme. The determination of S for Rubisco purified from the Vitis vinifera L. cvs Tempranillo and Manto Negro is described here for the first time. Rubisco extraction was made in Bicine buffer with the inclusion of polyethylene glycol (PEG), beta-mereaptoethanol, diethyldithio-carbamic acid (DIECA) and several protease inhibitors. Furthermore, in the same cultivars, the apparent in vivo specificity factor for Rubisco (S*), was obtained from gas exchange and chlorophyll fluorescence measurements. For both cultivars the values of S were close to 100 at 25 degreesC. However, in mature leaves, S* was about 67 for Manto Negro and 55 or 77 for Tempranillo, depending on leaf age. Leaves of plants under drought showed even lower values. These discrepancies between S and S* are ascribed to eq...
The effects of seasonal drought on the distribution of leaf area, photosynthesis and transpiratio... more The effects of seasonal drought on the distribution of leaf area, photosynthesis and transpiration within the canopy were evaluated for two Spanish grapevine cultivars. Both varieties were cultivated according to their typical training system. At early stages of drought, reduction of photosynthesis and transpiration was only observed in sun-exposed leaves. As drought intensified, even less sun-exposed leaves were affected. Severe drought reduced photosynthesis and transpiration in all locations of the canopy except for most shaded leaves in the inner part. However, those leaves were almost unproductive, and seemed to be insensitive to variation of both light intensity and drought. Leaf area was also reduced by drought, but the distribution of these reductions within the canopy differed between cultivars, possibly reflecting differences in the training system. Leaves from all locations of the canopy except those in the central part showed a similar radiation use efficiency, suggestin...
Climate change forecasts suggest temperature increases and lower rainfall rates, both of which ch... more Climate change forecasts suggest temperature increases and lower rainfall rates, both of which challenge viticulture, particularly in semi-arid areas where water availability is critical. In this scenario, the use of the genetic variability in grapevine varieties reported around the world represents an important strategy for the selection of climate-resilient cultivars. In this work, physiological and metabolomics analyses were conducted to compare the water deficit stress (WDS) responses of red and white, local and widespread grapevines cultivars. Leaf gas exchange, water use efficiency (WUE) and water relation parameters were determined in plants under well-watered and WDS conditions alongside assessment of the levels of foliar primary metabolites using gas-chromatography coupled to mass-spectrometry. Results denote that red and white local cultivars displayed more adapted physiological performance under WDS as compared to the widely-distributed ones. Multivariate analyses and spe...
In the Mediterranean region, grapevines usually deal with drought during their summer growth seas... more In the Mediterranean region, grapevines usually deal with drought during their summer growth season. Concurrently, grapevines are hosts to a large number of viruses from which grapevine leafroll associated virus-3 is one of the most widespread and provokes considerable economic losses in many vineyards. However, information concerning grapevine metabolic responses to the combination of drought and viral infection is scarce. Gas-chromatography coupled to massspectrometry based metabolite profiling was used in combination with growth analysis, viral loads and gas exchange data to perform an integrative study of the effects of individual and combined stress in two Majorcan grapevine varieties at two experimental years. Metabolic responses of both varieties to the combination of water stress and virus infection were specific and not predicted from the sum of single stress responses. Correlations between respiration, biomass and key metabolites highlight specific adjustments of respiratory and amino acid 2 metabolism possibly underlying the maintenance of carbon balance and growth in grapevines under stress combination.
The present work aims to study the interactive effect of drought stress and high vapour pressure ... more The present work aims to study the interactive effect of drought stress and high vapour pressure deficit (VPD) on leaf gas exchange, and especially on mesophyll conductance to CO 2 (g m), in two woody species of great agronomical importance in the Mediterranean basin: Vitis vinifera L. cv. Tempranillo and Olea europaea L. cv. Manzanilla. Plants were grown in specially designed outdoor chambers with ambient and below ambient VPD, under both well-irrigated and drought conditions. g m was estimated by the variable J method from simultaneous measurements of gas exchange and fluorescence. In both species, the response to soil water deficit was larger in g s than in g m , and more important than the response to VPD. Olea europaea was apparently more sensitive to VPD, so that plants growing in more humid chambers showed higher g s and g m. In V. vinifera, in contrast, soil water deficit dominated the response of g s and g m. Consequently, changes in g m /g s were more related to VPD in O. europaea and to soil water deficit in V. vinifera. Most of the limitations of photosynthesis were diffusional and especially due to stomatal closure. No biochemical limitation was detected. The results showed that structural parameters played an important role in determining g m during the acclimation process. Although the relationship between leaf mass per unit area (M A) with g m was scattered, it imposed a limitation to the maximum g m achievable, with higher values of M A in O. europaea at lower g m values. M A decreased under water stress in O. europaea but it increased in V. vinifera. This resulted in a negative relationship between M A and the CO 2 draw-down between substomatal cavities and chloroplasts in O. europaea, while being positive in V. vinifera.
... mesophyll and stomatal conductance to CO2 Jaume Flexas* , Josefina Bota , Antonio Diaz-Espejo... more ... mesophyll and stomatal conductance to CO2 Jaume Flexas* , Josefina Bota , Antonio Diaz-Espejo , Alex Gallé , Jeroni Galmés , Miquel Ribas-Carbo , Francesc Rosselló§, Magdalena Tomàs and Hipólito Medrano Research ...
The application of computer vision algorithms and chemometric fingerprinting using near-infrared ... more The application of computer vision algorithms and chemometric fingerprinting using near-infrared spectrometry (NIR) of plant leaves, offers enhanced capabilities for ampelography by providing more accurate methods to discriminate leaves based on morphological parameters, and chemometrics, respectively. This paper showed that machine learning algorithms based on morpho-colorimetric parameters and NIR analysis separately, were able to automatically classify leaves of 16 grapevine cultivars. The artificial neural network (ANN) model developed with morpho-colorimetric parameters as inputs (Model 1), and 16 cultivars as targets, rendered an accuracy of 94% to classify leaves for all cultivars studied. The ANN model obtained with the NIR spectra per leaf as inputs (Model 2), and the real classification as targets, rendered 92% accuracy. The automatic extraction of morphocolorimetric data, NIR chemical fingerprinting and machine learning modelling rendered rapid, accurate and non-destructive methods for cultivar classification, which can aid management practices.
Dark respiration measurements with open-flow gas exchange analyzers are often questioned for thei... more Dark respiration measurements with open-flow gas exchange analyzers are often questioned for their low accuracy as their low values often reach the precision limit of the instrument. Respiration was measured in five species, two hypostomatous (Vitis Vinifera L. and Acanthus mollis) and three amphistomatous, one with similar amount of stomata in both sides (Eucalyptus citriodora) and two with different stomata density (Brassica oleracea and Vicia faba). CO2 differential (ΔCO2) increased twofold with no change in apparent Rd, when the two leaves with higher stomatal density faced outside. These results showed a clear effect of the position of stomata on ΔCO2. Therefore, it can be concluded that leaf position is important to guarantee the improvement of respiration measurements increasing ΔCO2 without affecting the respiration results by leaf or mass units. This method will help to increase the accuracy of leaf respiration measurements using gas exchange analyzers.
Agraim a l'Institut Nacional d'Investigacio i 'Tecnologia Agraria i Alimentaria (INIA... more Agraim a l'Institut Nacional d'Investigacio i 'Tecnologia Agraria i Alimentaria (INIA) i als fons FEDER el financament d'aquest treballa traves del projecte RF2012- 00027-C5-00.
We review the photosynthetic responses to drought in field-grown grapevines and other species. As... more We review the photosynthetic responses to drought in field-grown grapevines and other species. As in other plant species, the relationship between photosynthesis and leaf water potential and/or relative water content in field-grown grapevines depends on conditions during plant growth and measurements. However, when light-saturated stomatal conductance was used as the reference parameter to reflect drought intensity, a common response pattern was observed that was much less dependent on the species and conditions. Many photosynthetic parameters (e.g. electron transport rate, carboxylation efficiency, intrinsic water-use efficiency, respiration rate in the light, etc.) were also more strongly correlated with stomatal conductance than with water status itself. Moreover, steady-state chlorophyll fluorescence also showed a high dependency on stomatal conductance. This is discussed in terms of an integrated down-regulation of the whole photosynthetic process by CO2 availability in the mes...
Leaf mesophyll conductance to CO 2 (g m ) has been recognized to be finite and variable, rapidly ... more Leaf mesophyll conductance to CO 2 (g m ) has been recognized to be finite and variable, rapidly adapting to environmental conditions. The physiological basis for fast changes in g m is poorly understood, but current reports suggest the involvement of protein-facilitated CO 2 diffusion across cell membranes. A good candidate for this could be the Nicotiana tabacum L. aquaporin NtAQP1, which was shown to increase membrane permeability to CO 2 in Xenopus oocytes. The objective of the present work was to evaluate its effect on the in vivo mesophyll conductance to CO 2 , using plants either deficient in or overexpressing NtAQP1. Antisense plants deficient in NtAQP1 (AS) and NtAQP1 overexpressing tobacco plants (O) were compared with their respective wild-type (WT) genotypes (CAS and CO). Plants grown under optimum conditions showed different photosynthetic rates at saturating light, with a decrease of 13% in AS and an increase of 20% in O, compared with their respective controls. CO 2 response curves of photosynthesis also showed significant differences among genotypes. However, in vitro analysis demonstrated that these differences could not be attributed to alterations in Rubisco activity or ribulose-1,5-bisphosphate content. Analyses of chlorophyll fluorescence and on-line 13 C discrimination indicated that the observed differences in net photosynthesis (A N ) among genotypes were due to different leaf mesophyll conductances to CO 2 , which was estimated to be 30% lower in AS and 20% higher in O compared with their respective WT. These results provide evidence for the in vivo involvement of aquaporin NtAQP1 in mesophyll conductance to CO 2 .
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