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Acoustic emission analysis is a promising technique to investigate the physiological events leading to drought-induced injuries and mortality. However, the nature and the source of the acoustic emissions are not fully understood and make the use of this technique difficult as a direct measure of the loss of xylem hydraulic conductance. In this study, acoustic emissions were recorded during severe dehydration in lavender plants and compared to the dynamics of embolism development and cell lysis. The timing and characteristics of acoustic signals from two independent recording systems were compared by principal component analysis. In parallel, changes in water potential, branch diameter, loss of hydraulic conductance and electrolyte leakage were measured to quantify drought-induced damages. Two distinct phases of acoustic emissions were observed during dehydration: the first one associated with a rapid loss of diameter and a significant increase in loss of xylem conductance (90%) and the second one with a significant increase in electrolyte leakage and slower diameter changes. This second phase corresponds to a complete loss of recovery capacity. The acoustic signals of both phases were discriminated by the third and fourth principal components. The loss of hydraulic conductance during the first acoustic phase suggests the hydraulic origin of these signals (i.e. cavitation events). For the second phase, the signals showed much higher variability between plants and acoustic systems suggesting that the sources of these signals may be plural, although likely including cellular damage. A simple algorithm was developed to discriminate hydraulic-related acoustic signals from other sources, allowing the reconstruction of dynamic hydraulic vulnerability curves. However, hydraulic failure precedes cellular damage and lack of whole plant recovery is associated to these latter., 6 figures, 1 table, + 2 supplementary figures and 1 supplementary table

Background and Aims Determining within-species large-scale variation in phenotypic traits is central to elucidate the drivers of species’ ranges. Intraspecific comparisons offer the opportunity to understand how trade-offs and biogeographical history constrain adaptation to contrasted environmental conditions. Here we test whether functional traits, ecological strategies from the CSR scheme and phenotypic plasticity in response to abiotic stress vary along a latitudinal or a center- margins gradient within the native range of Arabidopsis thaliana. Methods We experimentally examined the phenotypic outcomes of plant adaptation at the center and margins of its geographic range using 30 accessions from southern, central and northern Europe. We characterized the variation of traits related to stress tolerance, resource use, colonization ability, CSR strategy scores, survival and fecundity in response to high temperature (34 °C) or frost (- 6 °C), combined with a water deficit treatment. Key Results We found evidence for both a latitudinal and a center-margins differentiation for the traits under scrutiny. Age at maturity, leaf dry matter content, specific leaf area and leaf nitrogen content varied along a latitudinal gradient. Northern accessions presented a greater survival to stress than central and southern accessions. Leaf area, C-scores, R-scores and fruit number followed a center-margins differentiation. Central accessions displayed a higher phenotypic plasticity than northern and southern accessions for most studied traits. Conclusions Traits related to an acquisitive/conservative resource-use trade-off followed a latitudinal gradient. Traits associated with a competition/colonization trade-off differentiated along the historic colonization of the distribution range and then followed a center-margins differentiation. Our findings pinpoint the need to consider the joint effect of evolutionary history and environmental factors when examining phenotypic variation across the distribution range of a species.

The capacity of trees to tolerate and survive increasing drought conditions in situ will depend in part on their ability to acclimate (via phenotypic plasticity) key hydraulic and morphological traits that increase drought tolerance and delay the onset of drought-induced hydraulic failure. However, the effect of water-deficit acclimation in key traits that determine time to hydraulic failure (THF) during extreme drought remains largely untested. We measured key hydraulic and morphological traits in saplings of a hybrid poplar grown under well-watered and water-limited conditions. The time for plants to dry-down to critical levels of water stress (90% loss of stem hydraulic conductance), as well as the relative contribution of drought acclimation in each trait to THF, was simulated using a soil–plant hydraulic model (SurEau). Compared with controls, water-limited plants exhibited significantly lower stem hydraulic vulnerability (P50stem), stomatal conductance and total canopy leaf area (LA). Taken together, adjustments in these and other traits resulted in longer modelled THF in water-limited (~160 h) compared with well-watered plants (~50 h), representing an increase of more than 200%. Sensitivity analysis revealed that adjustment in P50stem and LA contributed the most to longer THF in water-limited plants. We observed a high degree of trait plasticity in poplar saplings in response to water-deficit growth conditions, with decreases in stem hydraulic vulnerability and leaf area playing a key role in delaying the onset of hydraulic failure during a simulated drought event. These findings suggest that understanding the capacity of plants to acclimate to antecedent growth conditions will enable better predictions of plant survivorship during future drought.

International audience; Polyploidy plays a major role in citrus plant breeding to improve the adaptation of polyploid rootstocks as well as scions to adverse conditions and to enhance agronomic characteristics. In Citrus breeding programs, triploidy could be a useful tool to react to environmental issues and consumer demands because the produced fruits are seedless. In this study, we compared the physiological, biochemical, morphological, and ultrastructural responses to water deficit of triploid and diploid citrus varieties obtained from 'Fortune' mandarin and `Ellendale' tangor hybridization. One diploid clementine tree was included and used as a reference. All studied scions were grafted on C-35 citrange rootstock. Triploidy decreased stomatal density and increased stomata size. The number of chloroplasts increased in 3x varieties. These cytological properties may explain the greater photosynthetic capacity (Pnet, gs, Fv/Fm) and enhanced water-holding capacity (RWC, proline). In addition, reduced degradation of ultrastructural organelles (chloroplasts and mitochondria) and thylakoids accompanied by less photosynthetic activity and low oxidative damages were found in 3x varieties. Triploid varieties, especially T40-3x, had a better ability to limit water loss and dissipate excess energy (NPQ) to protect photosystems. Higher starch reserves in 3x varieties suggest a better carbon and energy supply and increases in plastoglobuli size suggest less oxidative damage (H2O2, MDA), especially in T40-3x, and preservation of photosynthetic apparatus. Taken together, our results suggest that desirable cytological and ultrastructural traits induced by triploidy improve water stress response and could be a useful stress marker during environmental constraints.

Nutrient deficiency alters growth and the production of high-quality nutritious food. In Citrus crops, rootstock technologies have become a key tool for enhancing tolerance to abiotic stress. The use of doubled diploid rootstocks can improve adaptation to lower nutrient inputs. This study investigated leaf structure and ultrastructure and physiological and biochemical parameters of diploid common clementine scions (C) grafted on diploid (2x) and doubled diploid (4x) Carrizo citrange (C/CC2x and C/CC4x) and Citrumelo 4475 (C/CM2x and C/CM4x) rootstocks under optimal fertigation and after 7 months of nutrient deficiency. Rootstock ploidy level had no impact on structure but induced changes in the number and/or size of cells and some cell components of 2x common clementine leaves under optimal nutrition. Rootstock ploidy level did not modify gas exchanges in Carrizo citrange but induced a reduction in the leaf net photosynthetic rate in Citrumelo 4475. By assessing foliar damage, changes in photosynthetic processes and malondialdehyde accumulation, we found that C/CM4x were less affected by nutrient deficiency than the other scion/rootstock combinations. Their greater tolerance to nutrient deficiency was probably due to the better performance of the enzyme-based antioxidant system. Nutrient deficiency had similar impacts on C/CC2x and C/CC4x. Tolerance to nutrient deficiency can therefore be improved by rootstock polyploidy but remains dependent on the rootstock genotype.

International audience; Xylem hydraulic failure has been recognized as a pervasive factor in the triggering of drought-induced tree mortality. However, foundational evidence of the mechanistic link connecting hydraulic failure with living cell damage and tree death has not been identified yet, compromising our ability to predict mortality events. Meristematic cells are involved in the recovery of trees from drought, and focusing on their vitality and functionality after a drought event could provide novel information on the mechanistic link between hydraulic failure and drought-induced tree mortality. In this Opinion, we focus on the cell's critical hydration status for tree recovery from drought and how it links with the membrane integrity of the meristems.

Understanding variation in drought resistance traits is needed to predict the potential of trees to adapt to severe drought events. Xylem vulnerability to embolism is a critical trait related to drought-induced mortality that has large variability between species. Acclimation of this trait to environmental conditions implies changes in the xylem structure and organization, leading previous studies to investigate its variations under conditions that preserve growth. In European beech (Fagus sylvatica L.) saplings, we assessed the effect of drought on the species’ vulnerability to embolism in branches developed the next year during recovery. Newly formed branches displayed less vulnerability to embolism in plants that experienced the most severe droughts that led to native embolism. The pressure that induced 50% loss of conductance in severely drought affected plants versus control plants was −3.98 versus −3.1 MPa, respectively. These results argue for a lagged acclimation of this trait to drought events.

Knowledge on variations of drought resistance traits are needed to predict the potential of trees to acclimate to coming severe drought events. Xylem vulnerability to embolism is a key parameter related to such droughts, and its phenotypic variability relies mainly on environmental plasticity. We investigated the structural determinants controlling the plasticity of vulnerability to embolism, focusing on the key elements involved in the air bubble entry in vessels, especially the intervessel pits. Poplar saplings (Populus tremula x alba (Aiton) Sm., 1804) grown in contrasted water availability or light exposure exhibited differences in the vulnerability to embolism (P50) in a range of 0.76 MPa. We then characterized the structural changes in features related to pit quantity and pit structure, from the pit ultrastructure to the organization of xylem vessels, using different microscopy techniques (transmission electron microscopy, scanning electron microscopy, light microscopy). A multispectral combination of X-ray microtomography and light microscopy analysis allowed measuring the vulnerability of each single vessel and testing some of the relationships between structural traits and vulnerability to embolism inside the xylem. The pit ultrastructure did not change, whereas the vessel dimensions increased with the vulnerability to embolism and the grouping index and fraction of intervessel cell wall both decreased with the vulnerability to embolism. These findings hold when comparing between trees or between the vessels inside the xylem of an individual tree. These results evidenced that plasticity of vulnerability to embolism in hybrid poplar occurs through changes in the pit quantity properties such as pit area and vessel grouping rather than changes on the pit structure.

The triploidy has proved to be a powerful approach breeding programs, especially in Citrus since seedlessness is one of the main consumer expectations. Citrus plants face numerous abiotic stresses including water deficit, which negatively impact growth and crop yield. In this study, we evaluated the physiological and biochemical responses to water deficit and recovery capacity of new triploid hybrids, in comparison with diploid hybrids, their parents (“Fortune” mandarin and “Ellendale” tangor) and one clementine tree used as reference. The water deficit significantly decreased the relative water content (RWC) and leaf gas exchange (Pnet and gs) and it increased the levels of oxidative markers (H2O2 and MDA) and antioxidants. Compared to diploid varieties, triploid hybrids limited water loss by osmotic adjustment as reflected by higher RWC, intrinsic water use efficiency (iWUE Pnet/gs) iWUE and leaf proline levels. These had been associated with an effective thermal dissipation of excess energy (NPQ) and lower oxidative damage. Our results showed that triploidy in citrus enhances the recovery capacity after a water deficit in comparison with diploids due to better carboxylation efficiency, restored water-related parameters and efficient antioxidant system.

Nutrient deficiency alters growth and the production of high-quality nutritious food. In Citrus crops, rootstock technologies have become a key tool for enhancing tolerance to abiotic stress. The use of doubled diploid rootstocks can improve adaptation to lower nutrient inputs. This study investigated leaf structure and ultrastructure and physiological and biochemical parameters of common clementine scions (C) grafted on diploid (2x) and doubled diploid (4x) Carrizo citrange (C/CC2x and C/CC4x) and Citrumelo 4475 (C/CM2x and C/CM4x) rootstocks under optimal fertigation and after seven months of nutrient deficiency. Rootstock ploidy level had no impact on structure but induced changes in the number and/or size of cells and some cell components of common clementine leaves under optimal nutrition. Rootstock ploidy level did not modify gas exchanges in Carrizo citrange but induced a reduction in the leaf net photosynthetic rate in Citrumelo 4475. By assessing foliar damage, changes in photosynthetic processes and malondialdehyde accumulation, we found that C/CM4x were less affected by nutrient deficiency than the other scion/rootstock combinations. Their greater tolerance to nutrient deficiency was probably due to the better performance of the enzyme-based antioxidant system. Nutrient deficiency had similar impacts on C/CC2x and C/CC4x. Tolerance to nutrient deficiency can therefore be improved by rootstock polyploidy but remains dependent on the rootstock genotype.

Nutrient deficiency, in particular when this involves a major macronutrient (N, P, and K), is a limiting factor on the performance of plants in their natural habitat and agricultural environment. In the citrus industry, one of the eco-friendliest techniques for improving tolerance to biotic and abiotic stress is based on the grafting of a rootstock and a scion of economic interest. Scion tolerance may be improved by a tetraploid rootstock. The purpose of this study was to highlight if tolerance of a common clementine scion (C) (Citrus clementina Hort. ex Tan) to nutrient deficiency could be improved by several diploid (2×) and their tetraploid (4×) counterparts citrus genotypes commonly used as rootstocks: Trifoliate orange × Cleopatra mandarin (C/PMC2x and C/PMC4x), Carrizo citrange (C/CC2x and C/CC4x), Citrumelo 4475 (C/CM2x and C/CM4x). The allotetraploid FlhorAG1 (C/FL4x) was also included in the experimental design. The impact of nutrient deficiency on these seven scion/rootstock combinations was evaluated at root and leaf levels by investigating anatomical parameters, photosynthetic properties and oxidative and antioxidant metabolism. Nutrient deficiency affects foliar tissues, physiological parameters and oxidative metabolism in leaves and roots in different ways depending on the rootstock genotype and ploidy level. The best known nutrient deficiency-tolerant common clementine scions were grafted with the doubled diploid Citrumelo 4475 (C/CM4x) and the allotetraploid FlhorAG1 (C/FL4x). These combinations were found to have less foliar damage, fewer changes of photosynthetic processes [leaf net photosynthetic rate (Pnet), stomatal conductance (gs), transpiration (E), maximum quantum efficiency of PSII (Fv/Fm), electron transport rate (ETR), ETR/Pnet], and effective quantum yield of PSII [Y(II)], less malondialdehyde accumulation in leaves and better functional enzymatic and non-enzymatic antioxidant systems. Common clementine scions grafted on other 4× rootstocks did not show better tolerance than those grafted on their 2× counterparts. Chromosome doubling of rootstocks did not systematically improve the tolerance of the common clementine scion to nutrient deficiency.

Knowledge on variations of drought resistance traits are needed to predict the potential of trees to acclimate to coming severe drought events. Xylem vulnerability to embolism is a key parameter related to such droughts, and its phenotypic variability relies mainly on environmental plasticity. We investigated the structural determinants controlling the plasticity of vulnerability to embolism, focusing on the key elements involved in the air bubble entry in a vessel, especially the inter-vessel pits. Poplar saplings (Populus tremula x alba) grown in contrasted water availability or light exposure exhibited differences in vulnerability to embolism in a range of 0.76 MPa. We then characterized the structural changes related to qualitative and quantitative pit characteristics, from the pit structure to the organization of xylem vessels, using different microscopy techniques (TEM, SEM, light). X-ray microtomography analysis allowed observing the vessel vulnerability and testing some of the relationships between structural traits and vulnerability to embolism inside the xylem. The pit ultrastructure did not change, whereas the vessel dimensions increased with vulnerability to embolism and the grouping index and fraction of inter-vessel cell wall decreased with vulnerability to embolism. These findings holds when comparing trees or when comparing vessels inside the xylem. These results evidenced that plasticity of vulnerability to embolism occurs through changes in the quantitative pit properties such as pit area and vessel grouping rather than on the pit structure.

Knowledge on variations of drought resistance traits is needed to predict the potential of trees to adapt to severe drought events expected to be more intense and frequent. Xylem vulnerability to cavitation is among the most important traits related to drought-induced mortality and exhibits a large variability between species. Acclimation of this trait to environmental conditions implies changes in the xylem structure and organization, leading previous studies to investigate its variations under conditions preserving growth. In European beech saplings, we assessed the effect of droughts of on the vulnerability to cavitation in branches that develop during recovery. The newly formed branches displayed lower vulnerability to cavitation in the plants that underwent the severest droughts leading to native embolism; the pressure that induces 50% loss of conductance being of −3.98 MPa in severely droughted plants whereas it was of −3.1 MPa in control plants, respectively. Although unexpected, these results argue for an acclimation, and not a weakening, of this trait to drought events. Key message severe water stress make the future developed shoots less vulnerable to xylem cavitation.

International audience; Low temperatures during winter are one of the main constraints for citrus crop. Polyploid rootstocks can be used for improving tolerance to abiotic stresses, such as cold stress. Because the produced fruit are seedless, using triploid scions is one of the most promising approaches to satisfy consumer expectations. In this study, we evaluated how the triploidy of new citrus varieties influences their sensitivity to natural chilling temperatures. We compared their behavior to that of diploid citrus, their parents (Fortune mandarin and Ellendale tangor), and one diploid clementine tree, as reference, focusing on photosynthesis parameters, oxidative metabolism, and volatile organic compounds (VOC) in leaves. Triploid varieties appeared to be more tolerant than diploid ones to natural low temperatures, as evidenced by better photosynthetic properties (P-net, g(s), F-v/F-m, ETR/P-net ratio), without relying on a better antioxidant system. The VOC levels were not influenced by chilling temperatures; however, they were affected by the ploidy level and atypical chemotypes were found in triploid varieties, with the highest proportions of E-beta-ocimene and linalool. Such compounds may contribute to better stress adaptation.

International audience; In the context of climate change, determining the physiological mechanisms of drought-induced mortality in woody plants and identifying thresholds of drought survivorship will improve forecasts of forest and agroecosystem die-off. Here, we tested whether continuous measurements of branch diameter variation can be used to identify thresholds of hydraulic failure and physiological recoverability in lavender (Lavandula angustifoliaandLavandulaxintermedia) plants exposed to severe drought. Two parameters of branch diameter variation were tested: the percentage loss of diameter and the percentage loss of rehydration capacity. In two greenhouse experiments with different growth conditions, we monitored variation in branch diameter in the two lavender species exposed to a series of drought/rewatering cycles that varied in drought-stress intensity. Water potential, stomatal conductance, loss of xylem hydraulic conductance, and electrolyte leakage were also measured. We observed that plants were not able to recover when percentage loss of diameter reached maximum values of 21.3% +/- 0.6% during drought, regardless of species and growth conditions. A percentage loss of rehydration capacity of 100% was defined as the point of no recovery, and was observed with high levels of cellular damage as estimated by electrolyte leakage measured at 75.4% +/- 9.3% and occurred beyond 88% loss of xylem hydraulic conductance. Our study demonstrates that lavender plants are not able to recover from severe drought when they have used up their elastic water storage. Additionally, drought-induced mortality in these species was not linked to xylem hydraulic failure but rather to high levels of cell damage. Under extreme drought, death of lavender plants occurs when the water storage of the elastic compartment of the branch is exhausted.

Current climatic upheavals reduce water availability which impacts the growth and fruit quality of plants. In citrus crops, scion/rootstock combinations are used to ensure high fruit production and quality and a stress tolerance/resistance. Our objective was to assess the effect on the clementine scion (C) under natural mild water deficit of (i) polyploid rootstocks by comparing the allotetraploid FlhorAG1 (C/4xFLs, trifoliate orange + Willowleaf mandarin) with its diploid parents, trifoliate orange (C/2xTOs), and Willowleaf mandarin (C/2xWLs), and with a diploid genotype used as reference (Carrizo citrange, C/2xCCs), (ii) rootstock propagation methods by comparing trifoliate orange seedling (C/2xTOs) with cutting (C/2xTOc). A mild water deficit observed under orchard conditions during the summer period (July&ndash, August) induced a significant change in yield (except in C/2xTOs), fruit size, and quality. C/2xCCs, C/2xTOs, and C/2xWLs appeared less affected by water deficit as indicated by their lower reduction of predawn leaf water potential (&Psi, pd), relative water content (RWC), transpiration (E), and photosynthetic parameters (Pnet and gs). Their greater redox balance was probably due to their better antioxidant efficiency. Seedling rootstocks lead to a better adaptation of clementine scions to water deficit than cutting or allotetraploid rootstock. Improving the tolerance to water deficit requires taking into consideration the rootstock genotype, propagation method, and ploidy level.

While the xylem hydraulic properties, such as vulnerability to cavitation (VC), are of paramount importance in drought resistance, their genetic determinants remain unexplored. There is evidence that pectins and their methylation pattern are involved, but the detail of their involvement and the corresponding genes need to be clarified. We analysed the hydraulic properties of the 35S::PME1 transgenic aspen that ectopically under- or over-express a xylem-abundant pectin methyl esterase, PtxtPME1. We also produced and analyzed 4CL1::PGII transgenic poplars expressing a fungal polygalacturonase, AnPGII, under the control of the Ptxa4CL1 promoter that is active in the developing xylem after xylem cell expansion. Both the 35S::PME1 under and over-expressing aspen lines developed xylem with lower specific hydraulic conductivity and lower VC, while the 4CL1::PGII plants developed xylem with a higher VC. These xylem hydraulic changes were associated with modifications in xylem structure or in intervessel pit structure that can result in changes in mechanical behaviour of the pit membrane. This study shows that homogalacturonans and their methylation pattern influence xylem hydraulic properties, through its effect on xylem cell expansion and on intervessel pit properties and it show a role for PtxtPME1 in the xylem hydraulic properties.

In the context of climate changes, water availability is expected to severely decline. Consequently, there is a need to predict mortality of woody species, especially to find a physiological threshold to drought-induced mortality. Lavender species (Lavandula angustifolia and Lavandula x intermedia) which are important crops of the Mediterranean region are affected by a decline, notably caused by successive intense drought events. Lavender response to extreme drought events was monitored using continuous stem diameter measurements. Water potential, stomatal conductance, loss of xylem hydraulic conductivity and electrolyte leakage were also measured during desiccation, and recovery was evaluated after rewatering. Two parameters computed from stem diameter variations were related to stress intensity and resilience to stress: PLD (Percentage Loss of Diameter) and stem PLRC (Percentage Loss of Rehydration Capacity of the stem), respectively. We showed that plants did not recover when the PLD reached its maximal value (PLDmax) which was 21.27 ± 0.57% in both lavender species and whatever the growing conditions. This point of no return was associated with a high level of cell lysis evaluated by electrolyte leakage, and occurred far after the xylem hydraulic failure. We discussed the relevance of PLDmax as a threshold for drought-induced mortality and its physiological significance, in relation to the mortality mechanisms.One-sentence summaryUnder extreme drought, lavender death occurs when the water storage of the elastic compartment of the stem is exhausted.

International audience; Plants require essential minerals for their growth and development that are mainly acquired from soil by their roots. Nutrient deficiency is an environmental stress that can seriously affect fruit production and quality. In citrus crops, rootstock/scion combinations are frequently employed to enhance tolerance to various abiotic stresses. These tolerances can be improved in doubled diploid genotypes. The aim of this work was to compare the impact of nutrient deficiency on the physiological and biochemical response of diploid (2x) and doubled diploid (4x) citrus seedlings: Volkamer lemon, Trifoliate orange ´ Cleopatra mandarin hybrid, Carrizo citrange, Citrumelo 4475. Flhorag1 (Poncirus trifoliata + and willow leaf mandarin), an allotetraploid somatic hybrid, was also included in this study. Our results showed that depending on the genotype, macronutrient and micronutrient deficiency affected certain physiological traits and oxidative metabolism differently. Tetraploid genotypes, mainly Flhorag1 and Citrumelo 4475, appeared resistant compared to the other genotypes as indicated by the lesser decrease in photosynthetic parameters (Pnet, Fv/Fm and Gs) and the lower accumulation of oxidative markers (MDA and H202) in roots and leaves, especially after long-term nutrient deficiency. Their higher tolerance to nutrient deficiency could be explained by better activation of their antioxidant system. For the other genotypes, tetraploidization did not induce greater tolerance to nutrient deficiency.

International audience; Nutrient deficiency has economic and ecological repercussions for citrus fruit crops worldwide. Citrus crops rely on fertilization to maintain good fruit output and quality, whereas new crop management policy aims to reduce fertilizers input. New rootstocks are needed to meet to this constraint, and the use of new tetraploid rootstocks better adapted to lower nutrient intake could offer a promising way forward. Here we compared physiological, biochemical and anatomic traits of leaves in diploid (2x) and doubled-diploid (4x) Citrumelo 4475 (Citrus paradisi L. Macf. × Poncirus trifoliata L. Raf.) and Volkamer lemon (Citrus limonia Osb.) seedlings over 7 months of nutrient deficiency. Photosynthetic parameters (Pnet, Gs and Fv/Fm) decreased, but to a lesser extent in 4x genotypes than 2x. Degradation of the ultrastructural organelles (chloroplasts and mitochondria) and compound cells (thylakoids and starches) was also lower in 4x genotypes, suggesting that tetraploidy may enhance tolerance to nutrient deficiency. However, leaf surface (stomata, stomatal density and epithelial cells) showed no nutrient deficiency-induced change. In 4x Citrumelo 4475, the higher tolerance to nutrient deficiency was associated with a lower MDA and H2O2 accumulation than in the 2x, suggesting a more efficient antioxidant system in the 4x genotype. However, few differences in antioxidant system and oxidative status were observed between 2x and 4x Volkamer lemons.

The genome doubling of the allotetraploid somatic hybrid can confer greater tolerance to cold and light stress than the diploid parents and their respective tetraploid. Allopolyploids are generally known to display broader adaptation to abiotic stresses than their parental diploid species. In the Mediterranean area, Citrus species are subjected to abiotic constraints such as low temperature and high radiation. Tetraploids are known to resist these environmental constraints better, and so the use of new tetraploid rootstocks offers an alternative to overcome these threats to crop productivity. The objective of this study was to determine whether the use of an allotetraploid hybrid could provide greater tolerance to cold and light stresses than its diploid parents or respective doubled-diploid parents. We compared cold and light stress responses of the allotetraploid hybrid FlhorAG1 (FL-4x) with those of its diploid parents, the willow leaf mandarin (Citrus deliciosa Ten) (WLM-2x) and the Poncirus Pomeroy (Poncirus trifoliata (L.) Raf.) (POP-2x), and their respective doubled-diploids (WLM-4x and POP-4x, respectively) by measuring physiological and biochemical parameters. When subjected to cold and light stress, FL-4x showed lower photoinhibition (Fv/Fm) and less accumulation of oxidative markers (MDA and H2O2) than diploid and doubled-diploid WLM and POP genotypes. This was correlated with a greater increase for FL-4x in some antioxidant activities during cold stress (SOD, APX and GR) and light stress (SOD, APX and MDHAR mainly). Overall, our results suggest that greater antioxidant capability in FL-4x should make this allotetraploid hybrid more tolerant to low temperatures than the two WLM genotypes, and more tolerant to light stress than the two WLM and POP genotypes.

Drought-tolerance selection is a current challenge for breeding programs to ensure agrosystem resilience, particularly with intensity and frequency of drought increasing worldwide. In tree species, xylem vulnerability to cavitation is among the most important traits on which drought-induced mortality depends. It can be rapidly evaluated, enabling large-scale screening for drought resistance. Genetic variability and phenotypic plasticity for this trait have been studied in natural populations, but not yet for cultivated tree species. In this work, the genetic variability in xylem vulnerability to cavitation of six Persian walnuts (Juglans regia L.), six hybrid walnuts (J. regia × Juglans nigra), and seven walnut species was investigated. In the first step, the method for measuring xylem vulnerability to cavitation using the “Cavitron” centrifuge technique was improved on walnut samples to obtain more accurate results. The Cavitron technique was found well suited to assessing xylem vulnerability to cavitation on this species using a 0.38 m rotor, which is large enough to analyze samples with intact vessels. Despite differences in wood anatomical traits, xylem vulnerability to cavitation among the Persian walnuts studied was similar. Very narrow variations in xylem vulnerability to cavitation were also found among hybrid walnuts. Only slight differences were observed on comparing species among which some have shown differences in various traits. These results suggest uniform selection, leading to canalization in cavitation resistance for cultivated Juglans.

Trees; International audience; Citrus plants originate from southeastern Asia, in a large area with various climates characterized by a broad range of temperatures. Some species have been diversified in temperate climates, others in subtropical climates. Temperature is assumed to be a key factor in citrus species adaptation and diversification of basic cellular functions. In a field experiment, the tolerance of the three fundamental Citrus species C. medica L., C. reticulata Blanco and C. maxima (Burm.) Merr., and Fortunella japonica (Thunb.) Swing. to photooxidative stress caused by seasonal climatic changes was evaluated on adult trees by measuring net photosynthesis (Pnet), stomatal conductance (Gs), maximum photosynthesis (Pmax) and chlorophyll fluorescence (Fv/Fm). In addition, seasonal changes in oxidative status, antioxidant enzymes (superoxide dismutase, catalase, ascorbate peroxidase, monodehydroascorbate reductase, dehydroascorbate reductase and glutathione reductase) and antioxidant metabolites (ascorbate and glutathione) were monitored. Mandarin and pummelo appeared to be the most tolerant, showing the lowest down-regulation of photosynthetic parameters, and the lowest accumulation of oxidized compounds associated with efficiency of their antioxidant system. Kumquat showed intermediate behaviour, with a large diminution of photosynthetic parameters and marked accumulation of hydrogen peroxide, whereas the malondialdehyde content remained low, with a strong induction of glutathione synthesis. Finally, citron appeared to be the most sensitive genotype with a marked decrease in photosynthetic performance, the largest accumulation of oxidative parameters, insufficient induction of antioxidant enzymes and down-regulation of ascorbate and glutathione synthesis.

� Background and Aims The efficiency and safety functions of xylem hydraulics are strongly dependent on the pits that connect the xylem vessels. However, little is known about their biochemical composition and thus about their hydraulic properties. In this study, the distribution of the epitopes of different wall components (cellulose, hemicelluloses, pectins and lignins) was analysed in intervessel pits of hybrid poplar (Populus tremulaalba). � Methods Immunogold labelling with transmission electron microscopy was carried out with a set of antibodies raised against different epitopes for each wall polysaccharide type and for lignins. Analyses were performed on both immature and mature vessels. The effect of sap ionic strength on xylem conductance was also tested. � Key Results In mature vessels, the pit membrane (PM) was composed of crystalline cellulose and lignins. None of the hemicellulose epitopes were found in the PM. Pectin epitopes in mature vessels were highly concentrated in the annulus, a restricted area of the PM, whereas they were initially found in the whole PM in immature vessels. The pit border also showed a specific labelling pattern, with higher cellulose labelling compared with the secondary wall of the vessel. Ion-mediated variation of 24 % was found for hydraulic conductance. � Conclusions Cellulose microfibrils, lignins and annulus-restricted pectins have different physicochemical proper- ties (rigidity, hydrophobicity, porosity) that have different effects on the hydraulic functions of the PM, and these in- fluence both the hydraulic efficiency and vulnerability to cavitation of the pits, including ion-mediated control of hydraulic conductance. Impregnation of the cellulose microfibrils of the PM with lignins, which have low wettabil- ity, may result in lower cavitation pressure for a given pore size and thus help to explain the vulnerability of this species to cavitation.

eskimo1-5 (esk1-5) is a dwarf Arabidopsis (Arabidopsis thaliana) mutant that has a constitutive drought syndrome and collapsed xylem vessels, along with low acetylation levels in xylan and mannan. ESK1 has xylan O-acetyltransferase activity in vitro. We used a suppressor strategy on esk1-5 to screen for variants with wild-type growth and low acetylation levels, a favorable combination for ethanol production. We found a recessive mutation in the KAKTUS (KAK) gene that suppressed dwarfism and the collapsed xylem character, the cause of decreased hydraulic conductivity in the esk1-5 mutant. Backcrosses between esk1-5 and two independent knockout kak mutants confirmed suppression of the esk1-5 effect. kak single mutants showed larger stem diameters than the wild type. The KAK promoter fused with a reporter gene showed activity in the vascular cambium, phloem, and primary xylem in the stem and hypocotyl. However, suppression of the collapsed xylem phenotype in esk1 kak double mutants was not associated with the recovery of cell wall O-acetylation or any major cell wall modifications. Therefore, our results indicate that, in addition to its described activity as a repressor of endoreduplication, KAK may play a role in vascular development. Furthermore, orthologous esk1 kak double mutants may hold promise for ethanol production in crop plants.

To investigate the function of glutathione peroxidase (GPX) in plants, we produced transgenic tomato plants overexpressing an eukaryotic selenium-independent GPX (GPX5). We show here that total GPX activity was increased by 50% in transgenic plants, when compared to control plants transformed with the binary vector without the insert (PZP111). A preliminary two-dimensional electrophoretic protein analysis of the GPX overexpressing plants showed notably a decrease in the accumulation of proteins identified as rubisco small subunit I and fructose-1,6-bisphosphate aldolase, two proteins involved in photosynthesis. These observations, together with the fact that in standard culture conditions, GPX-overexpressing plants were not phenotypically distinct from control plants prompted us to challenge the plants with a chilling treatment that is known to affect photosynthesis activity. We found that upon chilling treatment with low light level, photosynthesis was not affected in GPX-overexpressing plants while it was in control plants, as revealed by chlorophyll fluorescence parameters and fructose-1,6-biphosphatase activity. These results suggest that overexpression of a selenium-independent GPX in tomato plants modifies specifically gene expression and leads to modifications of photosynthetic regulation processes.

Puroindolines are endosperm lipid binding proteins, which are separated by reversed phase-high-performance liquid chromatography or cation exchange chromatography into two isoforms, puroindoline-a (PIN-a) and puroindoline-b (PIN-b). Being very basic and close in molecular weight, PIN-a and PIN-b have never been separated using conventional isoelectric focusing and sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE). A two-dimensional electrophoresis method, linear immobiline pH gradient (IPGxSDS-PAGE), was developed, using 6-11 linear immobiline Dry Strips in the first dimension, which allowed the puroindolines to be focused between isoelectric point 10.5 and 11. Immunoblotting revealed that both PIN-a and PIN-b were each composed of several spots. Two-dimensional patterns from unrelated wheat varieties revealed that several spots can be highlighted among varieties. Matrix-assisted laser desorption/ionization-time of flight spectrometry allowed the majority of the spots revealed in the puroindoline zone to be identified. The two-dimensional IPGxSDS-PAGE of these very basic wheat endosperm proteins, puroindolines and related grain softness proteins should facilitate the identification of the proteins associated with wheat endosperm texture that have a strong effect on milling, dough properties and end-uses of wheats.

This study investigated the enzymatic function of two putative plant GPXs, GPXle1 from Lycopersicon esculentum and GPXha2 from Helianthus annuus, which show sequence identities with the mammalian phospholipid hydroperoxide glutathione peroxidase (PHGPX). Both purified recombinant proteins expressed in Escherichia coli show PHGPX activity by reducing alkyl, fatty acid and phospholipid hydroperoxides but not hydrogen peroxide in the presence of glutathione. Interestingly, both recombinant GPXle1 and GPXha2 proteins also reduce alkyl, fatty acid and phospholipid hydroperoxides as well as hydrogen peroxide using thioredoxin as reducing substrate. Moreover, thioredoxin peroxidase (TPX) activities were found to be higher than PHGPX activities in terms of efficiency and substrate affinities, as revealed by their respective Vmax and Km values. We therefore conclude that these two plant GPX-like proteins are antioxidant enzymes showing PHGPX and TPX activities.

MEDLINE:24918205; International audience; Background and Aims Various correlations have been identified between anatomical features of bordered pits in angiosperm xylem and vulnerability to cavitation, suggesting that the mechanical behaviour of the pits may play a role. Theoretical modelling of the membrane behaviour has been undertaken, but it requires input of parameters at the nanoscale level. However, to date, no experimental data have indicated clearly that pit membranes experience strain at high levels during cavitation events.Methods Transmission electron microscopy (TEM) was used in order to quantify the pit micromorphology of four tree species that show contrasting differences in vulnerability to cavitation, namely Sorbus aria, Carpinus betulus, Fagus sylvatica and Populus tremula. This allowed anatomical characters to be included in a mechanical model that was based on the Kirchhoff–Love thin plate theory. A mechanistic model was developed that included the geometric features of the pits that could be measured, with the purpose of evaluating the pit membrane strain that results from a pressure difference being applied across the membrane. This approach allowed an assessment to be made of the impact of the geometry of a pit on its mechanical behaviour, and provided an estimate of the impact on air-seeding resistance.Key Results The TEM observations showed evidence of residual strains on the pit membranes, thus demonstrating that this membrane may experience a large degree of strain during cavitation. The mechanical modelling revealed the interspecific variability of the strains experienced by the pit membrane, which varied according to the pit geometry and the pressure experienced. The modelling output combined with the TEM observations suggests that cavitation occurs after the pit membrane has been deflected against the pit border. Interspecific variability of the strains experienced was correlated with vulnerability to cavitation. Assuming that air-seeding occurs at a given pit membrane strain, the pressure predicted by the model to achieve this mechanical state corresponds to experimental values of cavitation sensitivity (P50).Conclusions The results provide a functional understanding of the importance of pit geometry and pit membrane structure in air-seeding, and thus in vulnerability to cavitation.

Intervessel pits are structures that play a key role in the efficiency and safety functions of xylem hydraulics. However, little is known about the components of the pit membrane (PM) and their role in hydraulic functions, especially in resistance to cavitation. We tested the effect of commercial chemicals including a cellulase, a hemicellulase, a pectolyase, a proteinase and DTT on xylem hydraulic properties: vulnerability to cavitation (VC) and conductance. The effects were tested on branch segments from Fagus sylvatica (where the effects on pit structure were analyzed using TEM) and Populus tremula. Cellulose hydrolysis resulted in a sharp increase in VC and a significant increase in conductance, related to complete breakdown of the PM. Pectin hydrolysis also induced a sharp increase in VC but with no effect on conductance or pit structure observable by TEM. The other treatments with hemicellulase, proteinase or DTT showed no effect. This study brings evidence that cellulose and pectins are critical components underpinning VC, and that PM components may play distinct roles in the xylem hydraulic safety and efficiency.

Plant species living in the understory increase carbon (C) allocation toward leaf production for maximizing light capture at the expense of roots and stems, with negative consequences for the whole-plant hydraulic conductance. Moreover, under some conditions, the high atmospheric evaporative demand occurring in Mediterranean areas may be not well buffered by the canopy, which might be the case for relict conifer Abies pinsapo Boiss. growing in the forest understory. We hypothesized that acclimation to combined understory shade and high atmospheric dryness can be achieved through the adjustment of water losses to cope with the restriction in water transport. The results reveal high structural plasticity in A. pinsapo that allows light harvesting of this species to maximize light capture in the forest understory, and maintain a positive C balance under low light conditions. However, growth in the understory resulted in reduced leaf-specific conductivity, up to approximately four to five times, implying decreased plant capacity to supply water to the leaves. In order to cope with the high atmospheric evaporative demand in the understory, there is an adjustment of the stomatal conductance to the hydraulic conductivity by means of a reduction in the stomatal density in understory individuals, which is due to the almost complete lack of stomata in the adaxial side of the needles. To the extent of our knowledge, such a drastic phenotypic response found in a conifer when growing under shaded conditions had not been previously reported. Published

†Background and Aims Extreme water stress episodes induce tree mortality, but the physiological mechanisms causing tree death are still poorly understood. This study tests the hypothesis that a potted tree’s ability to survive extreme monotonic water stress is determined by the cavitation resistance of its xylem tissue. †Methods Two species were selected with contrasting cavitation resistance (beech and poplar), and potted juvenile trees were exposed to a range of water stresses, causing up to 100 % plant death. †KeyResults The lethal dose of water stress, defined as the xylem pressure inducing 50 % mortality, differed sharply across species (1.75 and 4.5 MPa in poplar and beech, respectively). However, the relationships between tree mortality and the degree of cavitation in the stems were similar, with mortality occurring suddenly when .90 % cavitation had occurred. †Conclusions Overall, the results suggest that cavitation resistance is a causal factor of tree mortality under extreme drought conditions.

International audience; Gravity perception and gravitropic response are essential for plant development. In herbaceous species, it is widely accepted that one of the primary events in gravity perception involves the displacement of amyloplasts within specialized cells. However, the early signaling events leading to stem reorientation are not fully known, especially in woody species in which primary and secondary growth occur. Thirty-six percent of the identified proteins that were differentially expressed after gravistimulation were established as potential Thioredoxin targets. In addition, Thioredoxin h expression was induced following gravistimulation. In situ immunolocalization indicated that Thioredoxin h protein co-localized with the amyloplasts located in the endodermal cells. These investigations suggest the involvement of Thioredoxin h in the first events of signal transduction in inclined poplar stems, leading to reaction wood formation.

Times Cited: 0; Xylem cavitation resistance has profound implications for plant physiology and ecology. This process is characterized by a 'vulnerability curve' (VC) showing the variation of the percentage of cavitation as a function of xylem pressure potential. The shape of this VC varies from 'sigmoidal' to 'exponential'. This review provides a panorama of the techniques that have been used to generate such a curve. The techniques differ by (i) the way cavitation is induced (e.g. bench dehydration, centrifugation, or air injection), and (ii) the way cavitation is measured (e.g. percentage loss of conductivity (PLC) or acoustic emission), and a nomenclature is proposed based on these two methods. A survey of the literature of more than 1200 VCs was used to draw statistics on the usage of these methods and on their reliability and validity. Four methods accounted for more than 96% of all curves produced so far: bench dehydration-PLC, centrifugation-PLC, pressure sleeve-PLC, and Cavitron. How the shape of VCs varies across techniques and species xylem anatomy was also analysed. Strikingly, it was found that the vast majority of curves obtained with the reference bench dehydration-PLC method are 'sigmoidal'. 'Exponential' curves were more typical of the three other methods and were remarkably frequent for species having large xylem conduits (ring-porous), leading to a substantial overestimation of the vulnerability of cavitation for this functional group. We suspect that 'exponential' curves may reflect an open-vessel artefact and call for more precautions with the usage of the pressure sleeve and centrifugation techniques.

One adaptation of plants to cope with drought or frost stress is to develop wood that is able to withstand the formation and distribution of air bubbles (emboli) in its water conducting xylem cells under negative pressure. The ultrastructure of interconduit pits strongly affects drought-induced embolism resistance, but also mechanical properties of the xylem are involved. The first experimental evidence for a lower embolism resistance in stems of herbaceous plants compared to stems of their secondarily woody descendants further supports this mechanical-functional trade-off. An integrative approach combining (ultra)structural observations of the xylem, safety-efficiency aspects of the hydraulic pipeline, and xylem–phloem interactions will shed more light on the multiple adaptive strategies of embolism resistance in plants.

International audience; Despite the economic importance of citrus, insights on the genetic response to stress are scarce. The aim of the present study was to compare fundamental citrus species for their response to photooxidative stress. The experiment was conducted under orchard conditions on three fundamental citrus species C. medica L., C. reticulata Blanco and C. maxima (Burm.) Merr., and on Fortunella japonica (Thunb.) Swing. We examined their respective net photosynthesis (Pnet), stomatal conductance (Gs) and chlorophyll fluorescence (Fv/Fm) on sun-acclimated leaves and shade-acclimated leaves returned under natural sunlight irradiance. To compare the respective response mechanism, we analyzed changes in oxidative status (hydrogen peroxide (H2O2) and malondialdehyde (MDA)), reactive oxygen species (ROS)-scavenging enzymes (superoxide dismutase (SOD), catalase, ascorbate peroxidase), recycling enzymes (monodehydroascorbate reductase, dehydroascorbate reductase and glutathione reductase) and antioxidant metabolites (ascorbate and glutathione). Kumquat and pummelo exposed lower down-regulation and full recovery of photosynthetic parameters, lower accumulation of oxidized compounds associated with greater production of reduced glutathione (Gsh) and enhanced activity of the three ROS scavenging enzymes, especially SOD. Citron and mandarin showed a marked decrease and incomplete recovery in photosynthetic performance, mainly in Pnet and Fv/Fm, larger accumulation of oxidative parameters, slighter induction of antioxidant enzymes and down-regulation of reduced ascorbate (Asa) and Gsh synthesis. These results suggest that kumquat and pummelo have a greater tolerance to photooxidative stress than citron and mandarin.

Wood provides water transport and mechanical support of trees. Sap is transported under negative pressure in plant xylem conduits, which can be subject to embolism during severe drought. Typically, denser woods show thicker cell walls and stronger mechanical properties. Ten transgenic poplar lines modified for expression of genes involved in lignin metabolism were produced from the female clone 717-1B4 of Populus tremula × Populus alba to test the hypothesis of a possible trade-off between hydraulic and mechanical functions. Poplar lines underexpressed genes encoding for cinnamoyl alcohol dehydrogenase (CAD), cinnamoyl CoA reductase (CCR) and caffeic acid 3-O- methyltransferase (COMT), while new poplar lines underexpressed the CAD genes or overexpressed the MYB308 gene, encoding for a transcription factor repressing the phenylpropanoid metabolism. To maximize the contrast between line behaviors, these plants were grown under two different water regimes, and the impact on their hydraulic traits and xylem properties was analyzed to test for a link between water condition and mechanical and hydraulic properties. Our results show that the resistance to xylem cavitation was lower for the transgenic lines than for the control line 717-1b4 while they show neither a positive nor a negative tendency for the longitudinal Young's modulus between the transgenic lines and the control line. ASOMT10b and ASOMT2b, which possessed a down-regulated expression for all the genes, showed a lower value of the resistance to implosion index (t/b)2. No difference for xylem hydraulic conductivity between the lines was found. The changes in lignin metabolism in these transgenic lines did not affect the water transport, despite the change in the lignin content. Our data on the transgenic poplar lines do not therefore support the mechanical vs. hydraulic trade-off hypothesis and we point out that angiosperm trees have numerous ways to acclimate their internal structure in order to adjust their mechanical properties without hydraulic coupling. Moreover, we observed an acclimation to water stress for P50 but not for the Young's modulus. MYB308-25.1 showed better mechanical properties and vulnerability to cavitation than the control line 717-1b4. Finally, we present evidence that lignins are involved in the vulnerability to cavitation, probably through modifications of pit structure and behavior.

The function of selenium independent glutathione peroxidase (GPx) in response to biotic and abiotic stresses was investigated in transgenic tomato plants overexpressing an exogenous GPx and exhibiting a 50% increase in total GPx activity. GPx-overexpressing and control plants were challenged either by a mechanical stress or by infection with the biotrophic parasite Oidium neolycopersici or the necrotrophic parasite Botrytis cinerea. In mechanically stressed plants, internode growth was significantly less modified in GPx-overexpressing plants compared to controls. This stress resistant phenotype was not accompanied with any change in the global antioxidant response of the plants other than their increased GPx activity. Following infection by O. neolycopersici or by B. cinerea, lesion extension was increased in GPx-overexpressing plants compared with controls. These results showed that GPx overexpression provoked opposite effects in situations of biotic and abiotic challenges, suggesting a key role for this scavenger enzyme in controlling biotic and abiotic stress responses.

Xylem cavitation resistance is a key physiological trait correlated with species tolerance to extreme drought stresses. Little is known about the genetic variability and phenotypic plasticity of this trait in natural tree populations. Here we measured the cavitation resistance of 17 Fagus sylvatica populations representative of the full range of the species in Europe. The trees were grown in three field trials under contrasting climatic conditions. Our findings suggest that the genotypic variability of cavitation resistance is high between genotypes of a given population. By contrast, no significant differences were found for this trait across populations, the mean population cavitation resistance being remarkably constant in each trial. We found a significant site effect and a significant site×population interaction, suggesting that cavitation resistance has a high phenotypic plasticity and that this plasticity is under genetic control. The implications of our findings for beech forest management in a context of climate change are discussed. © The Author 2011.

The Cavitron spinning technique is used to construct xylem embolism vulnerability curves (VCs), but its reliability has been questioned for species with long vessels. This technique generates two types of VC: sigmoid 's'-shaped and exponential, levelling-off 'r'-shaped curves. We tested the hypothesis that 'r'-shaped VCs were anomalous and caused by the presence of vessels cut open during sample preparation. A Cavitron apparatus was used to construct VCs from samples of different lengths in species with contrasting vessel lengths. The results were compared with VCs obtained using other independent techniques. When vessel length exceeded sample length, VCs were 'r'-shaped and anomalous. Filling vessels cut open at both ends with air before measurement produced more typical 's'-shaped VCs. We also found that exposing segments of 11 woody species in a Cavitron at the pressure measured in planta before sampling considerably increased the degree of embolism above the native state level for species with long vessels. We concluded that open vessels were abnormally more vulnerable to cavitation than intact vessels. We recommend restricting this technique to species with short conduits. The relevance of our conclusions for other spinning techniques is discussed.

The Cavitron spinning technique is used to construct xylem embolism vulnerability curves (VCs), but its reliability has been questioned for species with long vessels. This technique generates two types of VC: sigmoid 's'-shaped and exponential, levelling-off 'r'-shaped curves. We tested the hypothesis that 'r'-shaped VCs were anomalous and caused by the presence of vessels cut open during sample preparation. A Cavitron apparatus was used to construct VCs from samples of different lengths in species with contrasting vessel lengths. The results were compared with VCs obtained using other independent techniques. When vessel length exceeded sample length, VCs were 'r'-shaped and anomalous. Filling vessels cut open at both ends with air before measurement produced more typical 's'-shaped VCs. We also found that exposing segments of 11 woody species in a Cavitron at the pressure measured in planta before sampling considerably increased the degree of embolism above the native state level for species with long vessels. We concluded that open vessels were abnormally more vulnerable to cavitation than intact vessels. We recommend restricting this technique to species with short conduits. The relevance of our conclusions for other spinning techniques is discussed.

Xylem vulnerability to cavitation is a key parameter in the drought tolerance of trees, but little is known about the control mechanisms involved. Cavitation is thought to occur when an air bubble penetrates through a pit wall, and would hence be influenced by the wall’s porosity. We first tested the role of wall-bound calcium in vulnerability to cavitation in Fagus sylvatica. Stems perfused with solutions of oxalic acid, EGTA, or sodium phosphate (NaPO 4 ) were found to be more vulnerable to cavitation. The NaPO 4 -induced increase in vulnerability to cavitation was linked to calcium removal from the wall. In contrast, xylem hydraulic conductance was unaffected by the chemical treatments, demonstrating that the mechanisms controlling vulnerability to cavitation and hydraulic resistance are uncoupled. The NaPO 4 solution was then perfused into stems from 13 tree species possessing highly contrasted vulnerability to cavitation. Calcium was found to be a major determinant of betweenspecies differences in vulnerability to cavitation. This was evidenced in angiosperms as well as conifer species, thus supporting the hypothesis of a common mechanism in drought-induced cavitation. In plants, long-distance sap transport occurs under negative pressures in xylem conduits. Sap flows between adjoining conduits through pits that form pores in the walls, and that facilitate the flow of water while preventing the passage of air bubbles. Under water stress conditions, xylem tensions increase and the conduits become vulnerable to cavitation. Cavitation provokes an air embolism that leads to a loss of hydraulic conductance, thus exacerbating plant water deficit. Species resistance to cavitation has been intensively studied over the past two decades, and is now ranked among the traits with the highest functional and ecological significance. In woody species for instance, xylem vulnerability to cavitation correlates tightly with species-specific drought tolerance (Pockman and Sperry, 2000; Tyree et al., 2003; Maherali et al., 2004), with more xerophilous species proving less vulnerable to cavitation. Substantial variations have also been found between genotypes of a different species (Cochard et al., 2007; Dalla-Salda et al., 2009). This implies that this trait could potentially be used in breeding programs to identify more drought-tolerant species or genotypes. However, efforts in this direction are still strongly impeded by a lack of understanding of the molecular and genetic basis of cavitation resistance. Our work represents a first significant step toward resolving this challenging issue. Understanding the fine mechanism of cavitation formation is a pivotal step toward identifying the key structures and the key genes coding for these structures, yet we currently have only partial insights. According to a hypothesis first formulated by Zimmermann (1983), water stress-induced cavitation

Xylem vulnerability to cavitation is a key parameter in understanding drought resistance of trees. We determined the xylem water pressure causing 50% loss of hydraulic conductivity (P(50)), a proxy of vulnerability to cavitation, and we evaluated the variability of this trait at tree and population levels for Fagus sylvatica. We checked for the effects of light on vulnerability to cavitation of stem segments together with a time series variation of P(50). Full sunlight-exposed stem segments were less vulnerable to cavitation than shade-exposed ones. We found no clear seasonal change of P(50), suggesting that this trait was designed for a restricted period. P(50) varied for populations settled along a latitudinal gradient, but not for those sampled along an altitudinal gradient. Moreover, mountainside exposure seemed to play a major role in the vulnerability to cavitation of beech populations, as we observed the differences along north-facing sides but not on south-facing sides. Unexpectedly, both north-facing mountainside and northern populations appeared less vulnerable than those grown on the southern mountainside or in the South of France. These results on beech populations were discussed with respect to the results at within-tree level.

Eight Hevea brasiliensis Muell. Arg. clones (GT1, YUNYAN77-4, IRCA707, IRCA317, PB217, PB260, PR107 and RRIM600) were compared for their tolerance towards chilling stress. Net photosynthesis (Pn), stomatal conductance (Gs), optimal and effective photochemical efficiencies (F(v)/F(m) and F'(v) = F'(m)), non-photochemical quenching, cellular lysis and leaf necrosis were measured on trees chilled at 10 degrees C for 96 h, as well as upon recovery at 28 degrees C. In addition, ascorbate peroxidase, catalase, dehydroascorbate reductase, glutathione reductase, monodehydroascorbate reductase and superoxide dismutase activities were monitored. Clone RRIM600 appeared to be the most tolerant, because it showed no cellular lysis or leaf necrosis and the best recovery as revealed by Pn, Gs, Fv/Fm and F'(v) = F'(m). Its ability to sustain chilling stress seemed related in part to the fast closure of stomata, suggesting an 'avoidance strategy' for this clone. IRCA707, GT1 and YUNYAN77-4 were also tolerant to the cold treatment as only a few leaf injuries were observed. However, YUNYAN77-4 showed a particular behaviour with a large stomata opening during the first hour of chilling, some photosynthetic activity after 96 h at 10 degrees C, but the slowest recovery in Pn. The greatest cell or leaf damage was observed on PB260, IRCA317, PR107 and PB217 clones, thus classified as sensitive to chilling. These clones showed the strongest decrease in Pn, F(v)/F(m) and F'(v) = F'(m) and the slowest recovery for F(v)/F(m) and F'(v) = F'(m), indicating a high sensitivity of photosystem II to cold temperatures. Punctual increases of various enzymatic activities were observed for all clones during chilling kinetics. During recovery, the strongest increases in enzymatic activity were observed for the most tolerant clones, suggesting that efficient reactive oxygen species elimination is a crucial step for determining chilling tolerance in Hevea although the enzymes implicated varied from one tolerant clone to another. This study points out contrasted strategies of the Hevea clones in copping with chilling stress and recovery.