Effects of prolonged drought on stem non-structural carbohydrates content and post-drought hydraulic recovery in Laurus nobilis L.: The possible link between carbon starvation and hydraulic failure.

Drought-induced tree decline is a complex event, and recent hypotheses suggest that hydraulic failure and carbon starvation are co-responsible for this process. We tested the possible role of non-structural carbohydrates (NSC) content on post-drought hydraulic recovery, to verify the hypothesis that embolism reversal represents a mechanistic link between carbon starvation and stem hydraulics. Measurements were performed in laurel plants subjected to similar water stress levels either over short or long term, to induce comparable embolism levels. Plants subjected to mild and prolonged water shortage (S) showed reduced growth, adjustment of turgor loss point driven by changes in both osmotic potential at full turgor and bulk modulus of elasticity, a lower content of soluble NSC and a higher content of starch with respect to control (C) plants. Moreover, S plants showed a lower ability to recover from xylem embolism than C plants, even after irrigation. Our data suggest that plant carbon status might indirectly influence plant performance during and after drought via effects on xylem hydraulic functioning, supporting the view of a possible mechanistic link between the two processes.

The contribution of vascular and extra-vascular water pathways to drought-induced decline of leaf hydraulic conductance.

Drought stress can impair leaf hydraulic conductance (Kleaf), but the relative contribution of changes in the efficiency of the vein xylem water pathway and in the mesophyll route outside the xylem in driving the decline of Kleaf is still debated. We report direct measurements of dehydration-induced changes in the hydraulic resistance (R=1/K) of whole leaf (Rleaf), as well as of the leaf xylem (Rx) and extra-vascular pathways (Rox) in four Angiosperm species. Rleaf, Rx, and Rox were measured using the vacuum chamber method (VCM). Rleaf values during progressive leaf dehydration were also validated with measurements performed using the rehydration kinetic method (RKM). We analysed correlations between changes in Rx or Rox and Rleaf, as well as between morpho-anatomical traits (including dehydration-induced leaf shrinkage), vulnerability to embolism, and leaf water relation parameters. Measurements revealed that the relative contribution of vascular and extra-vascular hydraulic properties in driving Kleaf decline during dehydration is species-specific. Whilst in two study species the progressive impairment of both vascular and extra-vascular pathways contributed to leaf hydraulic vulnerability, in the other two species the vascular pathway remained substantially unaltered during leaf dehydration, and Kleaf decline was apparently caused only by changes in the hydraulic properties of the extra-vascular compartment.

Diurnal changes in embolism rate in nine dry forest trees: relationships with species-specific xylem vulnerability, hydraulic strategy and wood traits.

Recent studies have reported correlations between stem sapwood capacitance (C(wood)) and xylem vulnerability to embolism, but it is unclear how C(wood) relates to the eventual ability of plants to reverse embolism. We investigated possible functional links between embolism reversal efficiency, C(wood), wood density (WD), vulnerability to xylem embolism and hydraulic safety margins in nine woody species native to dry sclerophyllous forests with different degrees of iso versus anisohydry. Substantial inter-specific differences in terms of seasonal/diurnal changes of xylem and leaf water potential, maximum diurnal values of transpiration rate and xylem vulnerability to embolism formation were recorded. Significant diurnal changes in percentage loss of hydraulic conductivity (PLC) were recorded for five species. Significant correlations were recorded between diurnal PLC changes and P50 and P88 values (i.e., xylem pressure inducing 50 and 88% PLC, respectively) as well as between diurnal PLC changes and safety margins referenced to P50 and P88. WD was linearly correlated with minimum diurnal leaf water potential, diurnal PLC changes and wood capacitance across all species. In contrast, significant relationships between P50, safety margin values referenced to P50 and WD were recorded only for the isohydric species. Functional links between diurnal changes in PLC, hydraulic strategies and WD and C(wood) are discussed.

Plant performance on Mediterranean green roofs: interaction of species-specific hydraulic strategies and substrate water relations.

Recent studies have highlighted the ecological, economic and social benefits assured by green roof technology to urban areas. However, green roofs are very hostile environments for plant growth because of shallow substrate depths, high temperatures and irradiance and wind exposure. This study provides experimental evidence for the importance of accurate selection of plant species and substrates for implementing green roofs in hot and arid regions, like the Mediterranean area. Experiments were performed on two shrub species (Arbutus unedo L. and Salvia officinalis L.) grown in green roof experimental modules with two substrates slightly differing in their water retention properties, as derived from moisture release curves. Physiological measurements were performed on both well-watered and drought-stressed plants. Gas exchange, leaf and xylem water potential and also plant hydraulic conductance were measured at different time intervals following the last irrigation. The substrate type significantly affected water status. Arbutus unedo and S. officinalis showed different hydraulic responses to drought stress, with the former species being substantially isohydric and the latter one anisohydric. Both A. unedo and S. officinalis were found to be suitable species for green roofs in the Mediterranean area. However, our data suggest that appropriate choice of substrate is key to the success of green roof installations in arid environments, especially if anisohydric species are employed.

Relax and refill: xylem rehydration prior to hydraulic measurements favours embolism repair in stems and generates artificially low PLC values.

Diurnal changes in percentage loss of hydraulic conductivity (PLC), with recorded values being higher at midday than on the following morning, have been interpreted as evidence for the occurrence of cycles of xylem conduits' embolism and repair. Recent reports have suggested that diurnal PLC changes might arise as a consequence of an experimental artefact, that is, air entry into xylem conduits upon cutting stems, even if under water, while under substantial tension generated by transpiration. Rehydration procedures prior to hydraulic measurements have been recommended to avoid this artefact. In the present study, we show that xylem rehydration prior to hydraulic measurements might favour xylem refilling and embolism repair, thus leading to PLC values erroneously lower than those actually experienced by transpiring plants. When xylem tension relaxation procedures were performed on stems where refilling mechanisms had been previously inhibited by mechanical (girdling) or chemical (orthovanadate) treatment, PLC values measured in stems cut under native tension were the same as those measured after sample rehydration/relaxation. Our data call for renewed attention to the procedures of sample collection in the field and transport to the laboratory, and suggest that girdling might be a recommendable treatment prior to sample collection for PLC measurements.

Coping with drought-induced xylem cavitation: coordination of embolism repair and ionic effects in three Mediterranean evergreens.

Embolism repair and ionic effects on xylem hydraulic conductance have been documented in different tree species. However, the diurnal and seasonal patterns of both phenomena and their actual role in plants' responses to drought-induced xylem cavitation have not been thoroughly investigated. This study provides experimental evidence of the ability of three Mediterranean species to maintain hydraulic function under drought stress by coordinating the refilling of xylem conduits and ion-mediated enhancement of stem hydraulic conductance (K stem). Vessel grouping indices and starch content in vessel-associated parenchyma cells were quantified to verify eventual correlations with ionic effects and refilling, respectively. Experiments were performed on stems of Ceratonia siliqua L., Olea europaea L. and Laurus nobilis L. Seasonal, ion-mediated changes in K stem (ΔK stem) and diurnal and/or seasonal embolism repair were recorded for all three species, although with different temporal patterns. Field measurements of leaf specific stem hydraulic conductivity showed that it remained quite constant during the year, despite changes in the levels of embolism. Starch content in vessel-associated parenchyma cells changed on diurnal and seasonal scales in L. nobilis and O. europaea but not in C. siliqua. Values of ΔK stem were significantly correlated with vessel multiple fraction values (the ratio of grouped vessels to total number of vessels). Our data suggest that the regulation of xylem water transport in Mediterranean plants relies on a close integration between xylem refilling and ionic effects. These functional traits apparently play important roles in plants' responses to drought-induced xylem cavitation.

Does citrus leaf miner impair hydraulics and fitness of citrus host plants?

Gas exchange and hydraulic features were measured in leaves of three different Citrus species (Citrus aurantium L., Citrus limon L., Citrus × paradisii Macfad) infested by Phyllocnistis citrella Staiton, with the aim to quantify the impact of this pest on leaf hydraulics and, ultimately, on plant fitness. Infested leaves were characterized by the presence on the leaf blade of typical snake-shaped mines and, in some cases, of a crumpled leaf blade. Light microscopy showed that leaf crumpling was induced by damage to the cuticular layer. In all three Citrus species examined: (a) the degree of infestation did not exceed 10% of the total surface area of infested plants; (b) control and infested leaves showed similar values of minimum diurnal leaf water potential, leaf hydraulic conductance and functional vein density; and (c) maximum diurnal values of stomatal conductance to water vapour, transpiration rate and photosynthetic rate (An) were similar in both control leaves and the green areas of infested leaves. A strong reduction of An was recorded only in mined leaf areas. Our data suggest that infestation with P. citrella does not cause conspicuous plant productivity reductions in young Citrus plants, at least not in the three Citrus species studied here.

Effects of NaCl addition to the growing medium on plant hydraulics and water relations of tomato.

This work reports on experimental evidence for the role of ion-mediated changes of xylem hydraulic conductivity in the functional response of Solanum lycopersicum L. cv. Naomi to moderate salinity levels. Measurements were performed in fully developed 12-week-old plants grown in half-strength Hoagland solution (control, C-plants) or in the same solution added with 35mM NaCl (NaCl-plants). NaCl-plants produced a significantly less but heavier leaves and fruits but had similar gas-exchange rates as control plants. Moreover, NaCl-plants showed higher vessel multiple fraction (FVM) than control plants. Xylem sap potassium and sodium concentrations were significantly higher in NaCl-plants than in control plants. When stems were perfused with 10mM NaCl or KCl, the hydraulic conductance of NaCl plants was nearly 1.5 times higher than in control plants. Accordingly, stem hydraulic conductance measured in planta was higher in NaCl- than in control plants. Our data suggest that tomato plants grown under moderate salinity upregulate xylem sap [Na+] and [K+], as well as sensitivity of xylem hydraulics to sap ionic content, thus, increasing water transport capacity.

Ion-mediated compensation for drought-induced loss of xylem hydraulic conductivity in field-growing plants of Laurus nobilis.

Xylem cavitation is a common occurrence in drought-stressed plants. Cavitation-induced embolism reduces xylem hydraulic conductivity (kxylem) and may lead to stomatal closure and reduction of photosynthetic rates. Recent studies have suggested that plants may compensate for kxylem loss through ion-mediated enhancement of the residual water transport capacity of functioning conduits. To test this hypothesis, field-grown laurel (Laurus nobilis L.) plants were subjected to mild drought stress by suspending irrigation. Drought treatment induced a significant increase in xylem embolism compared with control (well watered) plants. Xylem sap potassium concentration ([K+]) increased during the day both in control and water stressed plants. Midday values of sap [K+] were significantly higher in water stressed plants. The recorded increase in sap potassium concentration induced significant enhancement of residual kxylem when solutions with different [K+] were perfused through excised stems sampled in the field and measured in the laboratory. In planta measurements of stem hydraulic conductance revealed no change between water stressed plants and controls. Our data suggest that ion-mediated enhancement of residual kxylem buffered the actual loss of hydraulic conductance suffered by plants during the warmest hours of the day as well as under mild drought stress conditions.

Hydraulic connections of leaves and fruit to the parent plant in Capsicum frutescens (hot pepper) during fruit ripening.

BACKGROUND AND AIMS: The hydraulic architecture and water relations of fruits and leaves of Capsicum frutescens were measured before and during the fruiting phase in order to estimate the eventual impact of xylem cavitation and embolism on the hydraulic isolation of fruits and leaves before maturation/abscission. METHODS: Measurements were performed at three different growth stages: (1) actively growing plants with some flowers before anthesis (GS1), (2) plants with about 50 % fully expanded leaves and immature fruits (GS2) and (3) plants with mature fruits and senescing basal leaves (GS3). Leaf conductance to water vapour as well as leaf and fruit water potential were measured. Hydraulic measurements were made using both the high-pressure flow meter (HPFM) and the vacuum chamber (VC) technique. KEY RESULTS: The hydraulic architecture of hot pepper plants during the fruiting phase was clearly addressed to favour water supply to growing fruits. Hydraulic measurements revealed that leaves of GS1 plants as well as leaves and fruit peduncles of GS2 plants were free from significant xylem embolism. Substantial increases in leaf petiole and fruit peduncle resistivity were recorded in GS3 plants irrespective of the hydraulic technique used. The higher fraction of resistivity measured using the VC technique compared with the HPFM technique was apparently due to conduit embolism. CONCLUSIONS: The present study is the first to look at the hydraulics of leaves and fruits during growth and maturation through direct, simultaneous measurements of water status and xylem efficiency of both plant regions at different hours of the day.

Leafminers help us understand leaf hydraulic design.

Leaf hydraulics of Aesculus hippocastanum L. were measured over the growing season and during extensive leaf mining by the larvae of an invasive moth (Cameraria ohridella Deschka et Dimic) that specifically destroy the palisade tissue. Leaves showed seasonal changes in hydraulic resistance (R(lamina)) which were related to ontogeny. After leaf expansion was complete, the hydraulic resistance of leaves and the partitioning of resistances between vascular and extra-vascular compartments remained unchanged despite extensive disruption of the palisade by leafminers (up to 50%). This finding suggests that water flow from the petiole to the evaporation sites might not directly involve the palisade cells. The analysis of the temperature dependence of R(lamina) in terms of Q(10) revealed that at least one transmembrane step was involved in water transport outside the leaf vasculature. Anatomical analysis suggested that this symplastic step may be located at the bundle sheath where the apoplast is interrupted by hydrophobic thickening of cell walls. Our findings offer some support to the view of a compartmentalization of leaves into well-organized water pools so that the transpiration stream would involve veins, bundle sheath and spongy parenchyma, while the palisade tissue would be largely by-passed with the possible advantage of protecting cells from short-term fluctuations in water status.

Hydraulic characteristics and water relations in pigment-less mutant shoots of an orange tree.

Phenotypes more or less deficient in photosynthetic pigments show reduced productivity. Not much is known, however, about the influence of pigment-less twigs on the water balance of whole trees. We studied the water relations and hydraulic properties of normal and pigment-less (white) and 1-year-old shoots of 12-year-old Citrus sinensis L. trees. Compared with green leaves, white leaves showed a pronounced deficiency of pigments, higher stomatal density, the absence of chloroplasts in the guard cells and a different organization of leaf parenchyma. Stomatal conductance (gL) and transpiration rate (EL) were higher in white leaves than in green leaves during the hottest hours of the day, especially in July and September. The absence of chloroplasts in the stomatal guard cells seemed to be one of the factors causing insufficient stomatal control. Hydraulic conductance (KL) was higher in white leaves than in green leaves (16.96+/-2.24x10(-5) versus 11.26+/-0.66x10(-5) kg s-1 m-2 MPa). The ratio between the sum of the fourth power of xylem conduit radius (Sigmar4) (which determines theoretical conductance) and the total leaf area supplied by petioles and midribs was higher in white leaves than in green leaves. This was because of a smaller leaf area in white leaves and a statistically different distribution of lumen diameters of the conduits in midribs and petioles. The hydraulic properties of white twigs profoundly disturbed the water balance and physiology of the whole tree.

Impairment of leaf hydraulics in young plants of Citrus aurantium (sour orange) infected by Phoma tracheiphila.

Phoma trachephila (Petri) Kantschaveli et Gikachvili causes dieback of several Citrus species. The impact of this fungus on leaf hydraulics was studied in Citrus aurantium L. (sour orange) with the aim of identifying the primary mechanism of damage to leaves. Leaves inoculated with a conidial suspension were measured for conductance to water vapor (gL) and specific hydraulic conductance (Kleaf) every 3 days after inoculation. The earliest symptom of infection consisted of vein chlorosis. Functional vein density (FVD) was monitored and microscopic observations were made of major vein conduits. Impairment of vein hydraulics started 25 days after inoculation with a losses of Kleaf of 40% and gL of ~60%. Most minor veins within chlorotic areas were no longer functioning and some conduits of the major veins showed digested interconduit pits leading to vein cavitation. The close Kleaf-FVD relationship revealed that vein impairment caused drop of Kleaf and, consequently, of gL at chlorotic areas. Leaf infection was focused to veins that were first forced to embolise and then invaded by fungal hyphae. The vein embolism due to the Phoma amplifies the native dominant hydraulic resistance of leaf veins, and leads ultimately to early shedding of infected leaves.

Is rootstock-induced dwarfing in olive an effect of reduced plant hydraulic efficiency?

We investigated the hydraulic architecture of young olive trees either self-rooted or grafted on rootstocks with contrasting size-controlling potential. Clones of Olea europea L. (Olive) cv 'Leccino' inducing vigorous scion growth (Leccino 'Minerva', LM) or scion dwarfing (Leccino 'Dwarf', LD) were studied in different scion/rootstock combinations (LD, LM, LD/LD, LM/LM, LD/LM and LM/LD). Shoots growing on LD root systems developed about 50% less leaf surface area than shoots growing on LM root systems. Root systems accounted for 60-70% of plant hydraulic resistance (R), whereas hydraulic resistance of the graft union was negligible. Hydraulic conductance (K = 1/R) of LD root systems was up to 2.5 times less than that of LM root systems. Total leaf surface area (A(L)) was closely and positively related to root hydraulic conductance so that whole-plant hydraulic conductance scaled by A(L) did not differ between experimental groups. Accordingly, maximum transpiration rate and minimum leaf water potential did not differ significantly among experimental groups. We conclude that reduced root hydraulic conductance may explain rootstock-induced dwarfing in olive.

Kinetics of recovery of leaf hydraulic conductance and vein functionality from cavitation-induced embolism in sunflower.

The kinetics of leaf vein recovery from cavitation-induced embolism was studied in plants of sunflower cv. Margot, together with the impact of vein embolism on the overall leaf hydraulic conductance (Kleaf). During the air-dehydration of leaves to leaf water potentials (Psi L) of -1.25 MPa, Kleaf was found to decrease by about 46% with respect to values recorded in well-hydrated leaves. When leaves, previously dehydrated to Psi L= -1.1 MPa (corresponding to the turgor loss point), were put in contact with water, Kleaf recovered completely in 10 min and so did leaf water potential. Functional vein density was estimated in both dehydrating and rehydrating leaves in terms of total length of red-stained veins infiltrated with a Phloxine B solution per unit leaf surface area. Veins were found to embolize (unstained) with kinetics showing a linear relationship with Kleaf so that about a 70% loss of functional veins corresponded with a Kleaf loss of 46%. Cavitated veins recovered from embolism within 10 min from the beginning of leaf rehydration. These data indicate that: (a) leaves of sunflower underwent substantial vein embolism during dehydration; (b) vein embolism and leaf hydraulic efficiency apparently recovered from dehydration completely and rapidly upon rehydration; (c) vein refilling occurred while conduits were still at more negative xylem pressures than those required for spontaneous bubble dissolution on the basis of Henry's law. The possible consistent contribution of vital mechanisms for vein refilling is discussed.