Plant Science 172 (2007) 473–480 www.elsevier.com/locate/plantsci
Effects of sodium chloride on water potential components, hydraulic conductivity, gas exchange and leaf ultrastructure of Arbutus unedo plants A. Navarro c, S. Ban˜on a,c, E. Olmos b, M.J. Sa´nchez-Blanco b,c,* a
Departamento de Produccio´n Vegetal, Universidad Polite´cnica de Cartagena, 30203 Cartagena, Spain Centro de Edafologı´a y Biologı´a Aplicada del Segura (CSIC), P.O. Box 164, E-30100 Murcia, Spain c Horticultura Sostenible en Zonas A´ridas, Unidad Asociada al CSIC-CEBAS, Spain
b
Received 25 April 2006; received in revised form 9 October 2006; accepted 17 October 2006 Available online 7 November 2006
Abstract The purpose of this study was to evaluate the physiological and anatomical changes that occur in Arbutus unedo plants under saline conditions in order to understand the response of this species to salinity. A. unedo plants were grown in a greenhouse and submitted to three irrigation treatments using solutions containing 0, 52, and 105 mM NaCl with an electrical conductivity of 0.85 dS m1 (control treatment), 5.45 dS m1 (S.1) and 9.45 dS m1 (S.2). After 16 weeks, the leaf water relations, root hydraulic conductivity, gas exchange, ion concentrations and leaf ultrastructure were determined. Salinity induced a significant decrease in total biomass, leaf area and plant height. The concentration of Cl in leaves increased with increasing salinity and was higher than the corresponding concentration of Na+. Net photosynthesis (Pn) was reduced and the chloroplast ultrastructure was altered by salinity. Thylakoids were dilated and the number of plastoglobuli was greatly increased in both saline treatments compared with the control leaves. In addition, a reduction in the intercellular spaces of the lagunar mesophyll was observed in the saline treatments, affecting stomatal and mesophyll conductance to CO2. Root hydraulic resistance increased under saline conditions, affecting the water flow the root system. Pressure–volume analysis revealed osmotic adjustment values of 0.2 MPa at 52 mM and 0.5 MPa at 105 mM of NaCl, accompanied by 31 and 99% increases in the bulk tissue elastic modulus (e, wall rigidity) and resulting in turgor loss at the same relative water content in control and at 5.45 dS m1 and a higher relative water content at 9.45 dS m1. Osmotic adjustment and a high e together are seen as an effective means of counteracting the negative effects of salinity on the water balance of A. unedo plants. # 2006 Elsevier Ireland Ltd. All rights reserved. Keywords: Arbutus unedo; Elastic modulus; Leaf anatomy; Osmotic adjustment; Saline ions; Water relations
1. Introduction Low quality water is frequently used for irrigation in dry climates, because the intensive use of scarce water resources has increased the problems of salinity in the soil and in ground water. Increased salinity has an inverse relationship with net photosynthesis rate and dry matter production [1]. The growth of salt-treated plants is often limited by the ability of roots to extract water from the soil and transport it to the shoot due to
* Corresponding author at: Centro de Edafologı´a y Biologı´a Aplicada del Segura (CEBAS-CSIC), P.O. Box 164, E-30100 Murcia, Spain. Tel.: +34 968 396318; fax: +34 968 396213. E-mail address:
[email protected] (M.J. Sa´nchez-Blanco). 0168-9452/$ – see front matter # 2006 Elsevier Ireland Ltd. All rights reserved. doi:10.1016/j.plantsci.2006.10.006
the osmotic component of salinity [2]. Also, the quantity of water moving from the root to the shoot and its speed determine the concentration of substances reaching the shoot [3]. Under saline conditions, a low external water potential can be remedied by the uptake of electrolytes but may result in a gradual accumulation in the aerial parts, causing damage to plant metabolism, when no compartmentation of saline ions in the vacuole takes place. In species that maintain a positive water balance when subjected to salinity, osmotic adjustment maintains the positive turgor required for stomata opening and cell enlargement [4]. However, even if cell shrinkage occurs, the process of osmotic adjustment appears beneficial during cellular dehydration. The maintenance of turgor at a lower relative water volume tissue may be related also with elastic adjustment, preventing
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mechanical damage to plasma membranes [5]. A number of researchers have studied the water relations and osmotic and elastic adjustment of different crops under saline stress [6–9]. Nevertheless, the effects of this stress on tissue elasticity are not clear, with increases reported in some species [10,11] and decreases observed in others [12], some authors even suggesting that elastic adjustment does not play a role in the adaptation mechanism [13]. The use of native species of wild flora both in gardening and landscaping is of increasing interest because their characteristics and potential adaptation to adverse environmental conditions: drought, high day and low night temperatures, and salinity [6,14,15]. Among these, Arbutus unedo, an evergreen sclerophyll shrub, widely distributed in the Mediterranean Basin, is considered a good alternative for revegetation and restoration projects because it responds well to the conditions of Mediterranean summers, high temperature, low humidity and water stress at midday by closing its stomata [16]; it also shows intense vegetative activity in spring, when the conditions are not so restrictive. However, the changes in plant water relations that help maintain water uptake and any anatomical and ultrastructural modifications in response to salt stress have been little studied in A. unedo and Mediterranean ornamental species in general. Therefore, in the present study, the effects of NaCl on cell wall elasticity, osmotic adjustment, gas exchange, hydraulic conductivity and morphology were evaluated in A. unedo plants to ascertain the physiological and anatomical changes that occur in this species under salinity conditions.
2. Materials and methods 2.1. Plant material and treatments A. unedo L. (native of the province of Murcia, SE Spain) seedlings of an age sap (6 months to 1 year) were used. Seedlings were planted individually in 20 cm high pots, measuring 225 cm2 at the top and 121 cm2 at the base, containing a substrate of black peat, sand and a clay–loam soil (1:1:1) amended with osmocote plus (2 g L1 substrate). The experiment was conducted from December 2002 to May 2003 in a plastic greenhouse equipped with a cooling system, using drip irrigation with one emitter per plant, each delivering 3 L h1. During the experiment maximum/minimum average temperatures were 33/8 8C and the relative humidity ranged between 24 and 96%. The average maximum photosynthetically active radiation (PAR) was 805 mmol m2 s1. When they were 2 months old, 150 plants were submitted to three treatments, using irrigation water containing 0, 52, and 105 mM NaCl (control, S.1 and S.2 treatments, respectively) for 16 weeks. During the experimental period the average electrical conductivities of the irrigation solutions were 0.85, 5.45 and 9.45 dS m1, respectively. The irrigation water was applied in such a way as to maintain the electrical conductivity of the drainage water at about 10% EC of the irrigation water supplied for each treatment.
The experiment was laid out in a randomized block design, with two replicates per treatment. 2.2. Measurements of growth At the end of the experiment, the soil was gently washed from roots, and the plants were divided into shoots (stems and leaves) and roots. These were oven dried at 70 8C until they reached a constant mass to measure the respective dry weights. Ten plants per replicate were harvested and their height was measured. Leaf area was also measured using a Leaf Area Meter (DT Digital-Devices Ltd., Cambridge, England). 2.3. Determination of inorganic ions (Cl and Na+) The concentration of Cl was analysed by a chloride analyzer (Chloride Analyser Model 926, Sherwood Scientific Ltd.) in the aqueous extracts obtained when mixing 100 mg of dry vegetable powder with 40 ml of water, shaking later during 30 min and filtering. The concentration of Na+ was determined by an atomic absorption spectrometry (Shimadzu mod. AA6701, Shimadzu Co.) in a digestion extract with HNO3:HClO4 (2:1, v/v). Both were measured in the leaves of 10 plants per treatment at the end of the experimental period. 2.4. Transmission electron microscopy Leaf samples were fixed for 2.5 h at 4 8C in a 0.1 M Na phosphate-buffered (pH 7.2) mixture of 2.5% glutaraldehyde and 4% paraformaldehyde [17]. Tissue was post-fixed with 1% osmium tetroxide for 2 h. The samples were then dehydrated in a graded alcohol series and embedded in resin Spurr [18]. Blocks were sectioned on a Reichert ultramicrotome. Ultrathin sections (60–70 nm) for electron microscopy were placed on copper grids and stained with uranyl acetate followed by lead citrate. The ultrastructure of the tissue was observed with Zeiss EM10 and Zeiss EM109 electron microscopes. Semi-thin sections (0.5 mm) from the same blocks were stained with toluidine blue and examined with a Leica DMR light microscope. For morphometric analysis, a minimum of 10 leaves per treatment were studied. Parameters were measured using a Leica QM500 imaging analysis, as described previously by Olmos and Hellin [19]. 2.5. Water relations Predawn and midday leaf water potential (Cpd and Cmd), predawn and midday leaf osmotic potential (Cspd and Csmd), predawn and midday leaf turgor potential (Cppd and Cpmd), midday abaxial leaf conductance (gs), and midday net photosynthesis (Pn) were measured at the end of the experimental period in 10 plants per treatment. The leaf water potential was estimated according to the method described by Scholander et al. [20], using a pressure chamber (Soil Moisture Equipment Co, Santa Barbara, CA, USA), for which leaves were enclosed in a plastic bag and
A. Navarro et al. / Plant Science 172 (2007) 473–480
sealed in the chamber within 20 s of collection and pressurised at a rate of 0.02 MPa s1 [21]. Leaves from the (Cpd and Cmd) measurements were frozen in liquid nitrogen. After thawing, the sap was extracted from the sample with a small press, and them placed on a filter paper disc in the osmometer chamber and the values of the osmotic potential (Cspd and Csmd) was measured using a Wescor 5520 vapour pressure osmometer (Wescor Inc., Logan, UT, USA) according to Gucci et al. [22]. Estimates of leaf turgor potential (Cppd and Cpmd) were based on the difference between leaf water potential and leaf osmotic potential for each time. Stomatal conductance (gs) and the net photosynthetic rate (Pn) were determined on the same day and in the same plants as leaf water potential, using a gas exchange system (LI-6400, LICOR Inc., Lincoln, NE, USA). Measurements were made at midday on attached leaves. Root hydraulic conductivity (Lp) was measured in 10 plants per treatment according to Ramos and Kaufmann [23]. Plants were detopped with a razor blade and the soil was carefully washed away from the roots, which were immediately submerged in a container of water and placed in the pressure chamber with the cut stump exposed for 30 min. After a good seal was obtained, the air pressure was increased at an approximate rate of 0.4 MPa min1 up to a final pressure of 0.8 MPa. A small piece of plastic tubing was fitted to the stump, and every 5 min the exudate was collected and its volume measured. After the exudation measurements, the root systems were placed in an oven to 80 8C until they reached a constant mass (dry weight). Hydraulic root conductivity was calculated using the formula: Lp ¼
J ðP WÞ
where Lp is expressed in mg g1 s1 MPa1, P the applied hydrostatic pressure (MPa), W the dry weight of the root system (g) and J is the water flow rate through the entire root system (mg s1). Estimates of the bulk modulus of elasticity (e) at 100% RWC, osmotic potential at full turgor (C100s), and zero turgor (C0s), relative water content at zero turgor (RWC0) were obtained in three leaves per plant and 10 plants per treatment, via pressure–volume analysis of leaves, as outlined by Wilson et al. [24]. Bulk modulus of elasticity (e) at 100% RWC was calculated using the formula: e¼
ðRWC0 c100s Þ ð100 RWC0 Þ
where e is expressed in MPa, C100s the osmotic potential at full turgor (MPa) and RWC0 is the relative water content at zero turgor. Leaves were excised in the dark, placed in plastic bags and allowed to reach full turgor by dipping the petioles in distilled water overnight. Pressure–volume curves were obtained from periodic measurements of leaf weight and balance pressure as leaves dried on the bench at constant temperature of 20 8C. Drying-leaves period in each curve was about 3–5 h.
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2.6. Soil water content Substrate humidity was checked monthly by measuring the volumetric content of water in the substrate (uv) in the top 15 cm, by means of the time domain reflectometry technique (TDR) [25,26] using a TEKTRONIX 1502C in 10 replicates per treatment. The TDR method is based on the determination of the dielectric constant of the soil (Ka), which is related with the soil water content [27]. The direct in field manual analysis of the image that appears on the screen of the reflectometer allows obtaining the value of Ka before, uv (cm3/cm3) is calculated using the empiric equation: uv ¼ 5:3 102 þ 2:92 102 K a þ 5:5 104 Ka2 þ 4:3 106 Ka3 2.7. Statistical analysis The data were analysed by one-way ANOVA using Statgraphics Plus for Windows. Treatment means were separated with Tukey HSD’s multiple range test (P 0.05). 3. Results 3.1. Growth and mineral content The biomass data of A. unedo are shown in Table 1. Salinity induced a significant decrease in total plant biomass of 32% at 52 mM and 39% at 105 mM, in leaf area of 50% at 52 mM and 43% at 105 mM, and in height a reduction of 26% at 52 mM and 25% at 105 mM. Nevertheless, the differences between saline treatments were not statistically significant (Table 1). Visual symptoms of leaf necrosis appeared in plants from 52 and 105 mM treatment during the experimental period, being the level of injury smaller than 25% for 52 mM treatment and among 25–75% for the 105 mM. A tendency for the leaf dry weight/fresh weight ratio to increase in saline treatments was also observed (Table 1). At the end of the experimental period, salinity had led to a significant increase in Na+ and Cl ions levels in the leaf tissues of the treated plants (Fig. 1A and B). The concentration of Cl was higher than that of Na+ and increased with increasing salinity, reaching values of approximately 640 and Table 1 Total dry biomass (g plant1), height (cm), leaf area (cm2) and leaf dry weight/ fresh weight ratio, measures at the end of the experimental period in A. unedo seedlings, subjected to different saline treatments Treatments
Total dry biomass
Height
Leaf area
Leaf DW/ FW ratio
Control S.1. S.2.
5.10 b 3.46 a 3.12 a
21.23 b 15.73 a 15.90 a
5.63 b 2.76 a 3.22 a
0.37 a 0.39 a 0.44 a
Means within a column without a common letter are significantly different by Tukey0.05 test. Each value is the mean of 10 plants per treatment.
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Fig. 1. Contents of Na+ (A) and Cl (B) in leaves of Arbutus unedo at the end of the experimental period in control, S.1. and S.2. treatments. Each histogram represents the mean of 10 values and the vertical bars indicate standard errors.
910 mmol kg1 DW for 52 and 105 mM treatments, respectively. The sodium concentration was similar in the leaves of both saline treatments, reaching levels of about 157 mmol kg1 DW (Fig. 1A). 3.2. Leaf ultrastructure Chloroplast size was estimated as the length of the chloroplast cross-sections. No significant difference in the size of chloroplasts between treatments was found (Table 2), although, chloroplast ultrastructure was significantly altered. Thylakoids were dilated and the number of plastoglobuli was substantially higher in both saline treatments than in the control leaves (Table 2), showing a high increase of relative area filling the stroma of chloroplasts while the size of plastoglobuli increased more than three fold in the 105 mM treatment compared with 52 mM and control treatments (Table 2). The morphometrical data from cross-sections of control leaves pointed to internal anatomy with a well-defined palisade formed by two layer of cells, as well as a lagunar mesophyll four to five cells thick. A comparison of control plants with saline treated plants showed that the cell size of the first layer of palisade cells was not altered significantly (Table 2). However, the cell size of the second layer of palisade cells significantly increased with increased levels of salinity, with values of 777, 931 and 1387 mm2 for control, 52 and 105 mM treatments, respectively (Table 2). We also observed a substantial reduction in the intercellular spaces in the lagunar mesophyll in both
saline treatments (52 and 105 mM of NaCl) compared with control leaves (Table 2). 3.3. Water relations In salinized plants, predawn and midday leaf water potentials (Cpd and Cmd) were lower than in control plants (Fig. 2A and B). The Cpd values for both 52 and 105 mM treatments decreased from the beginning of salt application onwards. S.1 treatment had constant values throughout the experimental period. At 105 mM treatment values were lower than those of the 52 mM treatment and there was a decrease of Cpd at the third sampling time. Midday leaf water potential (Cmd) values showed little differences between treatments. Leaf osmotic potential (Fig. 2C and D) behaved in a similar way to Cpd and Cmd. Salinity had a significant effect on Cppd and Cpmd, both of which increased in salinized plants with respect to the control plants. At the end of experimental period, plants treated with 105 mM NaCl showed the highest leaf turgor potential values at predawn and midday (Fig. 2E and F). Parameters derived from the pressure–volume curve are shown in Table 3. At the end of experimental period, leaf osmotic potential values at full turgor (C100s) decreased in treated plants, which was indicative of the osmotic adjustment that occurred due to the salinity, the difference between the values obtained in the control and salinized plants were taken as an estimate of this adjustment (0.2 and 0.5 MPa for 52 and 105 mM treatments, respectively). Osmotic potential at turgor loss point (C0s) and relative water content at turgor loss point
Table 2 Quantitative analysis of morphometric data measurements at the end of the experimental period in leaves of A. unedo seedlings, subjected to different saline treatments Treatments
Chloroplast size, N = 15 (mm2)
Plastoglobuli size, N = 60 (mm2)
No. of plastglobuli, N = 40
Plast/Chl area, N = 15 (%)
Intercellular space, N = 10 (%)
Palisade cell (1) area, N = 40 (mm2)
Palisade cell (2) area, N = 40 (mm2)
Control S.1. S.2.
8.6 a 8.5 a 9.1 a
0.024 b 0.022 b 0.076 a
5.0 b 27.3 a 20.8 a
1.7 c 7.4 b 11.9 a
37.0 b 23.3 a 22.3 a
861 a 1051 a 982 a
777 c 931 b 1387 a
Means within a column without a common letter are significantly different by Tukey0.05 test. Palisade parenchyma is composed of two cell layers: (1) first layer of palisade cells in contact with epidermis. (2) Second layer of palisade cells in contact with lagunar mesophyll.
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Fig. 2. Leaf water potential (A and B), leaf osmotic potential (C and D) and leaf turgor potential (E and F) at predawn (A, C and E) and midday (B, D and F) after application of different saline treatments. (*) Control plants; ( ) S.1. (5.45 dS m1); ( ) S.2. (9.45 dS m1). Values are means (n = 10) and the vertical bars indicate standard errors.
(RWC0) of A. unedo were significantly affected by the highest NaCl treatment (Table 3). In this treatment C0s was lower and the point of zero turgor (RWC0) occurred at higher relative water content (87%) than in the control (80%). The values of bulk modulus of elasticity (e) increased with increasing salinity (Table 3). The response of hydraulic conductance (Lp) differed between control and saline treatments (Table 3), Lp decreasing proportionally to the saline concentration applied. Variations in the volumetric water content in the substrate of the pots during
the experimental period are shown in Fig. 3. A significant increase in the last sampling time was observed as the external salinity increased. Changes in stomatal conductance (gs) and the photosynthetic rate (Pn) are shown in Fig. 4A and B. In plants from both saline treatments the gs values decreased from the outset and significant differences between treatments were evident. The Pn also decreased in salinized plants, although the differences between treatments were lower than in the case of gs but more marked at the end of experimental period.
Table 3 Effects of the salinity on the osmotic potential at full turgor (C100s), and zero turgor (C0s), relative water content at zero turgor (RWC0), bulk modulus of elasticity (e) at 100% RWC and root hydraulic conductivity (Lp) measured at the end of the experimental period in A. unedo seedlings Treatments
c100s (MPa)
c0s (MPa)
RWC0 (%)
e (MPa)
Lp (mg s1 MPa1 g1)
Control S.1. S.2.
1.59 a 1.79 a,b 2.11 b
2.34 a 2.31 a 2.91 b
79.60 a 79.36 a 87.39 b
6.21 a 8.12 b 12.38 c
0.40 c 0.07 b 0.03 a
Means within a column without a common letter are significantly different by Tukey0.05 test. Each value is the mean of 10 plants per treatment.
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Fig. 3. Soil water content (TDR volumetric porcentage) after application of different saline treatments. (*) Control plants; ( ) S.1. (5.45 dS m1); ( ) S.2. (9.45 dS m1). Values are means (n = 10) and the vertical bars indicate standard errors.
Fig. 4. Net photosynthetic rate (Pn, A) and stomatal conductance (gs) after application of different saline treatments. (*) Control plants; ( ) S.1. (5.45 dS m1); ( ) S.2. (9.45 dS m1). Values are means (n = 10) and the vertical bars indicate standard errors.
4. Discussion The morphological differences in A. unedo due to exposure saline conditions were in terms of reduction in total biomass, height and leaf area (Table 1), perhaps, induced by the excess of
Na+ and Cl ions accumulated in the leaves (Fig. 1). The increase in Cl and Na+ concentrations in leaves, in addition to reducing whole leaf assimilation, also reduced leaf expansion with a subsequent decrease in leaf area and presumably in height plant, a response known from many ornamental plants [4,9,28,29]. The results of this study are consistent with the findings of Munns and Termaat [30], which suggested that this accumulation of Na+ and Cl ions is one of the causes of photosynthetic decline and therefore productivity falls below a level capable of sustaining further growth. As indicated in the results, the photosynthesis rates were significantly reduced 2 months after the saline application at 52 mM (20%) and even more so at 105 mM (40%) (Fig. 4B). It seems that the harmful consequences of NaCl salinity on Pn is associated with high leaf Cl values, since the reductions in Pn differed between both saline treatments as did the leaf Cl concentrations found [31]. This inhibition of photosynthesis in our experiment was reflected in inhibition of growth, indicating that photosynthesis could be the growth-limiting factor [4]. The number and size of plastoglobuli increase substantially in leaf chloroplasts (Table 2) following exposure to salinity [28,32] and drought [33]. It is assumed that plastoglobuli have a function in the storage of thylakoid components such as lipids, plastohydroquinone and tocopherol [34]. Recently, proteomic analysis of plastoglobuli has demonstrated that plastoglubuli in Arabidopsis contain several proteins involved in the metabolism of isoprenoid derived molecules, of lipids and carotenoids cleavage [35]. These authors suggested that plastoglubuli may play a role in the synthesis and recycling of lipophilic products and oxidative stress defense. Furthermore, it is well documented that chloroplast ultrastructure is altered by salinity [36], usually before other cell organelles. In addition, a reduction in the intercellular spaces in the lagunar mesophyll of the saline treated plants was observed [37]. These changes in mesophyll anatomy could have accounted for the decreased mesophyll conductance [38]. In this sense, salinity could reduce photosynthesis in the leaves by reductions in stomatal and by changes in mesophyll structure, which decreased the conductance to CO2 within the leaf [39]. At the last sampling time, the root hydraulic resistance markedly increased under saline conditions (Table 3). Some researchers have found that long-term exposure to sodium chloride affect the root permeability [40]. Such variations in root hydraulic conductivity as a result of saline stress affected the water flowing through the root system as could be seen from the soil water content. In the saline conditions, the water from the irrigation was kept on the substrate (Fig. 3). These changes in water flow could also explain the decreases in leaf water potential (Cpd and Cmd), reflecting the hydraulic signals that may be sent from the root to the shoot in salt-stressed plants. It is evident from leaf water relations (Table 3) that the plants underwent osmotic adjustment as a result of the salt stress imposed. Adaptative decreases in plant osmotic potential to maintain turgor in response to salinity have been widely reported [41,42]. The values of osmotic adjustment (0.2– 0.5 MPa) reported in this work (Table 3) are within those reported for other studies on Mediterranean ornamental plants
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submitted to saline stress [6,9,12]. Turgor maintenance can be mediated either through the accumulation of solutes to lower osmotic potential actively (Table 3), or by changes in wall elasticity [43]. Many species show osmotic adjustment and significant decreases in elasticity in response to drought [44,45], while some authors have demonstrated, in ornamental species, that the elasticity decreased as salinity increased [12,46]. In A. unedo, the osmotic adjustment was accompanied by 31 and 99% increase in the bulk modulus elasticity at 52 and 105 mM, respectively (Table 3), which resulted in turgor loss at the same relative water content in control and in the lower salinity level. In the plants exposed to the highest concentration of NaCl, the point of zero turgor (RWC0) occurred at higher relative water content than in the control. Similar results were obtained by Nabil and Coudret [12] in two Acacia nilotica subspecies in response to salt stress. With higher values of e, tissues became less elastic [47] and the effect is to allow a lower water potential to be reached for a given change in water volume [48]. An increase in e concomitant with osmotic adjustment is an effective means of counteracting the negative effects of salinity on water balance of A. unedo plants. In conclusion, the response of A. unedo to salinity shows an accumulation of salt in its tissues, which reduces stomatal and mesophyll conductance to CO2 and consequently photosynthesis, independently of maintaining the plant water balance. However, the origin of a wild species and its distribution does not necessarily define its degree of tolerance to salt or any other specific stress. Acknowledgment This work was supported by CICYT projects AGL 200505588-C02-1 and AGL 2005-05588-C02-2. References [1] W.J.S. Downton, W.J.R. Grant, S.P. Robinson, Photosynthetic and stomatal responses of spinach leaves to salt stress, Plant Physiol. 77 (1985) 85–88. [2] P. Rodrı´guez, J. Dell’Amico, D. Morales, M.J. Sa´nchez-Blanco, J.J. Alarco´n, Effects of salinity on growth, shoot water relations and root hydraulic conductivity in tomato plants, J. Agric. Sci. 128 (1997) 439– 444. [3] A.H. Markhart, B. Smit, Measurement of root hydraulic conductance, Hortic. Sci. 25 (1990) 282–287. [4] A. Torrecillas, P. Rodrı´guez, M.J. Sa´nchez-Blanco, Comparison of growth, leaf water relations and gas exchange of Cistus albidus and C. monspeliensis plants irrigated with water of different NaCl salinity levels, Sci. Hortic. 97 (2003) 353–368. [5] F. Shihe, T.J. Blake, E. Blumwald, The relative contribution of elastic and osmotic adjustments to turgor maintenance in woody species, Physiol. Plant. 90 (1994) 408–413. [6] M.J. Sa´nchez-Blanco, M.A. Morales, A. Torrecillas, J.J. Alarco´n, Diurnal and seasonal osmotic potential changes in Lotus creticus creticus plants grown under saline stress, Plant Sci. 136 (1998) 1–10. [7] M. Tattini, G. Montagni, L. Andreini, D. Remorini, R. Massai, R. Scott Johnson, Growth, gas exchange and ionic relations of peach rootstocks under root zone salinity stress, in: C.H. Chrisosto (Ed.), Proceedings of the 5th International Peach Symposium, vol. 592, Davis, California, USA, Acta Hortic. (2002) 553–556.
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