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Impact of salt stress on morpho-physiological and biochemical parameters of Solanum lycopersicum cv. Microtom leaves
SAJB-01583; No of Pages 6 South African Journal of Botany xxx (2016) xxx–xxx
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Impact of salt stress on morpho-physiological and biochemical parameters of Solanum lycopersicum cv. Microtom leaves H. Bacha a,⁎, M. Tekaya b, S. Drine a, F. Guasmi a, L. Touil a, H. Enneb a, T. Triki a, F. Cheour c, A. Ferchichi a a b c
Arid Land and Oasis Cropping Laboratory, Institute of Arid Land, Route Eljorf, 4119 Medenine, Tunisia Laboratory of Biochemistry, LR-NAFS/LR12ES05 ‘Nutrition—Functional Food & Vascular Health’, Faculty of Medicine of Monastir, 5019 Monastir, Tunisia Higher Institute of Applied Biology, Medenine, Tunisia
a r t i c l e
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Article history: Received 4 February 2016 Received in revised form 25 July 2016 Accepted 23 August 2016 Available online xxxx Edited by R Baraldi Keywords: Salt stress Solanum lycopersicum cv. Microtom Gas exchange parameters Water use efficiency Chlorophylls Phenols
a b s t r a c t The crops in arid and semi-arid regions are often exposed to adverse environmental factors such as high soil salinity. An experiment was conducted in order to determine the response of tomato (Solanum lycopersicum) cv. Microtom to salinity, a variety that has not been extensively studied yet. The investigated elements were the effects on gas exchange parameters, water use efficiency (WUE) as well as leaf area and the contents of total chlorophylls and phenols. At the stage of six leaves, salt stress was applied for 14 days, and three treatments were tested: T1: 0 mM NaCl (Control, irrigated only with rainwater); T2: 50 mM; T3: 150 mM. Sampling events included the collection of tomato leaf samples at two dates: 7 days and 14 days after salt stress application. The obtained results demonstrated that the period of salt stress and salinity treatments had significant effects on the studied parameters, being much more accentuated after two weeks of 150 mM salt treatment. Microtom showed different mechanisms of adaptation to saline stress. Under this stress, the adaptations are mainly morphological (by reducing leaf area), physiological (reduction in net CO2 assimilation rate, stomatal conductance and transpiration, and improvement of WUE) and biochemical responses (decrease of chlorophyll content). Thus, phenol accumulation was stimulated in the leaves of Microtom as a common defensive mechanism. These aspects allowed classifying Microtom as a tolerant cultivar to salinity. © 2016 SAAB. Published by Elsevier B.V. All rights reserved.
1. Introduction Salinity is a major abiotic factor limiting plant growth and fruit yield (Parada and Das, 2006). Currently, about 77 million hectares (5%) are affected by excessive salt concentrations (Sheng et al., 2008). This figure continues to increase from one year to another due to the poor quality of irrigation water (Villa-Castorena et al., 2003). Tunisia is among the countries threatened by a water shortage problem (especially when it comes to water quality). Faced with this problem, the country is obliged to come up with a solution by moving towards determining the actual water needs of different cultures and to the possibility of using saline water for irrigation. However, it was proven that about 30% of irrigated areas are affected by salts in different degrees (Hachicha, 2007). Soils affected by salts cover about 1.5 million hectares, which stands for around 10% of the total area of the country. Several studies have reported that salinity induces morphological, physiological, and biochemical changes in many crops (Ashraf and Foolad, 2007). Tolerance to various stresses differ depending to the species, varieties, and even ecotypes (Ullah et al., 2008). In response to stress, powerful plant antioxidant systems are activated, both ⁎ Corresponding author. E-mail address: [email protected] (H. Bacha).
enzymatic (e.g., superoxide dismutase, catalase, glutathione reductase, several peroxidases) and non-enzymatic (vitamins C and E, carotenoids, flavonoids, and other phenolic compounds, etc.) (Apel and Hirt, 2004). These phenomena have been observed in agricultural and horticultural crops, including tomato (Juan et al., 2005). The most general response to abiotic stresses consists of the accumulation of osmolytes which is a well-known phenomenon observed in all plants (Munns and Tester, 2008). Among the most common solutes which are accumulated in plants in response to abiotic stress, we identified glycine betaine and proline; and many works have been dedicated to these compounds, particularly in tomato plants (Kahlaoui et al., 2013). It has been observed in a previous work (Bacha et al., 2015) that S. lycopersicum plants of cv. Microtom, treated with different concentrations of salinity, accumulated the osmoprotectants glycine betaine and proline in large amounts as protective mechanisms to acclimate to the abiotic stress. Apart from that, there are many other compounds that were found accumulated in response to abiotic stress; among these, we can mention phenolic compounds which play a major role as powerful antioxidants (Petridis et al., 2012). Tomatoes (Solanum lycopersicum L.) are one of the most important vegetables in the human diet that can be eaten either fresh or processed. Its cultivation plays an important role in the Tunisian agricultural economy. Several studies have investigated the effects of salinity on tomato
http://dx.doi.org/10.1016/j.sajb.2016.08.018 0254-6299/© 2016 SAAB. Published by Elsevier B.V. All rights reserved.
Please cite this article as: Bacha, H., et al., Impact of salt stress on morpho-physiological and biochemical parameters of Solanum lycopersicum cv. Microtom leaves, South African Journal of Botany (2016), http://dx.doi.org/10.1016/j.sajb.2016.08.018
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crops (Sholi, 2012). As the response to salt stress is genotypedependent (Maggio et al., 2004), there has been a growing interest in studying the response of new cultivars to salt stress in order to evaluate their degree of tolerance to salinity and select the most resistant ones. The tomato cultivar Microtom was produced for ornamental purposes by crossing Florida Basket and Ohio 4013-3 cultivars, and is characterized by a dwarf phenotype with small red ripened fruits (Scott and Harbaugh, 1989). Because it is characterized by rapid growth and easy transformation, this cultivar has been proposed as a model system that allows studying the regulation of berry fruit development (Eyal and Levy, 2002). Only few studies have been carried out in investigating the response of this cultivar to salt stress (Knight et al., 1992). This work was conducted in the same context with the ultimate purpose of evaluating the response of tomato cultivar Microtom, a variety that has not been extensively studied yet, and its adaptation to salt stress. The factors that will be taken under study consist of modifications to key physiological parameters such as gas exchange parameters, water use efficiency, leaf area and chlorophyll content, as well as stress metabolites particularly total phenols. 2. Material and methods 2.1. Plant material and growth conditions
H2O m− 2 S−1) were measured at air temperature of 25 °C, with a photosynthetic photon flux density (PPFD) of 1200 μmol m−2 s−1 and a relative humidity of 60%. Moreover, instantaneous water use efficiency (WUEinstantaneous) (μmol CO2 mmol−1 H2O) and intrinsic water use efficiency (WUEintrinsic) (μmol CO2 mol− 1 H2O) were calculated from the gas exchange measurements as described by Polley (2002) and Thameur et al. (2012). 2.3.2. Leaf area Leaf area (cm2) was calculated from digitized images taken with a standard flat-bed scanner, using Mesurim software Pro 6 Version 3.2. (Madre, 1998). 2.3.3. Total chlorophyll content Total chlorophyll content was determined according to the method described by Arnon (1949). Ten leaf disks of 38.46 mm2 were taken and transferred to a 15 ml falcon tube in which 5 ml of acetone (80%) were added for chlorophyll extraction. The extracts were kept in the dark overnight at 4 °C. After centrifugation, absorbance was measured at 645 nm and 663 nm, and chlorophyll concentration was calculated using the formula given by MacKinney (1941), and then expressed in mg/g F.W.:
The plant material used in this study consists of a variety of tomato plants (S. lycopersicum L. cv. Microtom) derived from Peru. The experiment was conducted in a greenhouse located in the “Institute of Arid Regions”, 20 km southeast of Medenine (a major town in southeastern Tunisia, latitude 33°35′N; longitude 10°48′3″E; altitude 105 m). The climate of this region is Mediterranean, with hot, dry summers and mild winters. The greenhouse is equipped with a ventilation system which maintained the temperature at about 25 °C/18 °C (day/night) and the relative humidity at 70 °C/80% (day/night). The photosynthetic photon flux density (PPFD) was about 500 mmol m− 2 s− 1, and the radiation closely resembled ambient conditions for light intensity. Seeds of tomato plants were germinated at 28 °C in vermiculite; 10 days later, they were transferred to single-plant pots (30 cm long by 10 cm diameter). The pots had a volume of 5 L each and were filled with a mixture of sandy loam soil (90%) and manure (10%), having a pH of 7.5 and an electrical conductivity (E.C.) of 3.9 mΩ cm−1. A nutrient solution containing the following proportions of minerals (for 1 L water) was introduced: 250 g NH4NO3, + 400 g KNO3 + 100 ml H2PO4 (50%). Rainwater was used for irrigation, measuring a pH of 7.71, an E.C. of 845 μΩ cm−1 at T = 25 °C–26 °C, and a salinity equal to 0.5 g/L.
Total chlorophylls ðmg=LÞ ¼ 20:2 A645 þ 8:02 A663
2.2. Experimental procedure
3. Results and discussion
At the stage of six leaves (stage 16 on the Zadoks scale; Zadoks et al., 1974), salt stress was applied for 14 days; and a randomized block design with three blocks, three treatments and three replications for each treatment was used:
3.1. Gas exchange performance
• T1: 0 mM NaCl (Control, irrigated only with rainwater) • T2: 50 mM NaCl • T3: 150 mM NaCl Sampling events included the collection of tomato leaf samples at two dates: 7 days and 14 days after salt stress application. 2.3. Measurements 2.3.1. Gas exchange parameters and water use efficiency Flag leaf gas exchange parameters were measured using the photosynthetic apparatus LCi IRGA (LCi, IRGA; ADC Bioscientific Ltd.). Net photosynthesis (A, expressed in μmol CO2 m− 2 s−1), stomatal conductance (gs, mmol H2O m− 2.S− 1), and transpiration (Tr, mmol
where A is the absorbance of the extract at the respective wavelength. 2.3.4. Total phenol content Total phenols were extracted in a solution of methanol (90%) and were quantified colorimetrically according to the method described by Velioglu et al. (1998). The Folin–Ciocalteau reagent was added to a suitable aliquot of the leaf extracts, and the absorption of the solution at 765 nm was measured. Values are given as mg of gallic acid per gram of D.W. 2.4. Statistical analysis All statistical values were calculated using the Statistical Package for Social Sciences (SPSS) program, release 11.0 for Windows (SPSS, Chicago, IL, USA). Values are given as the mean ± standard deviations of three measurements. Duncan's multiple range tests were used to determine significant differences among data. The statistical significance level was fixed at p b 0.05. Pearson's correlation coefficients were used when calculating correlations between different studied parameters.
Photosynthesis is the most fundamental and intricate physiological process that all green plants undergo as it considerably affects the plant growth. Since the mechanism of photosynthesis involves various components, including photosynthetic pigments and photosystems, the electron transport system, and CO2 reduction pathways, any damage at any level caused by stress may reduce the overall photosynthetic capacity of a green plant (Ashraf and Harris, 2013). Photosynthesis measured in stressed and unstressed tomatoes was evaluated by net CO2 assimilation rate (A), stomatal conductance (gs), and transpiration (Tr) (Fig. 1.). As shown in Fig. 1B, it was observed that gs was noticeably decreased after a 1-week treatment with 50 and 150 mM NaCl, from 0.14 ± 0.012 mol H2O m−2 s−1 to 0.1 ± 0.012 mol H2O m−2 s−1 and to 0.09 ± 0.01 mol H2O m−2 s−1, respectively, whereas A decreased by about 17.86% and 29.57% respectively (Fig. 1A). After two weeks of salt stress application, the gas exchange was drastically reduced in both salt treatments; gs decreased by about 56.25% in plants subjected
Please cite this article as: Bacha, H., et al., Impact of salt stress on morpho-physiological and biochemical parameters of Solanum lycopersicum cv. Microtom leaves, South African Journal of Botany (2016), http://dx.doi.org/10.1016/j.sajb.2016.08.018
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Fig. 1. Measurements of net CO2 assimilation rate (A, A), stomatal conductance (gs, B) and transpiration (Tr, C) at 2 dates of leaf sampling (after 1 week and after 2 weeks of stress application). T1: control, 0 mM NaCl; T2: 50 mM NaCl; and T3: 150 mM NaCl. The results are expressed as means ± S.D. (n = 3). Different letters indicate significantly different values at p ≤ 0.05 according to Duncan test.
to T2 treatment while the reduction was by about 81.25% in plants subjected to T3 treatment. Statistical analysis showed the same trend in A levels, which strongly depressed under T2 and T3 treatments, respectively, by about 31.59% and 71.42%. As a consequence of the reduction of gs, transpiration decreased as well soon after salinity stress was imposed and continued to drop, reaching 34.33% and 82.47% of reduction under T2 and T3 treatments, respectively (Fig. 1C). These attained results are in consonance with those of Lycoskoufis et al. (2005), Niu et al. (2010), and Cheng-Jin et al. (2011) who found a significant decrease of gas exchange parameters under salt stress. In fact, the stomatal quick response of stressed plants can stand for a salinity tolerance mechanism (Jones, 1974). Salinity may limit net photosynthesis and stomatal conductance, either because of a CO2 supply limitation resulting from the partial closure of stomata (stomatal function), or by altering the biochemical mechanism of CO2 fixation (not a stomatal function), or by both procedures (Chaves et al., 2003). It was reported that cellular membranes are highly sensitive to stresses (Tayefi-Nasrabadi et al., 2011). Consequently, the accumulation of high concentrations of Na+ and Cl− in the chloroplasts under salinity stress is proven to damage thylakoid membranes (Omoto et al., 2010).
3.2. Changes in water use efficiency in response to salt stress The WUEinstantaneous (A/Tr) and WUEintrinsic (A/gs) were measured for leaf samples and the results are represented in Fig. 2. The WUEinstantaneous was affected by salinity only after a 2-week treatment (Fig. 2A). In fact, it increased significantly with the growing concentration of salt, by about 32.80% and 54.66%, in plants subjected to T2 and T3 treatments. This increase suggests that transpiration rates may have been more affected than CO2 assimilation rates. The WUEintrinsic (A/gs) showed the same trend as WUEinstantaneous (Fig. 2B); but this increase of WUE intrinsic in salt-stressed leaves occurred soon after salinity stress was imposed (1 week treatment). This indicates a greater reduction of gs than of A, and thus, increasing water use efficiency. Similar results were obtained by Thameur and Lachiheb (2012) in their work on saltstressed barley. In fact, in arid regions, WUE is of a vital importance in terms of water saving. According to Araus et al. (2002), high WUE is considered as a strategy to improve the performance of crops in arid conditions.
Please cite this article as: Bacha, H., et al., Impact of salt stress on morpho-physiological and biochemical parameters of Solanum lycopersicum cv. Microtom leaves, South African Journal of Botany (2016), http://dx.doi.org/10.1016/j.sajb.2016.08.018
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Fig. 2. Modification of instantaneous water use efficiency (WUEinstantaneous, A) and intrinsic water use efficiency (WUEintrinsic, B) at 2 dates of leaf sampling (after 1 week and after 2 weeks of stress application). T1: control, 0 mM NaCl; T2: 50 mM NaCl; and T3: 150 mM NaCl. The results are expressed as means ± S.D. (n = 3). Different letters indicate significantly different values at p ≤ 0.05 according to Duncan test.
3.3. Modifications of leaf area The leaf area represents a measure of plant growth, which can be affected by different stresses, including salt stress. Leaf area (LA) was measured for both control plants and plants subjected to salt stress
(Fig. 3.). It has been shown that this parameter increased over time but LA was higher when provided with favorable conditions (control plants) than the salt-stressed ones. For control plants, it increased from 54.74 ± 1.71 cm2 to 87.37 ± 6.38 cm2 between the first harvest and the second. Regarding the effect of salinity treatments, a little
Fig. 3. Leaf area (LA) of tomato cv. Microtom plants measured at 2 dates of leaf sampling (after 1 week and after 2 weeks of stress application). T1: control, 0 mM NaCl; T2: 50 mM NaCl; and T3: 150 mM NaCl. The results are expressed as means ± S.D. (n = 3). Different letters indicate significantly different values at p ≤ 0.05 according to Duncan test.
Please cite this article as: Bacha, H., et al., Impact of salt stress on morpho-physiological and biochemical parameters of Solanum lycopersicum cv. Microtom leaves, South African Journal of Botany (2016), http://dx.doi.org/10.1016/j.sajb.2016.08.018
H. Bacha et al. / South African Journal of Botany xxx (2016) xxx–xxx
decrease in LA was noticed as a consequence of the elevation of salt concentration, and this decrease was accentuated in the second harvest. In fact, the mean value of LA exhibited a reduction in the order of 15.36% and 41.3%, respectively, for the two treatments of 50 and 150 mM NaCl. The reduction in leaf area was due to the decline in leaf gas exchange during salt stress. We found good and significant correlations between LA and A (R = 0.892; p b 0.001 in the second sampling) and between LA and gs (R = 0.781; p b 0.001 in the first sampling and R = 0.809; p b 0.001 in the second sampling). In fact, the reduction of leaf area stands for a morphological adaptive strategy for salt tolerance. The present result is in agreement with the findings of Thameur and Lachiheb (2012) in barley, Vendeland et al. (1982) in soybean, Belaygue et al. (1996) in white clover, Clough and Milthorpe (1975) in tobacco, and NeSmith and Ritchie (1992) in maize, where a decrease in LA of stressed plants was reported. 3.4. Effect of salt stress on foliar chlorophyll content The principal pigment found in the majority of oxygenic photosynthetic organisms is known to be chlorophyll. Chlorophyll content is one of the main factors that reflect the photosynthetic rate (Mao et al., 2007). Some authors suggested that variation in pigment content can provide valuable insight into the physiological performance of leaves and indicates their photosynthetic capacity as well as the presence of stress or diseases (Boquera and Morales, 2010). In the present study, total chlorophyll content was measured for both stressed and unstressed tomato plants cv. Microtom, and the results were illustrated in Table 1. Statistics analysis showed that after a 1-week treatment, chlorophyll concentration exhibited a little reduction under 50 mM NaCl (from 1.13 ± 0.02 mg/g to 1.07 ± 0.03 mg/g) and 150 mM NaCl applications (from 1.13 ± 0.02 mg/g to 1.04 ± 0.03 mg/g). However, after a 2-week salt treatment, chlorophyll content decreased drastically with the increasing concentration of salt to reach 44.82% of reduction under 150 mM of salt concentration. It is probable that the changes in the chlorophyll concentration in tomato plants subjected to high concentration of NaCl, mainly under the 150-mM treatment, is a response to compensating for the loss of leaf area owing to a smaller leaf size. We also found a good and significant correlation (R = 0.896; p b 0.05) between leaf area and total chlorophyll content after a 2-week salt treatment. The chlorophyll content of crop plants is positively correlated with their photosynthetic activity (Gummuluru et al., 1989) and a reduction of chlorophyll level contributes to the inhibition of photosynthesis observed under abiotic stress conditions. This is also confirmed in our study since a good correlation was found between chlorophyll content and net photosynthetic rate (A) (R = 0.879; p b 0.001) at a 2-week salt-stress treatment. Our results are in agreement with those of Noreen et al. (2009) and Kaya et al. (2009) who observed a loss of chlorophyll content in plants subjected to salt stress. The salt-induced alterations in leaf chlorophyll content could be due to impaired biosynthesis or accelerated pigment degradation. The effect is ascribed to an increased level of the toxic cation, Na+ (Yang et al., 2011). Some studies suggest that chlorophyll accumulation under salt stress could be used as a biochemical marker for salt tolerance in different crops (Akram and Ashraf, 2011). However,
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in some other studies, this accumulation is not always associated with salt tolerance. For example, Juan et al. (2005), in their study on saltstressed tomato cultivars which differ in salinity tolerance, observed a weak correlation between photosynthetic pigments and leaf Na+. Thus, chlorophylls cannot be considered as good indicators for salt tolerance in tomatoes. 3.5. Response of phenols to salt-stress treatments In response to abiotic stress, plants have developed a wide variety of highly sophisticated and efficient mechanisms to sense, respond, and adapt to a wide range of environmental changes. A common defensive mechanism activated in plants exposed to stressing conditions is the production and accumulation of phenolic compounds. The antioxidant property of plant phenolic compounds, the metabolic pathways for their biosynthesis, and the involved enzymes are well documented in most important plant species (Balasundram et al., 2006). Some enzymes involved in phenolic metabolism such as polyphenol oxidase (PPO) and peroxidase (POD) generally respond actively to the presence of stress in the plant (Lotfi et al., 2010). In the present study, and after a 1-week treatment, total phenol concentration increased significantly in stressed Microtom leaves, only in response to a dose of 150 mM NaCl, by about 58.28% compared to control leaves (Table 1). However, when extending the period of salt stress to 2 weeks, we observed a remarkable increase of phenols in stressed plants of both tested treatments, by an increase of 70.45% for T2 leaves and 82.82% for T3 leaves. Our results are in consonance with Al Hassan et al. (2015) who observed a significant increase of phenolics in stressed tomato leaves under moderate and high salt concentrations in cherry tomato (S. lycopersicum L. cv. cerasiforme). The increase in antioxidant phenolic compound levels in leaves can be considered as part of the response induced to cope with oxidative stress induced by salinity. Thus, salt-stressed plants might represent potential sources of polyphenols, by increasing polyphenol concentration in the tissues, which is an issue that is directly tied with human health since these compounds are known to be bio-active compounds (Jemai et al., 2008). In fact, optimal polyphenol content would be obtained using stress-tolerant species (De Abreu and Mazzafera, 2005). 4. Conclusions The mechanism of adaptation to saline stress results in morphological, physiological, and biochemical responses. In this study, tomato plants of cv. Microtom adapted to salt stress by reducing leaf area, stomatal conductance, and minimizing water loss by transpiration. These led to greater reduction in photosynthetic rate, especially after 2 weeks of 150 mM salt treatment, and a decrease of the content of the major photosynthetic pigment (chlorophyll). Besides, Microtom has the ability to increase the efficiency of water use, which is of vital importance in terms of water saving and which represents a strategy to improve the performance of crops in arid conditions. In consequence to the enhanced production of ROS (reactive oxygen species) generated by salt stress, phenol accumulation was stimulated in the leaves of Microtom as a common defensive mechanism. These facts could be
Table 1 Variations in the foliar contents of chlorophylls and total phenols following increasing concentration of salinity and period of salt treatment. Parameters
Total chlorophylls (mg g
Sampling date (after stress application) −1
Total phenols (mg eq. GA g
FW)
−1
DW)
After 1 week After 2 weeks After 1 week After 2 weeks
Concentrations of salt treatments T1 (0 mM)
T2 (50 mM)
T3 (150 mM)
1.13 ± 0.02 a 1.16 ± 0.03 a 0.78 ± 0.12 b 0.80 ± 0.1 c
1.07 ± 0.03 a 0.92 ± 0.04 b 0.81 ± 0.11 b 2.64 ± 0.20 b
1.04 ± 0.03 b 0.64 ± 0.02 c 1.87 ± 0.21 a 4.54 ± 0.19 a
Results are expressed as means ± SD (n = 3). a, b, c, d: Values in the same row with different letters showed statistically significant differences (p b 0.05) according to Duncan test.
Please cite this article as: Bacha, H., et al., Impact of salt stress on morpho-physiological and biochemical parameters of Solanum lycopersicum cv. Microtom leaves, South African Journal of Botany (2016), http://dx.doi.org/10.1016/j.sajb.2016.08.018
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one of the strategies used by these plants to tolerate the severe conditions imposed by soils with a high salt concentration in arid lands. Though some other yet unidentified factors could also be involved in the survival of these plants under continuous salt stress.
Acknowledgments This study was supported by the Ministry of Higher Education and Scientific Research in Tunisia. We express our sincere thanks to the members of the laboratory “Aridoculture et Cultures Oasiennes” in the institute of Arid Regions in Medenine-Tunisia, and also to the members of “Department of Plant Nutrition, CEBAS–CSIC, Campus de Espinardo, Murcia Spain” for their support to this research. The authors declare that they have no conflict of interest. References Akram, N.A., Ashraf, M., 2011. Improvement in growth, chlorophyll pigments and photosynthetic performance in salt-stressed plants of sunflower (Helianthus annuus L.) by foliar application of 5-aminolevulinic acid. Agrochimica 55, 94–104. Al Hassan, M., Martinez Fuertes, M., Ramos Sanchez, F.J., Vicente, O., Boscaiu, M., 2015. Effects of salt and water stress on plant growth and on accumulation of osmolytes and antioxidant compounds in cherry tomato. Notulae Botanicae Horti Agrobotanici Cluj-Napoca 43, 1–11. Apel, K., Hirt, H., 2004. Reactive oxygen species: metabolism, oxidative stress, and signal transduction. Annual Review of Plant Biology 55, 373–399. Araus, J.L., Slafer, G.A., Reynolds, M.P., Royo, C., 2002. Plant breeding and water relations in C3 cereals: what should we breed for? Annals of Botany 89, 925–940. Arnon, D.I., 1949. Copper enzymes in isolated chloroplasts. Polyphenoloxidase in Beta vulgaris. Plant Physiology 24, 1–15. Ashraf, M., Foolad, M.R., 2007. Roles of glycinebetaine and proline in improving plant abiotic stress tolerance. Environmental and Experimental Botany 59, 206–216. Ashraf, M., Harris, P.J.C., 2013. Photosynthesis under stressful environments: an overview. Photosynthetica 51, 163–190. Bacha, H., Mansour, E., Guasmi, F., Triki, T., Ferchichi, A., 2015. Proline, glycine bétaïne et composition minérale des plantes de Solanum lycopersicum L. (var. Microtom) sous stress salin. Journal of New Sciences 2286–5314. Balasundram, N., Sundram, K., Samman, S., 2006. Phenolic compounds in plants and agriindustrial by-products: antioxidant activity, occurrence, and potential uses. Food Chemistry 99, 191–203. Belaygue, C., Wery, J., Cowan, A.A., Tardieu, F., 1996. Contribution of leaf expansion, rate of leaf appearance, and stolon branching to growth of plant leaf area under water deficit in white clover. Crop Science 36, 1240–1246. Boquera, M.L.E., Morales, P.L.V.C., 2010. Leaf chlorophyll content estimation in the monarch butterfly biosphere reserve. Revista Fitotecnia Mexicana 33, 175–181. Chaves, M.M., Maroco, J.P., Pereira, J.S., 2003. Understanding plant responses to drought—from genes to the whole plant. Functional Plant Biology 30, 239–264. Cheng-Jin, W., Sun, Y.L., Cho, D.H., 2011. Changes in photosynthetic rate, water potential, and proline content in kenaf seedlings under salt stress. Canadian Journal of Plant Science 311–319. Clough, B.F., Milthorpe, F.L., 1975. Effect of water deficit on leaf development in tobacco. Australian Journal of Plant Physiology 2, 291–300. De Abreu, I.N., Mazzafera, P., 2005. Effect of water and temperature stress on the content of active constituents of Hypericum brasilienne Choisy. Plant Physiology and Biochemistry 43, 241–248. Eyal, E., Levy, A.A., 2002. Tomato mutants as tools for functional genomics. Current Opinion in Plant Biology 5, 112–117. Gummuluru, S., Hobbs, S.L.A., Jana, S., 1989. Genotypic variability in physiological characters and its relationship to drought tolerance in durum wheat. Canadian Journal of Plant Science 69, 703–711. Hachicha, M., 2007. Les sols sales et leur mise en valeur en Tunisie. Secheresse 18, 45–50. Jemai, H., Bouaziz, M., Fki, I., El Feki, A., Sayadi, S., 2008. Hypolipidimic and antioxidant activities of oleuropein and its hydrolysis derivative-rich extracts from Chemlali olive leaves. Chemico-Biological Interactions 176, 88–98. Jones, H.G., 1974. Assessment of stomatal control of plant water status. The New Phytologist 73, 851–859. Juan, M., Rivero, R.M., Romero, L., Ruiz, J.M., 2005. Evaluation of some nutritional and biochemical indicators in selecting salt-resistant tomato cultivars. Environmental and Experimental Botany 54, 193–201.
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Please cite this article as: Bacha, H., et al., Impact of salt stress on morpho-physiological and biochemical parameters of Solanum lycopersicum cv. Microtom leaves, South African Journal of Botany (2016), http://dx.doi.org/10.1016/j.sajb.2016.08.018
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Impact of salt stress on morpho-physiological and biochemical parameters of Solanum lycopersicum cv. Microtom leaves H. Bacha a,⁎, M. Tekaya b, S. Drine a, F. Guasmi a, L. Touil a, H. Enneb a, T. Triki a, F. Cheour c, A. Ferchichi a a b c
Arid Land and Oasis Cropping Laboratory, Institute of Arid Land, Route Eljorf, 4119 Medenine, Tunisia Laboratory of Biochemistry, LR-NAFS/LR12ES05 ‘Nutrition—Functional Food & Vascular Health’, Faculty of Medicine of Monastir, 5019 Monastir, Tunisia Higher Institute of Applied Biology, Medenine, Tunisia
a r t i c l e
i n f o
Article history: Received 4 February 2016 Received in revised form 25 July 2016 Accepted 23 August 2016 Available online xxxx Edited by R Baraldi Keywords: Salt stress Solanum lycopersicum cv. Microtom Gas exchange parameters Water use efficiency Chlorophylls Phenols
a b s t r a c t The crops in arid and semi-arid regions are often exposed to adverse environmental factors such as high soil salinity. An experiment was conducted in order to determine the response of tomato (Solanum lycopersicum) cv. Microtom to salinity, a variety that has not been extensively studied yet. The investigated elements were the effects on gas exchange parameters, water use efficiency (WUE) as well as leaf area and the contents of total chlorophylls and phenols. At the stage of six leaves, salt stress was applied for 14 days, and three treatments were tested: T1: 0 mM NaCl (Control, irrigated only with rainwater); T2: 50 mM; T3: 150 mM. Sampling events included the collection of tomato leaf samples at two dates: 7 days and 14 days after salt stress application. The obtained results demonstrated that the period of salt stress and salinity treatments had significant effects on the studied parameters, being much more accentuated after two weeks of 150 mM salt treatment. Microtom showed different mechanisms of adaptation to saline stress. Under this stress, the adaptations are mainly morphological (by reducing leaf area), physiological (reduction in net CO2 assimilation rate, stomatal conductance and transpiration, and improvement of WUE) and biochemical responses (decrease of chlorophyll content). Thus, phenol accumulation was stimulated in the leaves of Microtom as a common defensive mechanism. These aspects allowed classifying Microtom as a tolerant cultivar to salinity. © 2016 SAAB. Published by Elsevier B.V. All rights reserved.
1. Introduction Salinity is a major abiotic factor limiting plant growth and fruit yield (Parada and Das, 2006). Currently, about 77 million hectares (5%) are affected by excessive salt concentrations (Sheng et al., 2008). This figure continues to increase from one year to another due to the poor quality of irrigation water (Villa-Castorena et al., 2003). Tunisia is among the countries threatened by a water shortage problem (especially when it comes to water quality). Faced with this problem, the country is obliged to come up with a solution by moving towards determining the actual water needs of different cultures and to the possibility of using saline water for irrigation. However, it was proven that about 30% of irrigated areas are affected by salts in different degrees (Hachicha, 2007). Soils affected by salts cover about 1.5 million hectares, which stands for around 10% of the total area of the country. Several studies have reported that salinity induces morphological, physiological, and biochemical changes in many crops (Ashraf and Foolad, 2007). Tolerance to various stresses differ depending to the species, varieties, and even ecotypes (Ullah et al., 2008). In response to stress, powerful plant antioxidant systems are activated, both ⁎ Corresponding author. E-mail address: [email protected] (H. Bacha).
enzymatic (e.g., superoxide dismutase, catalase, glutathione reductase, several peroxidases) and non-enzymatic (vitamins C and E, carotenoids, flavonoids, and other phenolic compounds, etc.) (Apel and Hirt, 2004). These phenomena have been observed in agricultural and horticultural crops, including tomato (Juan et al., 2005). The most general response to abiotic stresses consists of the accumulation of osmolytes which is a well-known phenomenon observed in all plants (Munns and Tester, 2008). Among the most common solutes which are accumulated in plants in response to abiotic stress, we identified glycine betaine and proline; and many works have been dedicated to these compounds, particularly in tomato plants (Kahlaoui et al., 2013). It has been observed in a previous work (Bacha et al., 2015) that S. lycopersicum plants of cv. Microtom, treated with different concentrations of salinity, accumulated the osmoprotectants glycine betaine and proline in large amounts as protective mechanisms to acclimate to the abiotic stress. Apart from that, there are many other compounds that were found accumulated in response to abiotic stress; among these, we can mention phenolic compounds which play a major role as powerful antioxidants (Petridis et al., 2012). Tomatoes (Solanum lycopersicum L.) are one of the most important vegetables in the human diet that can be eaten either fresh or processed. Its cultivation plays an important role in the Tunisian agricultural economy. Several studies have investigated the effects of salinity on tomato
http://dx.doi.org/10.1016/j.sajb.2016.08.018 0254-6299/© 2016 SAAB. Published by Elsevier B.V. All rights reserved.
Please cite this article as: Bacha, H., et al., Impact of salt stress on morpho-physiological and biochemical parameters of Solanum lycopersicum cv. Microtom leaves, South African Journal of Botany (2016), http://dx.doi.org/10.1016/j.sajb.2016.08.018
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crops (Sholi, 2012). As the response to salt stress is genotypedependent (Maggio et al., 2004), there has been a growing interest in studying the response of new cultivars to salt stress in order to evaluate their degree of tolerance to salinity and select the most resistant ones. The tomato cultivar Microtom was produced for ornamental purposes by crossing Florida Basket and Ohio 4013-3 cultivars, and is characterized by a dwarf phenotype with small red ripened fruits (Scott and Harbaugh, 1989). Because it is characterized by rapid growth and easy transformation, this cultivar has been proposed as a model system that allows studying the regulation of berry fruit development (Eyal and Levy, 2002). Only few studies have been carried out in investigating the response of this cultivar to salt stress (Knight et al., 1992). This work was conducted in the same context with the ultimate purpose of evaluating the response of tomato cultivar Microtom, a variety that has not been extensively studied yet, and its adaptation to salt stress. The factors that will be taken under study consist of modifications to key physiological parameters such as gas exchange parameters, water use efficiency, leaf area and chlorophyll content, as well as stress metabolites particularly total phenols. 2. Material and methods 2.1. Plant material and growth conditions
H2O m− 2 S−1) were measured at air temperature of 25 °C, with a photosynthetic photon flux density (PPFD) of 1200 μmol m−2 s−1 and a relative humidity of 60%. Moreover, instantaneous water use efficiency (WUEinstantaneous) (μmol CO2 mmol−1 H2O) and intrinsic water use efficiency (WUEintrinsic) (μmol CO2 mol− 1 H2O) were calculated from the gas exchange measurements as described by Polley (2002) and Thameur et al. (2012). 2.3.2. Leaf area Leaf area (cm2) was calculated from digitized images taken with a standard flat-bed scanner, using Mesurim software Pro 6 Version 3.2. (Madre, 1998). 2.3.3. Total chlorophyll content Total chlorophyll content was determined according to the method described by Arnon (1949). Ten leaf disks of 38.46 mm2 were taken and transferred to a 15 ml falcon tube in which 5 ml of acetone (80%) were added for chlorophyll extraction. The extracts were kept in the dark overnight at 4 °C. After centrifugation, absorbance was measured at 645 nm and 663 nm, and chlorophyll concentration was calculated using the formula given by MacKinney (1941), and then expressed in mg/g F.W.:
The plant material used in this study consists of a variety of tomato plants (S. lycopersicum L. cv. Microtom) derived from Peru. The experiment was conducted in a greenhouse located in the “Institute of Arid Regions”, 20 km southeast of Medenine (a major town in southeastern Tunisia, latitude 33°35′N; longitude 10°48′3″E; altitude 105 m). The climate of this region is Mediterranean, with hot, dry summers and mild winters. The greenhouse is equipped with a ventilation system which maintained the temperature at about 25 °C/18 °C (day/night) and the relative humidity at 70 °C/80% (day/night). The photosynthetic photon flux density (PPFD) was about 500 mmol m− 2 s− 1, and the radiation closely resembled ambient conditions for light intensity. Seeds of tomato plants were germinated at 28 °C in vermiculite; 10 days later, they were transferred to single-plant pots (30 cm long by 10 cm diameter). The pots had a volume of 5 L each and were filled with a mixture of sandy loam soil (90%) and manure (10%), having a pH of 7.5 and an electrical conductivity (E.C.) of 3.9 mΩ cm−1. A nutrient solution containing the following proportions of minerals (for 1 L water) was introduced: 250 g NH4NO3, + 400 g KNO3 + 100 ml H2PO4 (50%). Rainwater was used for irrigation, measuring a pH of 7.71, an E.C. of 845 μΩ cm−1 at T = 25 °C–26 °C, and a salinity equal to 0.5 g/L.
Total chlorophylls ðmg=LÞ ¼ 20:2 A645 þ 8:02 A663
2.2. Experimental procedure
3. Results and discussion
At the stage of six leaves (stage 16 on the Zadoks scale; Zadoks et al., 1974), salt stress was applied for 14 days; and a randomized block design with three blocks, three treatments and three replications for each treatment was used:
3.1. Gas exchange performance
• T1: 0 mM NaCl (Control, irrigated only with rainwater) • T2: 50 mM NaCl • T3: 150 mM NaCl Sampling events included the collection of tomato leaf samples at two dates: 7 days and 14 days after salt stress application. 2.3. Measurements 2.3.1. Gas exchange parameters and water use efficiency Flag leaf gas exchange parameters were measured using the photosynthetic apparatus LCi IRGA (LCi, IRGA; ADC Bioscientific Ltd.). Net photosynthesis (A, expressed in μmol CO2 m− 2 s−1), stomatal conductance (gs, mmol H2O m− 2.S− 1), and transpiration (Tr, mmol
where A is the absorbance of the extract at the respective wavelength. 2.3.4. Total phenol content Total phenols were extracted in a solution of methanol (90%) and were quantified colorimetrically according to the method described by Velioglu et al. (1998). The Folin–Ciocalteau reagent was added to a suitable aliquot of the leaf extracts, and the absorption of the solution at 765 nm was measured. Values are given as mg of gallic acid per gram of D.W. 2.4. Statistical analysis All statistical values were calculated using the Statistical Package for Social Sciences (SPSS) program, release 11.0 for Windows (SPSS, Chicago, IL, USA). Values are given as the mean ± standard deviations of three measurements. Duncan's multiple range tests were used to determine significant differences among data. The statistical significance level was fixed at p b 0.05. Pearson's correlation coefficients were used when calculating correlations between different studied parameters.
Photosynthesis is the most fundamental and intricate physiological process that all green plants undergo as it considerably affects the plant growth. Since the mechanism of photosynthesis involves various components, including photosynthetic pigments and photosystems, the electron transport system, and CO2 reduction pathways, any damage at any level caused by stress may reduce the overall photosynthetic capacity of a green plant (Ashraf and Harris, 2013). Photosynthesis measured in stressed and unstressed tomatoes was evaluated by net CO2 assimilation rate (A), stomatal conductance (gs), and transpiration (Tr) (Fig. 1.). As shown in Fig. 1B, it was observed that gs was noticeably decreased after a 1-week treatment with 50 and 150 mM NaCl, from 0.14 ± 0.012 mol H2O m−2 s−1 to 0.1 ± 0.012 mol H2O m−2 s−1 and to 0.09 ± 0.01 mol H2O m−2 s−1, respectively, whereas A decreased by about 17.86% and 29.57% respectively (Fig. 1A). After two weeks of salt stress application, the gas exchange was drastically reduced in both salt treatments; gs decreased by about 56.25% in plants subjected
Please cite this article as: Bacha, H., et al., Impact of salt stress on morpho-physiological and biochemical parameters of Solanum lycopersicum cv. Microtom leaves, South African Journal of Botany (2016), http://dx.doi.org/10.1016/j.sajb.2016.08.018
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Fig. 1. Measurements of net CO2 assimilation rate (A, A), stomatal conductance (gs, B) and transpiration (Tr, C) at 2 dates of leaf sampling (after 1 week and after 2 weeks of stress application). T1: control, 0 mM NaCl; T2: 50 mM NaCl; and T3: 150 mM NaCl. The results are expressed as means ± S.D. (n = 3). Different letters indicate significantly different values at p ≤ 0.05 according to Duncan test.
to T2 treatment while the reduction was by about 81.25% in plants subjected to T3 treatment. Statistical analysis showed the same trend in A levels, which strongly depressed under T2 and T3 treatments, respectively, by about 31.59% and 71.42%. As a consequence of the reduction of gs, transpiration decreased as well soon after salinity stress was imposed and continued to drop, reaching 34.33% and 82.47% of reduction under T2 and T3 treatments, respectively (Fig. 1C). These attained results are in consonance with those of Lycoskoufis et al. (2005), Niu et al. (2010), and Cheng-Jin et al. (2011) who found a significant decrease of gas exchange parameters under salt stress. In fact, the stomatal quick response of stressed plants can stand for a salinity tolerance mechanism (Jones, 1974). Salinity may limit net photosynthesis and stomatal conductance, either because of a CO2 supply limitation resulting from the partial closure of stomata (stomatal function), or by altering the biochemical mechanism of CO2 fixation (not a stomatal function), or by both procedures (Chaves et al., 2003). It was reported that cellular membranes are highly sensitive to stresses (Tayefi-Nasrabadi et al., 2011). Consequently, the accumulation of high concentrations of Na+ and Cl− in the chloroplasts under salinity stress is proven to damage thylakoid membranes (Omoto et al., 2010).
3.2. Changes in water use efficiency in response to salt stress The WUEinstantaneous (A/Tr) and WUEintrinsic (A/gs) were measured for leaf samples and the results are represented in Fig. 2. The WUEinstantaneous was affected by salinity only after a 2-week treatment (Fig. 2A). In fact, it increased significantly with the growing concentration of salt, by about 32.80% and 54.66%, in plants subjected to T2 and T3 treatments. This increase suggests that transpiration rates may have been more affected than CO2 assimilation rates. The WUEintrinsic (A/gs) showed the same trend as WUEinstantaneous (Fig. 2B); but this increase of WUE intrinsic in salt-stressed leaves occurred soon after salinity stress was imposed (1 week treatment). This indicates a greater reduction of gs than of A, and thus, increasing water use efficiency. Similar results were obtained by Thameur and Lachiheb (2012) in their work on saltstressed barley. In fact, in arid regions, WUE is of a vital importance in terms of water saving. According to Araus et al. (2002), high WUE is considered as a strategy to improve the performance of crops in arid conditions.
Please cite this article as: Bacha, H., et al., Impact of salt stress on morpho-physiological and biochemical parameters of Solanum lycopersicum cv. Microtom leaves, South African Journal of Botany (2016), http://dx.doi.org/10.1016/j.sajb.2016.08.018
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Fig. 2. Modification of instantaneous water use efficiency (WUEinstantaneous, A) and intrinsic water use efficiency (WUEintrinsic, B) at 2 dates of leaf sampling (after 1 week and after 2 weeks of stress application). T1: control, 0 mM NaCl; T2: 50 mM NaCl; and T3: 150 mM NaCl. The results are expressed as means ± S.D. (n = 3). Different letters indicate significantly different values at p ≤ 0.05 according to Duncan test.
3.3. Modifications of leaf area The leaf area represents a measure of plant growth, which can be affected by different stresses, including salt stress. Leaf area (LA) was measured for both control plants and plants subjected to salt stress
(Fig. 3.). It has been shown that this parameter increased over time but LA was higher when provided with favorable conditions (control plants) than the salt-stressed ones. For control plants, it increased from 54.74 ± 1.71 cm2 to 87.37 ± 6.38 cm2 between the first harvest and the second. Regarding the effect of salinity treatments, a little
Fig. 3. Leaf area (LA) of tomato cv. Microtom plants measured at 2 dates of leaf sampling (after 1 week and after 2 weeks of stress application). T1: control, 0 mM NaCl; T2: 50 mM NaCl; and T3: 150 mM NaCl. The results are expressed as means ± S.D. (n = 3). Different letters indicate significantly different values at p ≤ 0.05 according to Duncan test.
Please cite this article as: Bacha, H., et al., Impact of salt stress on morpho-physiological and biochemical parameters of Solanum lycopersicum cv. Microtom leaves, South African Journal of Botany (2016), http://dx.doi.org/10.1016/j.sajb.2016.08.018
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decrease in LA was noticed as a consequence of the elevation of salt concentration, and this decrease was accentuated in the second harvest. In fact, the mean value of LA exhibited a reduction in the order of 15.36% and 41.3%, respectively, for the two treatments of 50 and 150 mM NaCl. The reduction in leaf area was due to the decline in leaf gas exchange during salt stress. We found good and significant correlations between LA and A (R = 0.892; p b 0.001 in the second sampling) and between LA and gs (R = 0.781; p b 0.001 in the first sampling and R = 0.809; p b 0.001 in the second sampling). In fact, the reduction of leaf area stands for a morphological adaptive strategy for salt tolerance. The present result is in agreement with the findings of Thameur and Lachiheb (2012) in barley, Vendeland et al. (1982) in soybean, Belaygue et al. (1996) in white clover, Clough and Milthorpe (1975) in tobacco, and NeSmith and Ritchie (1992) in maize, where a decrease in LA of stressed plants was reported. 3.4. Effect of salt stress on foliar chlorophyll content The principal pigment found in the majority of oxygenic photosynthetic organisms is known to be chlorophyll. Chlorophyll content is one of the main factors that reflect the photosynthetic rate (Mao et al., 2007). Some authors suggested that variation in pigment content can provide valuable insight into the physiological performance of leaves and indicates their photosynthetic capacity as well as the presence of stress or diseases (Boquera and Morales, 2010). In the present study, total chlorophyll content was measured for both stressed and unstressed tomato plants cv. Microtom, and the results were illustrated in Table 1. Statistics analysis showed that after a 1-week treatment, chlorophyll concentration exhibited a little reduction under 50 mM NaCl (from 1.13 ± 0.02 mg/g to 1.07 ± 0.03 mg/g) and 150 mM NaCl applications (from 1.13 ± 0.02 mg/g to 1.04 ± 0.03 mg/g). However, after a 2-week salt treatment, chlorophyll content decreased drastically with the increasing concentration of salt to reach 44.82% of reduction under 150 mM of salt concentration. It is probable that the changes in the chlorophyll concentration in tomato plants subjected to high concentration of NaCl, mainly under the 150-mM treatment, is a response to compensating for the loss of leaf area owing to a smaller leaf size. We also found a good and significant correlation (R = 0.896; p b 0.05) between leaf area and total chlorophyll content after a 2-week salt treatment. The chlorophyll content of crop plants is positively correlated with their photosynthetic activity (Gummuluru et al., 1989) and a reduction of chlorophyll level contributes to the inhibition of photosynthesis observed under abiotic stress conditions. This is also confirmed in our study since a good correlation was found between chlorophyll content and net photosynthetic rate (A) (R = 0.879; p b 0.001) at a 2-week salt-stress treatment. Our results are in agreement with those of Noreen et al. (2009) and Kaya et al. (2009) who observed a loss of chlorophyll content in plants subjected to salt stress. The salt-induced alterations in leaf chlorophyll content could be due to impaired biosynthesis or accelerated pigment degradation. The effect is ascribed to an increased level of the toxic cation, Na+ (Yang et al., 2011). Some studies suggest that chlorophyll accumulation under salt stress could be used as a biochemical marker for salt tolerance in different crops (Akram and Ashraf, 2011). However,
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in some other studies, this accumulation is not always associated with salt tolerance. For example, Juan et al. (2005), in their study on saltstressed tomato cultivars which differ in salinity tolerance, observed a weak correlation between photosynthetic pigments and leaf Na+. Thus, chlorophylls cannot be considered as good indicators for salt tolerance in tomatoes. 3.5. Response of phenols to salt-stress treatments In response to abiotic stress, plants have developed a wide variety of highly sophisticated and efficient mechanisms to sense, respond, and adapt to a wide range of environmental changes. A common defensive mechanism activated in plants exposed to stressing conditions is the production and accumulation of phenolic compounds. The antioxidant property of plant phenolic compounds, the metabolic pathways for their biosynthesis, and the involved enzymes are well documented in most important plant species (Balasundram et al., 2006). Some enzymes involved in phenolic metabolism such as polyphenol oxidase (PPO) and peroxidase (POD) generally respond actively to the presence of stress in the plant (Lotfi et al., 2010). In the present study, and after a 1-week treatment, total phenol concentration increased significantly in stressed Microtom leaves, only in response to a dose of 150 mM NaCl, by about 58.28% compared to control leaves (Table 1). However, when extending the period of salt stress to 2 weeks, we observed a remarkable increase of phenols in stressed plants of both tested treatments, by an increase of 70.45% for T2 leaves and 82.82% for T3 leaves. Our results are in consonance with Al Hassan et al. (2015) who observed a significant increase of phenolics in stressed tomato leaves under moderate and high salt concentrations in cherry tomato (S. lycopersicum L. cv. cerasiforme). The increase in antioxidant phenolic compound levels in leaves can be considered as part of the response induced to cope with oxidative stress induced by salinity. Thus, salt-stressed plants might represent potential sources of polyphenols, by increasing polyphenol concentration in the tissues, which is an issue that is directly tied with human health since these compounds are known to be bio-active compounds (Jemai et al., 2008). In fact, optimal polyphenol content would be obtained using stress-tolerant species (De Abreu and Mazzafera, 2005). 4. Conclusions The mechanism of adaptation to saline stress results in morphological, physiological, and biochemical responses. In this study, tomato plants of cv. Microtom adapted to salt stress by reducing leaf area, stomatal conductance, and minimizing water loss by transpiration. These led to greater reduction in photosynthetic rate, especially after 2 weeks of 150 mM salt treatment, and a decrease of the content of the major photosynthetic pigment (chlorophyll). Besides, Microtom has the ability to increase the efficiency of water use, which is of vital importance in terms of water saving and which represents a strategy to improve the performance of crops in arid conditions. In consequence to the enhanced production of ROS (reactive oxygen species) generated by salt stress, phenol accumulation was stimulated in the leaves of Microtom as a common defensive mechanism. These facts could be
Table 1 Variations in the foliar contents of chlorophylls and total phenols following increasing concentration of salinity and period of salt treatment. Parameters
Total chlorophylls (mg g
Sampling date (after stress application) −1
Total phenols (mg eq. GA g
FW)
−1
DW)
After 1 week After 2 weeks After 1 week After 2 weeks
Concentrations of salt treatments T1 (0 mM)
T2 (50 mM)
T3 (150 mM)
1.13 ± 0.02 a 1.16 ± 0.03 a 0.78 ± 0.12 b 0.80 ± 0.1 c
1.07 ± 0.03 a 0.92 ± 0.04 b 0.81 ± 0.11 b 2.64 ± 0.20 b
1.04 ± 0.03 b 0.64 ± 0.02 c 1.87 ± 0.21 a 4.54 ± 0.19 a
Results are expressed as means ± SD (n = 3). a, b, c, d: Values in the same row with different letters showed statistically significant differences (p b 0.05) according to Duncan test.
Please cite this article as: Bacha, H., et al., Impact of salt stress on morpho-physiological and biochemical parameters of Solanum lycopersicum cv. Microtom leaves, South African Journal of Botany (2016), http://dx.doi.org/10.1016/j.sajb.2016.08.018
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one of the strategies used by these plants to tolerate the severe conditions imposed by soils with a high salt concentration in arid lands. Though some other yet unidentified factors could also be involved in the survival of these plants under continuous salt stress.
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Please cite this article as: Bacha, H., et al., Impact of salt stress on morpho-physiological and biochemical parameters of Solanum lycopersicum cv. Microtom leaves, South African Journal of Botany (2016), http://dx.doi.org/10.1016/j.sajb.2016.08.018