Introduction

Salt stress is one of the major abiotic stress factors that affect almost every aspect of physiology and biochemistry of a plant, resulting in a reduction in its yield (Foolad, 2004; Tatar et al., 2010; Babu et al., 2012, Hamdia and Shaddad, 2010, 2014, 2016). Thus it is a serious threat to agricultural productivity especially in arid and semi-arid regions (Parvaiz and Satyawati, 2008). Salt stress causes hyperosmotic stress and ion disequilibrium, thereby disabling the vital cellular functions of a plant. Reduced availability of water, increased respiration rate, altered mineral distribution, membrane instability, failure in the maintenance of turgor pressure are some of the events that prevails during this stress episode. Plants try to withstand these stresses either by tolerating it or by adopting a dormant stage (Cuartero et al., 2006). Salt tolerance is a complex trait which involves numerous genes and various physiological and biochemical mechanisms (Cuartero and Moreno, 2006). Proline content of salt-stressed plants was previously reported in different species (Özcan et al., 2000; Turan et al., 2007). Lutts et al. (1999) suggested that proline accumulation in rice under salinity was most likely a symptom of injury rather than an indicator of increased tolerance. However, Singh et al. (1996) emphasized the protective role of proline under salinity. Mansour (1998) also suggested additive effects of proline upon salt tolerance via increased cell membrane stability. Based on our results, we cannot distinguish between higher proline concentrations in leaf cells being an indicator of injury or a protection mechanism. The decrease in K⁺/ Na⁺ ratio and proline content in leaves (indicated that the proline content of leaves may act as an indicator of Na⁺ uptake and allow the identification of tolerant varieties, which effectively exclude Na⁺ from leaf tissue either by exclusion or compartmentalization in root.

Phytohormones suggested playing important roles in stress responses and adaptation (Sharma et al., 2005; Hamdia and Shaddad, 2013, 2014; Hamdia, 2016). It is thought that the repressive effect of salinity on seed germination and plant growth could be related to a decline in endogenous levels of phytohormones (Debez et al., 1997; Hamdia, 2016). Thus the present work carried out to study the response of maize, wheat, cotton, broad bean and parsley plants to salinity and phtohormnal treatments concerning proline and sodium as osmotic stress signal components.

1 Results and Analysis

1.1 Dry matter

Dry matter of shoots and roots of maize plant was progressively decreased with rise of osmotic pressure, the percent of reduction at-1.2 MPa NaCl level was 27.3% and 40% in shoots and roots respectively (Table 1). In shoots and roots of wheat plant while dry matter tended to smoothly increase in shoots, it tended to smoothly decrease in roots up-0.6 MPa NaCl level, afterthat a significantly reduction was detected at the higher levels of osmotic pressure. The percent of reduction was 15.4% and 32.9% of shoots and roots of wheat plant at -1.2 MPa NaCl level (Table 1). It is an elevation trend on the production of dry matter at -0.3 MPa NaCl level of shoots and roots of cotton plant, afterthen a reduction was observed, the percent of reduction at -1.2 MPa NaCl level was 32.9% and 28.6% in shoots and roots (Table 1). In broad bean plant dry matter remain more or less unchanged up to -0.9 MPa NaCl level, obovethat a reduction was recorded. In broad bean plant, there is an enhancement effect in the production of dry matter at -0.3 MPa NaCl level, then a reduction was recorded, the percent of reduction was 20.8% and 13.9% in shoots and roots of broad bean plants (Table 1). While dry matter run with stable values from control up to -0.9 MPa NaCl level, it shows a smooth reduction at -1.2 MPa NaCl level in shoots of parsley plant. However, a significant reduction in dry matter was observed in roots of parsley plant.There is a surprising data that dry matter generally in shoots of maize, wheat, broad bean and shoots and roots of parsley produce more or less a stable trend with increasing osmotic stress.

1.2 Proline

Proline content was significantly decreased in both shoots and roots of maize plants; this reduction was most obvious in roots starting up to -0.3 MPa NaCl level than in shoots (Figure 1a; Figure 1b). The percent of reduction was 20% and 66% in shoots and roots respectively. However, in shoots and roots of wheat plant proline was markedly increased with increasing osmotic pressure especially at higher osmotic stress (Figure 2a; Figure 2b). This activation was more pronounced in shoots than in roots, the percent of activation was 233.3% and 109.5% at -1.2 MPa NaCl level in shoots and roots respectively as compared with control plants. While proline content was markedly decreased in shoots of cotton plant, in roots it remain more or less unchanged up to -0.9 MPa NaCl level, after this levels a reduction was recorded at -1.2 MPa NaCl level (Figure 3a; Figure 3b). In broad bean an elevation trend with stable values was detected in shoots of broad bean plant starting -0.3 MPa NaCl level (Figure 4a; Figure 4b). In broad bean roots proline tended to decrease at -0.3 MPa NaCl level and progressively decreased at -0.6 MPa, - 0.9 MPa and -1.2 MPa NaCl level (Figure 4). The percent of reduction at -1.2 MPa was 43.6% MPa NaCl level. There is a smooth increase trend in shoots and roots of parsley plant, the percent of activation at -1.2 MPa NaCl level was 25% and 25.5% in shoots and roots respectively (Figure 5a; Figure 5b). Also, there is more or less a stable trend in proline content of shoots maize, broad bean and shoots and roots of parsley plants.

1.3 Sodium

There is an interval increase between the five tested plants in sodium content of shoots and roots (Figure 1; Figure 2; Figure 3; Figure 4; Figure 5). The increase values in sodium content at -1.2 MPa NaCl level reach 26 - folds, and 21.5- folds, 20- folds, 25- folds, 22- folds, 8- folds, 49- folds, 30- folds, 49- folds and 14.5- folds of shoots and roots of maize, wheat, cotton, broad bean and parsley plants respectively. The higher values of increase in sodium content were represented in shoots of parsley and broad bean plant. The opposite situation, the lower values of increase was represented in shoots of wheat plants.The higher increase values in sodium content were represented in the roots of broad bean plant. The lower increase values were represented in the roots of cotton plants.

1.4 Phytohormones

There are a marked and significant increase in the production of dry matter in shoots and roots of maize, wheat, cotton, broad bean and parsley plants when spraying the vegetative parts with 100 ppm either GA₃ or kinetin (Table 1). It is worthy to note that the production of dry matter at -1.2 MPa NaCl level produce a values mostly higher than those of control plants in shoots and roots of maize, wheat, cotton, broad bean and parsley plants when treated with GA₃ or kinetin (Table 1; Table 2; Table 3; Table 4). Except of this trend roots of maize, shoots of wheat and broad bean reach near the values of control and roots of parsley plants (Table 1).When treated salinized maize, wheat, cotton, broad bean and parsley plants with either 100 ppm GA₃ or kinetin showed a different response on the accumulation of proline. Plants treated with GA₃ reduced the accumulation of proline in shoots and roots of maize broad bean and parsley plants while tended to increase in shoots and roots of wheat and cotton as compared with control plants (Figure 1; Figure 2; Figure 3; Figure 4; Figure 5). Kinetin treatment induced in most cases a reduction in proline content of wheat roots, shoots and roots of broad bean and parsley plants. While stimulation in the accumulation was observed in shoots of maize and shoots and roots of cotton plants was induced, proline content remain unchanged in roots of maize and shoots of wheat. Phytohormonal application reduced the accumulation of sodium in shoots and roots of maize, wheat, cotton, broad bean and parsley plants (Figure 1; Figure 2; Figure 3; Figure 4; Figure 5). Except of this trend sodium content showed an increase of Na⁺ content when treated with GA₃ in roots of maize plants (Figure 1b).

2 Discussion

Based on our results, we cannot distinguish between higher proline concentrations in plant cells being as an indicator of injury or a protection mechanism. The increase of Na⁺ and proline content in these tested plants indicated that the proline content may act as an indicator of Na⁺ uptake and allow the identification of tolerant plants, which effectively exclude Na⁺ from plant either by exclusion or compartmentalization (Zhu 2001; Kumara et al., 2003; Garthwaite et al., 2005; Garacia et al., 1997; Garacia et al., 2004). In maize and cotton plants while Na⁺ increased in shoots and roots, proline content decreased in shoot and roots of these plants. This concomitant with reduction of dry matter in both organs of the maize and cotton plants. i.e. proline cannot be considered as a signal of high osmotic stress. The percent of reduction in proline content and dry matter of shoots and roots at -1.2 MPa NaCl level was 20%, 66%, 28%, 40% for maize and 60%, 44%, 33% and 29% for cotton plants. This indicate that a-Root is the most organ exposed to high osmotic stress in the soil and the most sensitive than shoot to saline injury in maize plants and in cotton shoot is the most sensitive organ. B- Proline was more translocated form root to shoot with mass flow in maize while it accumulated in root of cotton plant. This reflected an increase the accumulation of Na⁺ reached 26 - folds, 21.5- folds for shoots and roots of maize plants and 27- folds and 8- folds for cotton plants respectively compared with control plants.

On the other side the ratio of increase was the same in shoots and roots of parsley plants. This reflected a reduction in dry matter of these three plants especially in parsley roots. Moreover this reduction was more in roots organ of wheat and parsley plants than in shoots while an opposite situation was detected in broad bean plants. This strategy of proline and dry matter was concomitant with the increase of Na⁺ content. The organ that the most reduced dry matter accumulated Na⁺ only as in wheat, another accumulated both Na⁺ and proline was recorded as in shoots of broad bean. However, the organ which more reduces dry matter accumulated more proline as roots parsley. The percentage of increasing in Na⁺ was 20%, 25%, 49%, 30%, 49% and 14.5 - folds for shoots and roots of three tested plants. Proline accumulation, a common metabolic response of plants subjected to salinity stress, is considered to be involved in stress-tolerance mechanisms (Majid et al., 2012; Lutts et al., 1999; Nanjo et al., 1999). In the present study, a significant increase in proline content was found. One distinctive feature of most plants growing in saline environments is the accumulation of proline (Kumara et al., 2003) and it has been inferred that there may be a relationship between cellular proline level and cell turgidity via osmotic adjustment (Hayashi et al., 2000; Kumara et al., 2003). The ability to exclude sodium from the shoot is an important determinant of salt tolerance in both monocots and dicots (Garthwaite et al., 2005; Zhu, 2001). However, the salt sensitive Arabidopsis ecotype Columbia grown at moderate salt concentrations keeps the shoot Na⁺ concentration much lower than do coastal, more salt tolerant ecotypes (Rus et al., 2006). This strategy has a limited capacity since growth of Arabidopsis ecotype Colombia at high salt is severely limited due to a strong increase in shoot Na⁺ concentration suggested that Arabidopsis may use mechanisms involved with Na⁺ tissue tolerance, such as intracellular compartmentation and increased accumulation of compatible solutes, as was hypothesized before Munns and Rester (2008) and apparently, some of the potato cultivars respond in this way. Phytohormonal application was markedly and significant reduced mostly the accumulation both stress markers sodium ion and proline in shoots and roots of maize, wheat, cotton, broad bean and parsley plants.This can be reflected on the accumulation of metabolities which finally affected on the production of dry matter in shoots and roots of the five tested plants. Shevyakova et al., (2013) study ABA protective action under salinity can be realized through the weakening of oxidative stress (a decrease in MDA content) and the regulation of PA, proline, and CK metabolism, which has a great significance in plant adaptation to injurious factors. Hamdia (2013) Hamdia and Shaddad (2014) and Hamdia (2016) can be concluded that the GA₃, Kinetin and IAA regulate the disturbances of metabolities and neglected the negative effects of the accumulation of ethylene especially in plants treated with IAA under stress conditions which in turn resulted in a pronounced alleviated the drastic effects of salt.

3 Materials and Methods

Five plant species Maize (Zea mays), Wheat (Triticum aestivum), Broad bean (Vicia faba ), Cotton (Gossypium herbaceum ) and Parsley plants (Petroselinum crispum) were grown in plastic pots in the soil without NaCl (control) and under salinization levels corresponding to osmotic potential of NaCl solution, -0.3 MPa, -0.6 MPa, -0.9 MPa and -1.2 MPa at the first group. Saline solutions were added to the soil in such a way that the soil solution acquired the assigned salinization levels at field capacity. Plants were irrigated every other day with 1/10 Pfeffer’s nutrient solution for two weeks. The previous experiment was repeated for GA₃ and kinetin (100 ppm) treatments as the second and third groups. Solutions were sprayed three times (5 intervals) by spraying the shoots system of the growing plants (each pot with 10 cm³ of GA₃ or kinetin solutions). The control plants were sprayed with distilled water. A week after the later spraying, the plants was used for analysis after 45-days. Dry matter was determined after drying plants in an aerated oven at 70ºC to constant mass. Proline content was determined according to Bates (1973). Sodium was determined by Flame-photometer according to method of Williams and Twin (1960).

The experimental data were subjected to the one way analysis of variances (ANOVA test) using the SPSS version 11.0 to quantify and evaluate the source of variation and the means were separated by the least significant differences, L.S.D. at P level of 0.05% (Steel and Torrie, 1960). The percentage presented in the following tables was calculated by the data of survival plants with salinization treatments and with hormonal applications either GA₃ or kinetin 200 ppm. This by measuring dry matter, sodium and proline content at control plants and hormonal treatments of shoots and roots of maize, wheat, cotton, broad bean and parsley plants. The data was compared by plants grow at control (untreated) and the other with GA₃ or kinetin applications.

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