Introduction

French marigold (Tageta patula) belongs to the Asteraceae family (Hassani, 1998), which is used as a cut or pot flower and marginal planting in landscape. Nowadays, marigold is commercially grown for the extraction of carotene pigments, especially xanthophylls. The leaves of marigolds contain special aromatic oils. Marigold oil has therapeutic applications, and is insect repulsive. Marigold cultivation is also useful for reduction of nematode populations in the soil. The application of marigold in rotation of cultivation programs can be a replacement for nematicides to reduce populations of scabies-agent nematode (Meloidogyne spp.). Nowadays, the existing species of marigolds are cultured in various climates, where problems of environmental stresses such as freezing, salinity and drought are imposed to plants after their establishments. Research on ornamental and non-bearing plants is usually limited due to low economic benefit of these plants. However, the environmental benefits, purification role, aesthetic appeal, soil fixation, erosion prevention, windbreaker role and air modification of these plants highlight the importance of basic studies on these plants.

With regards to the utilization of marigold as a summer flower in green spaces and city landscapes, especially in regions with saline soil, it is important to investigate the response of this plant to salt stresses. Plants uses two mechanisms to cope with the stress factor: in resistance mechanism when plants encounter to slight stresses they keep their metabolic activities at a high level (similar non-stress conditions), on the other hand when they encounter to severe stresses their metabolic activities decreases. In avoidance mechanism, plant reduces its metabolic activities and shows dormancy under extreme stress conditions. In agriculture, scientist paid more attention to stress avoidance mechanism by using plant breeding methods. In nature, both mechanisms are used by different plant species. Salinity and drought have attracted more attention among abiotic stresses (Alizadeh, 2005). Salinity in the agricultural sciences is one of the characteristics of soil or water. It’s due to excessive presence of ions in soil or water. Among ions, one valence types such as sodium (Na) and potassium (K), despite their differences, have effective and determinant roles in plant biochemical properties. Saline soils are characterized with EC>4 mmos/cm, exchangeable sodium percentage (ESP) less than 15 and pH<8.5. These soils are known as ‘white alkali’—in Russia ‘Solonchak’— and, according to the new American taxonomy procedure, ‘Solarthia’ (Baybordi and Koohestani, 2004).

Salinity stress leads to reductions in transpiration rate and consequently delays plant growth and reduces crop yield. Generally, salinity stress could be defined as the general effects of salts on plant growth (Hasheminia et al., 1997). Results of previous studies showed that the influence of salinity on plant growth and metabolism results from the reduction of osmotic potential due to salt accumulation (Levitt, 1980) and ion toxicity (Wiggans and Gardner, 1989).

Salinity stress affects various aspects of plant growth and metabolism. Photosynthesis (Staples and Toenniessen, 1984), nitrate reduction (Polard and Wynjones, 1979) and internal unbalancing of plant growth regulators (Shah and Loomis, 1965) are the most important parameters that have been evaluated for plant responses to stress conditions. In saline media, high concentrations of ions in the rhizosphere can affect nutrient uptake by the root (Birendra et al., 1996), which has been mainly attributed to the presence of high concentrations of chloride and sodium (Abo-Kaseam et al., 1995). Decline in the uptake of potassium (Nassery, 1979), calcium, magnesium (Al-Harbi, 1995), ammonium and nitrate (Soliman et al., 1994) are examples of negative effects of salinity stress in relation to plant nutrition. Potassium is involved in stomata conductance, enzyme activity, and transportation within the plant. It has been proved that different strategies are used by different plant species to tolerate salinity stress.

Marigold is a very tolerant plant species to salinity stress based on vegetative growth and flower size, but based on flower number it is a very sensitive plant (Decital and Morris, 1987). Depends on plant species, presence of ions in soil or water can stimulate, inhibit or counteract with germination. Salinity stress generally leads to a delay in germination and a reduction in plantlet growth (Mirmohammadi-Meibodi and Ghareyazi, 2002). Influence of salinity on plant growth varies at different stages of plant growth; also plant reaction to salinity is also changed at different stages of plant growth. Furthermore, response of different species and even their cultivars are also different in response to salinity stress. For example, rice can tolerate high concentrations of salt at germination stage (more than 30 ds/m) but it is sensitive to salinity at the primary stage of plantlet growth and flowering (Sarrafi, 1993). Sugar beet is sensitive at the germination stage but it is tolerant at the other stages (Shannon, 1998); maize is tolerant at the germination stage, its sensitivity to salinity increases at the plantlet stage and decreases at the clustering and seed filling stages (Mass and Hoffman, 1987).

Salinity stress can affect different stages of plant growth, however, the sensitivity of some stages are more than the others. Germination is one of the most critical and sensitive stages of plant growth to salinity stress. Low germination in the saline soils causes poor establishment and weak production of the plant. At the germination stage Tolerant plants to salinity finish vegetative stage with more success. Pill and Killan (2000) evaluated effects of osmo-priming on germination percentage and germination rate of parsley in saline stress conditions by different treatments. They showed that in the saline soils, uptake of some elements such as calcium, magnesium and potassium reduces because of competition with sodium (Yeo and Flowers, 1986). High sodium concentrations reduces uptake of other cations and as a result unbalances cations uptake into the plant. For instance, declines in calcium, magnesium and potassium uptake have been reported as a consequence of high sodium concentration in the root medium (Lauchi and Epsetin, 1984). In barley, excessive NaCl concentrations lead to calcium deficiency in plant tissues (Brown and Hayward, 1970). It has been reported that in saline conditions, sodium chloride competes with other cations leading to growth retardation. Apart from osmotic stress, growth of bean, pea and sweet orange is reduced due to potassium defficiency (Hassan-Shahi et al., 2008). Salinity in the form of NaCl, causes potassium deficiency in the lower parts of squash and sweet clover plants. Therefore, the main reason for growth disorders by NaCl is due to the inhibition of other minerals uptake in competition with sodium. Although high concentrations of sodium can decrease potassium uptake, Its low concentrations may increase potassium uptake. High concentrations of sodium can also inhibit calcium uptake (Mirmohammadi-Meibodi and Ghareyazi, 2002).

Nitrogen plays a key role in plant nutrition. In fact, plant life is not possible without this element. Contrary to other elements, nitrogen is not available in the soil materials and rainwater and atmosphere (through biological agents such as nitrogen-fixing bacteria) are the sources of nitrogen for plants. In addition to nitrogen uptake by plants, leaching, erosion, ammonia sublimation and elemental nitrogen are the sources of nitrogen loss from the soil (Ghazan-Shahi, 1999). Usually increasing uptake and accumulation of Cl^- is associated with a reduction in NO₃^- in the plant shoot. Many researchers attribute this subject to the antagonistic effect of Cl^- on NO₃^- uptake (Kafkafi et al., 1982). Negative effects of salinity on root water uptake have been proposed as another reason for NO₃^- deficiency following salinity stress (Lehle et al., 1992). It has been reported that in saline conditions, the uptake of nitrate and ammonium ions reduces, and finally the plant adapts to nitrogen deficiency. This causes a reduction in amino acids and protein synthesis. Furthermore, nitrate reduction in the root is hampered by exposure to salinity stress (Odegbaro and Smith, 1969). This study aimed to investigate the interactions between salinity and ammonium nitrate on seed germination, vegetative growth and sodium and potassium concentrations in French marigold flowers (Tegeta patula).

Material and Methods

To assess the effect of ammonium nitrate on the reduction of salinity stress in French marigolds (Tageta patula), an experiment was conducted as a factorial arrangement based on a completely randomized design (CRD). The first factor was different levels of ammonium nitrate including 0, 15 and 30 g/l and the second factor was different concentrations of NaCl salinity , including solutions with electrical conductivity of 0.49 (tap water as control treatment), 2, 4, 6 and 8 mmos/cm with four replications. The marigold seeds were prepared from a seed production centre in Esfahan, Iran. Benomyl was used to the seeds before culture. Three seeds were sown in 10 l pots containing manure, sand and agricultural soil (1:1:1) and were irrigated daily using the salt solutions. In this study, germination rate, plant height, lateral shoot number, root and shoot fresh weights, root and shoot dry weights, and amount of sodium and potassium were measured. To determine germination rate, the germinated seeds were daily counted, and germination rate was calculated by Equation (1) (Khan et al., 1997), where GR, Ni and Di are germination rate, number of germinated seeds, and number of days from initiation of the experiment, respectively.

 (1)

In order to measure sodium and potassium levels, the shoot samples were dried in oven 70 °C for 24 h. Then 0.2 g of the powdered sample was placed in 550 °C for 5 h until discoloration of the samples. The plant extracts were then prepared using 2 ml of HCl and boiling water and 50 ml of distilled water. Sodium and potassium concentrations were measured by a flame photometer device (Aboutalebi et al., 2008). The obtained data was analysed by using MSTAT-C software and the mean averages were compared by Duncan’s multiple range test (DMRT).

Results and Discussion

The plants irrigated with a solution of 8 mmos/cm were died after the germination stage, Therefore, this salinity level was removed from the analysis.

Germination rate (GR)

Based on the results of analysis of variance, salinity and ammonium nitrate levels had no significant influence on GR. There is no significant interaction between these factors (Table 1). According to the results of the mean comparisons, the highest GR was observed at a salinity of 2 mmos/cm, but it was not significantly different compared to the other concentrations (Table 2). In all growth stages, salinity stress negatively affected the plant growth, but variation in the sensitivity was observed for different stages. Germination is one of the most critical and sensitive stages of plant growth due to exposure to salinity stress (Table 3).

Low germination in the saline soils resulted in poor establishment and production of the plants; at this stage tolerant plants were successfully finished their vegetative stage. The results of this study showed that salinity levels had no significant influence on GR. The highest GR was observed in plants treated with 30 g/l ammonium nitrate but no significant difference was observed among ammonium nitrate levels (Table 2). In evaluation of the interactions between the two factors, the highest GR was observed in 30 g/l ammonium nitrate and control treatments (0.84) and the lowest was observed in 15 g/l ammonium nitrate and control treatments (0.62) (Table 4).

Table 1. Analysis of variance for the measured parameters

^ns not significant; ^* significant difference at P<0.05 and ^** significant difference at P<0.01

Plant height

Salinity levels had a significant influence (P<0.01) on plant height, but no significant effect was observed for application of different levels of ammonium nitrate. Moreover, significant interaction (P<0.05) was observed between ammonium nitrate and salinity stress (Table 1). Longest plants (14.6 cm) were observed in the control treatment (Table 3). Negative effects of salinity stress can be due to water deficiency, ion toxicity, ion unbalancing, or combination of them, however, it is hard to exactly conclude which of these problems is the main factor that influenced the plant growth. In the present study plant height was decreased under saline conditions. Growth limitations resulting from salinity stress can be as a result of water stress and ion deficiency (Table 3). Based on the result of previous studies, low water potential in the external plant medium is the more important limiting factor for growth inhibition in several species than ion toxicity under salinity stress conditions (Liu and Staden, 2001). The main effect of sodium acetate is reduction in root volume, leading to decline in water absorption and nutrient uptake (Salisbury and Ross, 1992). Flowers and Yeo (1995) divided the effect of salinity into two short-term and long-term effects: short-term effects can be seen following few days of salinity stress that usually results in shoot growth inhibition due to salt-induced water deficiency. Salinity negatively influences growth of leaf cells. Reduction in leaf growth may occur even if ions desorptions and absorptions are equal, because maintenance of a suitable ion gradient for ion distribution and for the production of different solvents require consumption of energy (Volkmar et al., 1998). Hung and Cox (1988) reported that marigolds showed symptoms of toxicity when EC of the irrigation water exceed 7.9 ds/m. In such situations, leaves turned to green-yellow and bronze two weeks after imposing of salinity stress, and necrotic symptoms appeared in the margin of its mature leaves.

The Tallest and shortest plants (10.07 and 9.22 cm) were observed in 30 and 15 g/l ammonium nitrate, respectively (Table 2). In evaluating the effect of interactions between ammonium nitrate and salinity, the tallest plants were observed by application of 30 g/l ammonium nitrate and control treatment for the salinity (15.0 cm). In control and salinity 2 mmos/cm, plant height was increased in high concentrations of ammonium nitrate. Salinity decreased plant height. Average of plant height was 14.6 cm in the control treatment, while it decreased to 5.3 cm in plants exposed to 6 mmos/cm salinity (Table 4). There was no significant difference among the levels of ammonium nitrate in each salinity level.

Number of lateral shoots

Salinity levels had a significant influence (P<0.01) on the number of lateral shoots, while there was no significant effect resulting from the application of different levels of ammonium nitrate. Interactions between ammonium nitrate and salinity were not significant (Table 1). Highest number of lateral shoots was obtained in control treatment (6.8 shoots) and the lowest was seen in the plants exposed to 6.0 mmos/cm salinity (Table 3). Number of lateral shoot was decreased by increasing salt concentrations in the irrigation water. Different plants species and varieties show different levels of resistance to salinity. Salt composition, soil properties, weather, moisture, and plant variety can influence harmful effects of salt on the plants (Mass and Hoffman, 1977). Differences in tolerance to salinity have been observed between plants and even between different species of one plant (Hughes et al., 1975). Number of lateral shoot decreases with increasing levels of ammonium nitrate (Table 2). The highest number of lateral shoot was observed in the control treatment and 15 g/l ammonium nitrate (7.5 shoots). By increasing salinity level, the application of ammonium nitrate had no influence on the number of lateral shoot (Table 4).

Root fresh weight (RFW)

Salinity levels had a significant influence (P<0.01) on RFW but there was no significant effect from application of different levels of ammonium nitrate. Interactions between ammonium nitrate and salinity were significant (P<0.01; Table 1). The highest RFW was observed in the control treatment (1.84 g) and the lowest RFW was observed in plants that treated with6.0 mmos/cm salinity (0.27 g; Table 3). Among ammonium nitrate levels, the highest RFW was obtained from 0 g/l and the lowest RFW was observed in plants treated with 30 g/l ammonium nitrate (Table 2).

Table 2. Effects of different levels of ammonium nitrate on vegetative parameters and potassium and sodium levels in okra

^†Means in each row with similar letters are not significantly different (P<0.05) according to DMRT.

Table 3. Effects of different levels of electrical conductivity on vegetative parameters and potassium and sodium levels in marigold plants

^†Means in each row with similar letters are not significantly different (P<0.05) according to DMRT.

Table 4. Interactions between ammonium nitrate (AN) and salinity levels in marigold plants

^†Means in each column with similar letters are not significantly different (P<0.05) according to DMRT.