Physiological Adaptation of Three Halophytes from the Same Ecological Habitat and Analysis of their Supporting Soil

Photosynthetic pigment

Chl content was estimated in fresh leaves as described by Witham et al. (1971).^[29]

Antioxidant enzyme activity

• a)Peroxidase activity : measured after 2 min in fresh, weighted leaves using the method described by Fehrmann and Dimond (1967). ^[30]

• b)Phenoloxidase activity : measured after 45 min in fresh, weighted leaves that were extracted according to Hammad and Ali (2014). ^[31]

Proline, total carbohydrates, soluble sugars, and free amino acids (protein)

The total contents of proline, total carbohydrates and soluble sugars, and free amino acid concentrations were estimated according to the methods described by Bates et al. (1973),^[32] Dubois et al. (1956),^[33] and Rosen (1957),^[34] respectively.

Soil texture

The soil texture analysis at the surface (0-15 cm) and the subsurface (15-30 cm) revealed that the soil under L. shawii and P. undulata was overall moderately coarse sandy at both depths. In contrast, the soil under T. nilotica was fine sandy at both depths [Table 1].

Bulk density (Db)

Bulk density analysis showed the highest value in L. shawii at the soil’s subsurface (15-30 cm) with a mean of 1.71 g cm^{- 3}, whereas the surface soil (0-15 cm) for the same plant had a mean of 1.54 g cm^{- 3}. On the other hand, T. nilotica soil surface recorded the lowest bulk density of 1.36 g cm^-3, and the soil subsurface for the same plant was 1.6 g cm^-3 [Table 2].

Dp

DP analysis revealed a range of 2.36-2.54 g cm^-3 in the surface soil and 2.59 to 2.72 g cm^-3 in the subsurface soil. The highest values were observed in L. shawii soil at the soil’s subsurface (15-30 cm), while the lowest values were recorded for T. nilotica soil at the soil’s surface (0-15 cm) [Table 2].

Total porosity (P%)

The highest total porosity was found in T. nilotica soil, whereas L. shawii had the lowest. The P% of T. nilotica soil was 42.63% and 40.23% at the soil’s surface and subsurface, respectively. Whereas L. shawii had a P% of 39.53% at the surface and 37.17% at the subsurface [Table 2].

HC

HC analysis showed high values of 23.33-26.33 cm h^-1 at the soil surface and 23.67-27 cm h^-1 at the subsurface. The lowest HC at the subsurface level was seen in T. nilotica soil, whereas the highest was found in P. undulata [Table 2].

pH

The pH of all selected soils was 7.8, indicating slightly alkaline soil at 25°C [Table 3].

EC

The EC analysis showed the highest EC value at the soil surface for L. shawii (1,460 ppm), whereas P. undulata had the lowest EC value (231.67 ppm). On the soil’s subsurface 15-30 cm depth, T. nilotica showed the highest EC of 10,900 ppm, while P. undulata had the lowest 1,066.67 ppm [Table 3].

Soil cations (Ca ²⁺, Mg ²⁺, Na⁺, K⁺)

Cation analysis revealed that Na⁺ was the most abundant, followed by Ca2⁺, Mg ²⁺, and K⁺. Furthermore, Ca ²⁺ concentration in T. nilotica was the highest at the subsurface soil (30.47 mEq L^-1), whereas P. undulata exhibited the lowest concentration (3.63 mEq L^-1). Mg ²⁺ estimation showed the highest level in L. shawii at 0-15 cm depth (2.5 mEq L^-1), while P. undulata had the lowest (0.2 mEq L^-1). Similarly, P. undulata recorded the lowest at 15-30 cm depth (2.0 mEq L^-1), whereas T. nilotica showed the highest concentration of Mg ²⁺ (17.93 mEq L^-1). K ⁺ was the lowest cation among the others. T. nilotica soil demonstrated the highest level at subsurface, whereas P. undulata had the lowest at both surface and subsurface [Table 3].

Soil anions (CO₃^-2, HCO₃^-, Cl^-, SO₄^-2)

Anions’ analysis showed Cl^- had the highest level among the anions in all halophytes, followed by SO₄^-2 and CO₃^-2 + HCO₃^-, respectively. Furthermore, L. shawii had the highest Cl ^- level at the soil’s surface (15.27 mEq L^-1), whilst T. nilotica had the highest at the subsurface (120.13 mEq L^-1). On the other hand, P. undulata showed the lowest Cl^- levels at both surface and subsurface (2.5 mEq L^-1 and 11.23 mEq L^-1, respectively) [Table 3].

Water sample analysis

The water sample showed a pH value of 7.16 and an EC of 112,233.3 ppm. Moreover, the soluble cations analysis showed that the soluble Na⁺ was the most abundant (1,345 mEq L^-1), whereas the soluble K⁺ was the lowest (21.3 mEq L^-1). Furthermore, Cl^- had the highest estimation among the soluble anions (1,480.3 mEq L^-1), whilst CO₃^--+HCO₃^- estimation was the lowest (23.3 mEq L^-1) [Table 4].

SAR

The water sample is classified as fourth class, with the highest Na⁺ content [Table 4].

CaCO₃

CaCO₃ analysis showed the highest value in L. shawii soil surface (6.07%), whereas P. undulata had the lowest concentration (4.4%). At the soil subsurface, T. nilotica had the highest level (5.83%), while P. undulata had the lowest (4.47%) [Table 5].

CEC

CEC estimation revealed the highest value in T. nilotica soil subsurface (12.3 Meq/100g) and 11.27 Meq/100g at the soil surface, whereas L. shawii had the lowest at both depths [Table 5].

OM

OM content was relatively high in the soil of T. nilotica, reaching 0.83% and 0.74% at the soil surface and subsurface, respectively. Whereas the soils of the P. undulata and L. shawii showed similar OM values [Table 5].

GC

The GC analysis indicated close values, with a low range of 0.36-0.78% in the selected soils. T. nilotica was slightly higher than the others [Table 5].

ESP

ESP analysis showed a wide range of 6.57-7.63% at the soil surface and 7.7-11.4% at the soil subsurface in selected halophytes. The highest values were observed in L. shawii soil at the soil’s subsurface (15-30 cm), while the lowest values were recorded for p. undulata soil at the soil’s surface (0-15 cm) [Table 5].

Growth characters

L. shawii showed the largest leaf area compared to the other plants, with an average of 2.34 cm ²/plant. In contrast, the T. nilotica has the smallest leaf area, with an average of 0.66 cm²/plant [Table 6, Figure 5].

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Figure 5:

Estimation of leaf area of the selected halophytes. L. shawii had the largest leaf area of 2.34 cm²/plant, whereas T. nilotica had the smallest (0.66 cm²/plant).

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Photosynthetic pigment

Photosynthesis pigment estimation showed that P. undulata had the highest Chl A and B concentrations, as well as carotene, total Chl, and the Chl/carotene ratio, compared to the others. In comparison, L. shawii showed the lowest in all [Table 6, Figure 6].

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Figure 6:

Estimation of Chl pigments (A and B), total Chl, and carotene in the selected halophytes. P. undulata had the highest concentration of Chl A, Chl B, carotene, and total Chl/carotene, while L. shawii had the lowest. On the other hand, T. nilotica had the highest Chl A/B, whereas P. undulata had the lowest.

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Antioxidants

Peroxidase enzyme estimation showed the highest in L. shawii (477.33 u/g fresh weight/h) and the lowest in P. undulata (325 u/g fresh weight/h). Additionally, P. undulata showed the highest in polyphenol oxidase analysis (38.94 u/g fresh weight/h), whereas T. nilotica had the lowest (30.72 u/g fresh weight/h) [Table 7, Figure 7].

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Figure 7:

Estimation of antioxidant levels in the selected halophytes. L. shawii showed the highest concentration of peroxidase, and P. undulata had the lowest. However, polyphenol oxidase activity was similar among the selected halophytes, ranging from 30-38 µg fresh wt/h.

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Proline

Proline estimation demonstrated the highest level in L. shawii (46.66 mg/100g dry weight) and the lowest in T. nilotica (25.49 mg/100g dry weight) [Table 7].

Total carbohydrates and soluble sugars

Carbohydrate estimation showed the highest content in T. nilotica (31.5%), while the L. shawii had the lowest (16.8%). Furthermore, soluble sugar analysis showed the highest value in T. nilotica (640.48 mg/100g dry wt), whereas L. shawii showed the lowest (278.55 mg/100g dry wt) [Table 7, Figure 8].

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Figure 8:

Estimation of total carbohydrates and protein in the selected halophytes. T. nilotica showed the highest estimation in both, whereas P. undulata had the lowest protein content, and L. shawii had the lowest in total carbohydrates.

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Free amino acids (Protein)

Protein analysis showed the lowest protein content in P. undulata (13.06%), while T. nilotica demonstrated the highest (18.95%) [Table 7, Figure 8].

Salinity significantly alters soil characteristics. Hence, elevated soil salt levels can lead to increased bulk density, soil compaction, compromised soil structure, clay dispersion, and the formation of surface crusts.^[35] Table 1 shows that sand content is high across all soils due to the desert environment, where sandy soils are more prevalent. Moreover, the prevalence of sand contributes to soil porosity and total pore volume, allowing well-structured soil to not impose restrictions on the growth of the tested plant roots, which aligns with the soil texture investigation by Ibraheem et al. (2021) in selected halophytes.^[36]

Db analysis showed the lowest in T. nilotica, which may be attributed to the soil texture under T. nilotica, which is fine sandy, and to higher total porosity. In addition, T. nilotica harbors a higher clay content, which also contributes to lower Db values. In contrast, the soil under L. shawii is moderately coarse sandy, thus exhibiting the highest Db and Dp values at both depths [Table 2]. Furthermore, P% analysis showed the highest value in T. nilotica soil at both depths, likely due to its soil texture and high OM and clay content. In contrast, L. shawii soil had the lowest P% at both depths, likely due to its low levels of both OM and clay content [Table 2]. According to Abd El-Wahab and Al-Salameen (2015), soil’s porosity increases with a higher OM%, while it decreases with increasing soil depth; hence, at the subsurface level, due to the increased pressure from the upper layers, leading to particle compaction and reduced interstitial spaces.^[37] Our OM determination showed the highest value in T. nilotica soil at both depths [Table 5], which is attributed to its clay and silt content. It is worth mentioning that the soil’s OM content decreased as the depth increased, reflecting the presence of biological processes in the surface layer, hence making it higher in OM. Overall, the selected halophytes showed low levels of OM (below the reference range of 0.86%),^[38] which may be due to rapid decomposition of OM in the soil caused by high temperatures, reduced rainfall, and increased soil aeration.^[39] Similar findings by Ibraheem et al. (2021) were reported, in which the soil in most areas exhibited extremely low levels of organic matter, falling below the detectable limit in the selected halophytes.^[36]

HC investigation revealed high values ranging from 23.33-26.33 cm h^-1 at the soil surface and 23.67-27 cm h^-1 at the subsurface [Table 2]. These values are generally elevated, indicating very rapid water movement, which is likely due to the overall sandy texture of the studied soils, which commonly have macropores between particles with little resistance to water movement, as well as fast drainage and low water retention. Moreover, T. nilotica had the lowest HC at 15-30 cm, which may be due to its fine, sandy soil and its high silt/clay ratio. Similar findings were reported by Bocuti et al. (2020), who found that decreased HC is affected by the high silt-clay ratio and fine particles.^[40]

pH was the same for all selected halophytes and was slightly alkaline (7.8), which is attributed to the high soluble salts in saline soils [Table 3]. Similar results were reported by Ibraheem et al. (2022), who reported that the pH of the halophyte soil ranged from 7-8.^[41]

Cation analysis showed Na⁺ with the highest concentration, followed by K⁺. On the other hand, anion analysis showed that Cl- had the highest concentration, and CO₂+HCO₃ had the lowest [Table 3]. Similar findings were reported by Basyoni and Aref (2016), where Na⁺ was the highest among cations and Cl^- among anions in Sabkhat Jizan.^[42] Furthermore, plant exposure to salinity is reported to lead to a significant rise in Na⁺ and Cl^- levels in the soil, resulting in the induction of osmotic stress.^[43] Moreover, EC analysis indicated variations from one depth to another for the same plant. These variations can be attributed to rainfall, which leaches excess salt from the soil and precipitates it in lower layers. Additionally, there are overall differences in EC among the soils of the studied plants. For instance, T. nilotica exhibited the highest at the soil’s subsurface, likely due to its proximity to saline water accumulation (the lake), resulting in higher soil salinity as well as its fine sandy-textured soil. Furthermore, it may also be attributed to its high silt and clay content, which reflects a higher retention of salts in the soil. On the other hand, P. undulata showed the lowest EC at both depths, as it is farther from the saline water accumulation [Table 3]. Additionally, the soils of P. undulata and L. shawii are moderately coarse sandy, which results in easier leaching of salt by rainfall compared to fine sand.^[44] Overall, all soil analyses of CaCO₃ were within the normal range (<8%) due to the soil’s slight alkalinity (pH 7.8).^[45] The water sample is classified as fourth class, with the highest Na⁺ content (SAR mean of 99.8) [Table 4]. Hence, it is unsuitable for irrigation for all crops.^[46]

CEC estimation showed the highest level in T. nilotica, which may be attributed to its high clay and OM content. Hence, the smaller the soil particles, the larger the specific surface area, and consequently, the CEC increases.^[47] According to Audah and Shamsham (2008), soils with high clay and OM content generally exhibit higher CEC than sandy or low-OM soils. Furthermore, CEC is influenced by various factors, including soil texture, the quantity and type of clay minerals, OM, pH, and the soil’s chemical composition. Therefore, CEC is considered an estimate of soil fertility, since exchangeable cations serve as the primary source of essential nutrients for plant growth. In addition, the GC in the selected soils ranged from 0.36-0.78%, within the expected range for sandy soils [Table 5]. Gypsum is widely distributed in arid and semi-arid regions. Gypsum crystals partially dissolve in water, with an estimated solubility of 2.6 g L^-1. Thus, determining the soil content of gypsum helps identify and characterize soil properties.^[45]

ESP analysis showed the highest values in T. nilotica and L. shawii, due to their higher Na⁺ concentrations than other cations [Table 5], as indicated by the soil chemical analysis. It’s worth mentioning that there are noticeable differences in ESP between the depths of the soil for L. shawii. This difference is attributed to leaching of the soil surface after rainfall, leading to salt deposition in lower soil layers. Moreover, leaching of high-salt content is a good indicator that allows seeds to germinate, roots to grow, and facilitates nutrient availability for the plant.^[48]

L. shawii showed the largest leaf area compared to the other plants, with an average of 2.34 cm²/plant [Table 6, Figure 5]. Despite being subjected to considerable abiotic stress, the soil salinity beneath is estimated at EC 9.8 mΩ (6313.33 ppm) [Table 3]. In contrast, T. nilotica has the smallest leaf area, averaging 0.66 cm²/plant, which can be attributed to its severe salinity stress, as the salinity level in its subsurface soil is extremely high (17 mΩ = 10900 ppm), causing the plant to reduce transpiration and water loss by minimizing leaf area [Table 6, Figure 5]. Furthermore, salt stress limits cell elongation in the growing tissue, leading to decreased leaf area and reduced dry matter assimilation in the plant. Moreover, prolonged exposure to salinity reduces photosynthesis rates, diminishes nutrient storage, and decreases growth hormone production.^[43] Additionally, salt glands, such as those found in young leaves of T. nilotica, are also present in most halophytes,^[10] serving for ion isolation due to the presence of small vacuoles in these leaves.^[49]

Photosynthesis pigment estimation showed that P. undulata had the highest Chl A and Chl B concentration, as well as total Chl, carotene, and the Chl/carotene ratio, whereas L. shawii showed the lowest in all. Maybe due to its less exposure to stress [Table 6, Figure 6]. Additionally, the high carotenoid content in P. undulata may have aided the protection of photosynthetic pigments by quenching singlet oxygen (¹O₂) and peroxyl radicals generated by excess excitation of Chl under intense light and high temperatures.^[50,51] Carotenoids serve a crucial function as non-enzymatic antioxidants, protecting the photosynthetic system. Ghanem et al. (2021) demonstrated notable increases in carotenoid levels with elevated NaCl concentrations in selected halophytes. This upsurge in carotenoid concentration could be a strategic response to preserve Chl levels, preventing their decrease under varying salinity concentrations. Generally, saline and arid conditions significantly reduce pigment synthesis, thereby lowering the net photosynthetic rate. It is worth mentioning that P. undulata exhibited the lowest Chl A/B (2.45). At the same time, T. nilotica recorded the highest value of (2.94) [Table 6, Figure 6], which is attributed to the fact that under saline stress conditions, the plant tends to develop an increase in this ratio, which, in turn, leads to a reduction in the photosynthetic process.^[18] which further confirms that P. undulata experiences less stress than other plants. Furthermore, Pérez-Labrada et al. (2019) reported that under saline stress, the plant exhibits an elevated Chl A/B ratio compared to non-stress conditions, leading to an imbalance that hinders photosynthesis.^[52] In addition, photosynthesis is among the most vulnerable physiological processes to ROS induced by salinity. This susceptibility is attributed to the fact that the photosynthetic photosystem reaction centers (PSI and PSII) are the primary sites for ROS generation within chloroplasts.^[53,50] Therefore, the presence of salt stress has the potential to diminish the Chl content in leaves, consequently impacting the efficiency of photosynthesis. Zhang et al. (2022) have reported similar observations: a decline in Chl A, B, and overall Chl content resulting from increased salt concentration.^[54]

Notably, T. nilotica showed low Chl content, although it had the highest carbohydrate estimation. Maybe because carbohydrates can be generated independently of Chl via several metabolic processes, such as fat oxidation. L. shawii had the lowest Chl parameters, except for the Chl A/B ratio (which is a stress indicator), which may be attributed to its high soil salinity, which inhibits leaf photosynthetic rate and increases CO₂ concentration inside cells, thus reducing carbohydrate synthesis [Figure 8]. Additionally, TSS estimation showed the highest in T. nilotica and the lowest in L. shawii. Soluble sugars are vital in the osmotic stress response, helping plants cope with high salinity, increase water uptake, and provide nutrients for growth.^[1] Similar findings were reported by Guo et al. (2020), who found that soluble sugar levels increased in leaves and stems after treatment with 200 mM NaCl.^[55] In addition, high levels of NaCl were found to inhibit starch accumulation, thereby impacting photosynthesis. Moreover, stress-induced responses in halophytes generally involve: 1) the buildup of essential compounds (such as soluble sugars and amino acids) to resist osmotic stress and maintain cellular osmotic potential and metabolism; 2) secondary metabolites, serving as potential antioxidants and regulatory substances.^[56]

Proline estimation showed the lowest in T. nilotica and the highest in L. shawii [Table 7]. Proline is reported to serve as a nitrogen source for plants during recovery phases and as a promoter of antioxidant enzyme activities. Hence, the high estimated proline level in L. shawii may be attributed to the high peroxidase enzyme activity. Additionally, proline levels are mediated by Ca; therefore, they may contribute to the Ca levels at the soil surface reported in the previous chemical analysis section, as well as to peroxidase.^[57,58] Furthermore, plants accumulate osmotic modulators, such as proline, to maintain osmotic pressure under salt stress.^[59,60] Moreover, Ghanem et al. (2021) have reported an increase in proline synthesis in halophytes at high salinity levels, which may be considered as a coping mechanism to compensate for the reduction in carotene and flavonoids, hence aiding the free radical scavenging activity.^[18] Furthermore, protein analysis showed the highest content in T. nilotica, whereas P. undulata had the lowest [Table 7, Figure 8]. There is an inverse correlation between protein and proline content since proline is generated from protein breakdown. Therefore, we observed that T. nilotica had the highest protein content and the lowest proline content.^[61]

Lastly, excessive ROS generation in halophytes results from salinity stress, leading to imbalances in redox reactions.^[43] In addition, ROS production induces carbohydrate oxidation and pigment breakdown.^[36] Therefore, we found increased levels of the antioxidant enzymes peroxidase and polyphenol oxidase to cope with elevated ROS levels [Table 7, Figure 7]. Similar results were reported by Ghanem et al. (2021), who found that peroxidase levels were high in halophytes, helping decrease free radical concentrations.^[18]