Enhancing drought tolerance in cumin through physiological and genetic insights from a synthetic variety

Drought stress exerts numerous adverse effects on plants, including the inhibition of photosynthesis and growth, as well as impacts on osmotic regulators and enzymatic activity. Additionally, severe drought can result in reduced crop yields and plant^1,2. Developing and cultivating drought-tolerant varieties is a strategic approach to mitigate the adverse effects of drought stress on plants. The synthetic cumin variety developed in this study showed improved drought tolerance and seed yield compared to the parental genotypes. These improvements are attributed to increased photosynthetic pigment retention, higher antioxidant enzyme activity, and greater osmoregulatory compound accumulation. These findings suggest that the synthetic variety represents an excellent genetic resource for agricultural production under arid and semi-arid conditions (Fig. 3).

In cumin, seed yield is the most critical and economically relevant trait. It is controlled by numerous physiological and morphological processes, environmental conditions, and the plant’s genetic structure¹⁹. Improved seed yield under both normal and drought conditions demonstrates the genetic advantage of the synthetic variety. Compared to parental genotypes, the synthetic variety achieved a yield increase of up to 71.59% under drought stress, supported by higher photosynthetic pigment retention and osmoregulatory efficiency observed in this study. This highlights the role of physiological traits as key factors in drought resilience.

The findings are consistent with earlier studies on medicinal plants and their drought tolerance. Drought stress significantly decreased seed yield by impacting various traits, aligning with the findings of Noori et al.²⁰, and Yadav and Yadav²¹in cumin. Factors contributing to yield reduction under drought stress include reduced carbon metabolism, stomatal conductance, and water uptake due to impaired root growth³⁰. Under optimal irrigation, increased photosynthesis and assimilate production enhance the seed filling rate, seed weight, and ultimately yield. Drought stress leads to reduced leaf water content, stomatal closure, and decreased photosynthesis, along with impaired enzymatic activities, resulting in lower seed yield. Additionally, drought stress causes flower drop and reduced seed weight, further diminishing seed yield¹. The observed increase in antioxidant enzyme activities (e.g., catalase, peroxidase, and ascorbate peroxidase) likely contributed to the synthetic variety’s ability to detoxify reactive oxygen species under drought conditions, protecting cellular structures and maintaining physiological processes. Additionally, higher proline and soluble sugar levels in the synthetic variety supported osmotic adjustment, improving water uptake and turgor maintenance.

One of the primary breeding goals for cumin is to increase seed yield, which can be achieved through the development of synthetic varieties. The synthetic cumin variety exhibited the highest seed yield under both stress and non-stress conditions (Table 2). Furthermore, this variety was less negatively affected by drought stress compared to the parental genotypes. The enhanced seed yield of the synthetic variety over the parental genotypes can be attributed to increased genetic diversity and recombination²². The combination of genes from the parental genotypes provides a broader genetic base and potential for new gene combinations in the synthetic variety, increasing yield potential, especially under adverse environmental conditions²³. This enhances the suitability of the synthetic variety for cultivation in arid and semi-arid regions. Significant seed yield increases through the development of synthetic varieties have also been reported by other researchers²³.

Photosynthetic pigment content is a crucial and straightforward physiological indicator for assessing drought stress impact and determining drought tolerance. Chlorophyll, located within the thylakoid membrane, is essential for photosynthesis, and drought stress triggers the rapid translocation of nitrogen and carbohydrates from vegetative organs to seeds, accelerating their maturation². According to Table 2, drought stress resulted in a significant reduction in photosynthetic pigment content. Similar reductions in cumin under drought conditions have been documented by Noori et al.²⁰, Timachi et al.²⁴, and Pishva et al.²⁵. Thus, photosynthetic pigment content can be effectively used alongside seed yield to screen for drought tolerance. In this study, the synthetic variety achieved the highest seed yield under both normal and drought stress conditions (Table 2). Additionally, the synthetic variety showed the least reduction in chlorophyll a and carotenoid content under drought stress, maintaining higher levels of photosynthetic pigments and carotenoids than the parental genotypes. This indicates that the synthetic variety sustains higher net photosynthesis and water content, showcasing greater drought tolerance compared to the parental genotypes. The reduced decline in photosynthetic pigment levels in the synthetic variety under drought stress optimizes the expression of stress-related genes, which plays a critical role in enhancing the plant’s resilience to drought. This presents a promising approach for improving agricultural sustainability in water-scarce environments.

The initial physiological response of plants to drought stress involves osmotic regulation and stomatal closure, which are crucial for maintaining cellular moisture and enhancing water uptake from the environment. This process helps preserve normal physiological and biochemical functions within cells²⁰. Osmotic regulation, by generating a negative osmotic potential within cells, enhances the plant’s ability to absorb water from the soil, thereby improving its drought tolerance. In our study, the levels of three osmotic regulators proline, sugar, and protein exhibited differential changes under drought stress (Table 2). Both proline and sugar levels increased, with the highest accumulation observed in the synthetic cumin variety, suggesting its superior drought tolerance.

Proline functions as an antioxidant, maintaining cellular redox balance, reducing lipid peroxidation, and inducing the expression of stress-responsive genes while activating antioxidant enzymes¹. Soluble sugars are vital for cellular biosynthesis and metabolism, serving as essential energy sources during developmental processes. Under drought conditions, soluble sugars aid in osmotic regulation, reduce water potential, and protect cells from oxidative damage by stabilizing cellular membranes and proteins¹.

Drought stress led to a reduction in soluble protein concentration in the leaves. Nevertheless, the synthetic variety exhibited higher protein levels under both normal and drought conditions. This reduction in protein concentration can be attributed to decreased protein synthesis or increased proteolysis due to heightened protease activity under drought conditions²⁶. Drought stress enhances protease activity, resulting in protein degradation and an increase in free amino acids, including proline. The activation of protease-encoding genes promotes protein degradation and the synthesis of compatible solutes²⁶.

Previous studies on cumin have similarly reported an accumulation of proline and sugar, along with a decrease in protein content under drought stress, aligning with our findings^24,25. Therefore, it can be concluded that the higher accumulation of proline and soluble sugars and the reduction in protein content in the synthetic variety, compared to the parental genotypes, significantly contribute to the increased drought tolerance of cumin. This adaptation facilitates improved physiological activity and reduced membrane lipid peroxidation, offering a promising strategy for enhancing agricultural sustainability in water-limited environments.

Drought stress led to an increase in the essential oil content of cumin (Table 2). This observation aligns with the findings of previous studies, which reported that drought stress enhances the essential oil percentage in cumin¹⁹. Medicinal plants tend to produce more essential oils under drought conditions as a defensive mechanism to prevent cellular oxidation¹⁹. Our study showed that the synthetic variety had the highest essential oil content under both normal and drought stress conditions, suggesting improved drought tolerance.

The chemical composition of essential oils is influenced by the plant’s genetic makeup and environmental conditions²⁷. Analysis of the chemical constituents in cumin essential oil indicated that the synthetic variety had higher levels of the primary active ingredient compared to the parental genotypes under both normal and drought stress conditions (Table 3). Although the changes in essential oil components due to drought stress did not follow a consistent trend, the combination of superior parental genotypes in the synthetic variety resulted in increased levels of most compounds, particularly under drought stress, compared to the parental genotypes.

This enhanced chemical profile under stress conditions indicates that the synthetic variety is more adept at producing valuable metabolites, which may contribute to its superior drought tolerance. This finding underscores the potential of using synthetic varieties to improve the resilience and productivity of crops in water-limited environments.

The antioxidant system in plants is composed of enzymatic and non-enzymatic antioxidants²⁸. Enzymatic antioxidants are pivotal in neutralizing free radicals and maintaining the balance of ROS within plant cells². In this study, drought stress enhanced the activities of antioxidant enzymes, including ascorbate peroxidase, peroxidase, and catalase (Table 2). The synthetic variety demonstrated the highest antioxidant activity under drought condition. Elevated antioxidant enzyme activity strengthens the plant’s physiological and biochemical defense mechanisms against drought, aiding in the maintenance of cellular water content and reducing drought-induced damage.

Under drought stress, the synthetic variety exhibited increases of 131.81%, 122.05%, and 264.78% in catalase, peroxidase, and ascorbate peroxidase activities, respectively, compared to the average of the parental genotypes (Table 2). Catalase, an iron-containing enzyme, is crucial for the detoxification of hydrogen peroxide²⁹. Enhanced catalase activity in response to drought stress has also been observed in plants by other researchers^1,2.

Peroxidase aids in eliminating ROS to prevent excessive plasma membrane damage, with malondialdehyde serving as an indicator of lipid peroxidation due to ROS damage. Peroxidase helps remove hydrogen peroxide and malondialdehyde, preserving cellular membrane integrity. In this study, drought stress resulted in a twofold increase in peroxidase activity. Similar increases in peroxidase activity due to drought stress have been observed in various other plants^1,2. Ascorbate peroxidase reduces oxidative stress damage by scavenging free radicals, particularly hydrogen peroxide. The primary detoxification system for hydrogen peroxide in plant chloroplasts involves the ascorbate-glutathione cycle, where ascorbate peroxidase is a key enzyme. It catalyzes the conversion of hydrogen peroxide to water using ascorbate as an electron donor²⁸. Overexpression of ascorbate peroxidase in various plant species underscores its critical role in defending plants against abiotic stress^1,2. Thus, the increased antioxidant enzyme levels in the synthetic cumin variety suggest enhanced tolerance to oxidative stress and better adaptability to environmental challenges.

Recent studies on drought-tolerant cereal crops have emphasized the critical role of enzymatic antioxidant systems in improving drought resilience. For instance, Reza et al.¹ reported that improved catalase and peroxidase activities in wheat (Triticum boeoticum) significantly reduced ROS damage under drought conditions, leading to enhanced yield stability. Similarly, Bhardwaj et al.² demonstrated that ion homeostasis and mesophyll regulation in barley play vital roles in maintaining cellular integrity and photosynthetic activity during water stress. These findings align with our results, where catalase, peroxidase, and ascorbate peroxidase activities increased significantly in the synthetic cumin variety under drought conditions. The universal relevance of these mechanisms across plant species underscores their importance in developing drought-tolerant varieties.

A comprehensive understanding of genetic diversity is crucial for breeding programs aimed at exploiting heterosis and increasing crop resilience. ISSR primers revealed substantial genetic diversity between the synthetic cumin variety and the parental genotypes, confirming the findings of Ebrahimiyan et al.³⁰ on the high genetic diversity of Iranian cumin genotypes. Primers ISSR_06 and ISSR_05, which demonstrated high diversity according to PIC, EMR, and MI indices, are particularly useful for analyzing the subsequent generations of the synthetic variety (Table 4).

Cluster analysis of physiological and molecular data revealed that the parental genotypes and the synthetic variety were grouped into two distinct clusters (Fig. 1). The cluster analysis results were consistent with the PCoA diagram, which illustrated the distribution of the parental genotypes and the synthetic variety (Fig. 2). The greater genetic distance observed during the random crossing of parental genotypes in the polycross test for producing the synthetic variety increased the number of heterozygous loci and the potential for heterosis. Moreover, genetic distance might be associated with desirable traits³¹. Therefore, it can be concluded that the parental genotypes were aptly selected, facilitating the transfer of superior genes to the synthetic variety.

The findings of this study demonstrate that the synthetic cumin variety exhibits significantly enhanced seed yield and drought tolerance compared to parental genotypes. This improvement is attributed to a combination of physiological adaptations, such as increased antioxidant enzyme activity and osmoregulatory compound accumulation, as well as the broader genetic base achieved through synthetic variety breeding. These traits are not only pivotal for the resilience of cumin but can also serve as a model for improving other medicinal and aromatic crops cultivated in arid and semi-arid regions.

The breeding strategy employed here could be applied to other cross-pollinated crops where hybridization is less feasible, providing a promising approach for improving seed yield and stress tolerance. Additionally, the integration of physiological, biochemical, and molecular markers demonstrated in this study can be adopted in breeding programs targeting climate-resilient crops.

Beyond cumin, these findings hold relevance for addressing global food security challenges, particularly in regions experiencing increasing water scarcity due to climate change. Developing drought-tolerant varieties of crops with low water requirements will contribute to sustainable agricultural practices and ensure the productivity of economically valuable plants under adverse conditions.

While this study provides a comprehensive evaluation of the synthetic cumin variety under drought stress, further investigations are warranted to fully exploit its potential. Future research should focus on unraveling the molecular pathways and gene networks that confer drought tolerance, which could lead to the identification of novel genetic markers for marker-assisted selection. Additionally, multi-location and multi-year trials are essential to confirm the stability and adaptability of the synthetic variety across diverse agro-ecological zones. Moreover, investigating the effects of other abiotic stresses, such as salinity or extreme temperatures, on the synthetic variety could expand its utility and relevance in stress-prone regions, ensuring broader applications in sustainable agriculture.