The Mediterranean climate is characterized by a dry period, varying in length depending on the latitude. Plants develop numerous morphological adaptations to cope with these periods of water scarcity [1]. The genus Pistacia is part of this vegetation, which includes xeric species with significant ecological and economic potential.

The species of the genus Pistacia exhibit a highly complex taxonomy. Therefore, the taxonomic relationships among its different species remain a subject of controversy and are not yet well-defined. The number of recognized species varies among authors, ranging from 9 to 11 species [2–6].

The Atlas Pistachio tree (Pistacia atlantica Desf.) is a fascinating deciduous tree that holds a significant place in the Algerian steppe and Saharan ecosystems. Its ecological range stretches from the heart of the Sahara to the margins of humid bioclimates [7]. It adapts to diverse climatic conditions, and its contribution to Algerian ecosystem biodiversity is invaluable. However, this plant heritage is endangered, with its degradation resulting from several factors such as habitat loss, climate change, overexploitation, and diseases that threaten the survival of this species protected by Algerian law [8] (Fig. 1). Therefore, a comprehensive understanding of this species is essential to establish and develop effective and sustainable conservation strategies. Hence, studying intraspecific and intersex variability in the species is valuable for explaining and predicting population adaptation to climate change and understanding mechanisms for conserving and restoring endangered species. However, research focusing on inter-population variability and between-sex differences in Pistacia species is considered negligible compared to interspecific variability. Notable works in this area include studies by [9–14].

In this context, given the complexity of taxonomic affinities within the genus Pistacia and their behaviors in response to understudied environmental heterogeneity, the present study contributes to the investigation of diversity and variability among 7 ecotypes of P. atlantica sampled from various bioclimates. The study aims to explore intra-specific micro and macromorphological leaf variability and even inter-sex variability to understand how this species adapts to extreme conditions in its environment (aridity and altitude). This is achieved by examining morphological and ultrastructural leaf traits, particularly using scanning electron microscopy (SEM) for leaf epidermal characteristics (density, size, distribution, and position of stomata, type and density of trichomes, etc.), and potentially providing criteria for its systematic classification. Our study will also enhance our understanding of Pistacia species, which are largely understudied and therefore underutilized in afforestation and desertification control programs such as the “Green Dam,” despite their ecological and economic significance.

Plant responses to drought vary depending on the adaptive mechanisms expressed by the plants, as they have adapted by implementing stress-adaptive strategies favoring drought tolerance and/or survival [17–20]. Most of these characteristics manifest in leaves as the primary organ for regulating water loss through transpiration.

At the aerial level, reducing water loss can be achieved through the regulation of stomatal opening or leaf surface area [18,21]. Thus, modifying the number of stomata [22,23] and/or reducing stomatal conductance [24–26] allows for the control of water fluxes. Additionally, reducing leaf size decreases the plant’s evaporative surface area [27]. Some species or populations develop leaf adaptations to protect leaf surfaces from intense solar radiation, creating a moister environment around stomata that slows water loss. These adaptations include wax accumulation on leaf cuticles [28], dense trichomes as seen in Olea europaea [29], or leaf rolling as observed in Oryza sativa [30]. Analyzing leaf morphology, especially epidermal features, has thus been necessary to better understand plant adaptive responses to fluctuating environmental conditions.

This study aimed to enhance our understanding of Pistacia species, particularly Pistacia atlantica, which remains poorly defined in terms of taxonomy and ecology. We focused on macro-and micro-morphological leaf traits to explore potential criteria for Pistacia species’ taxonomy and understand the observed intraspecific and inter-sexual heterogeneity and the adaptive processes they reveal among different P. atlantica populations located in various bioclimatic regions in Algeria. Unfortunately, intraspecific variability is often considered negligible compared to interspecific variability across broad species pools. However, numerous studies highlight the importance of considering intraspecific variability in functional traits to improve our understanding of plant community structure and distribution [31–36], which can be highly significant among populations of the same species along ecological gradients [37]. Similar studies regarding inter-population variability in Pistacia species have been reported for populations of different origins studied by various authors, in P. atlantica [38–41], and in P. lentiscus [11,13].

Regarding quantitative leaf macro-morphological traits, a strong correlation was observed between leaf and terminal leaflet biometrics for all P. atlantica populations. These results align with those of [42] in a study on morphological diversity among three Pistacia species in Turkey and [43] for P. atlantica. Indeed, they note strong correlations between leaf and leaflet dimension variables. Our results also reveal a positive correlation between leaf and terminal leaflet dimensions with the aridity index of collection sites; however, they are negatively correlated with altitude. Reference [44] similarly noted strong correlations between leaf dimensions, terminal leaflet length, and altitude, while the number of leaflets is negatively correlated with altitude.

In the present study, remarkable variability in quantitative leaf traits was observed among different ecotypes of P. atlantica. This could be due to individual sexes or environmental parameters (aridity and altitude). Indeed, in P. atlantica, leaves are longer at Oz (11.16 cm) (IA: 0.35; Altitude 779 m) and Bg (10.97 cm) (IA: 0.32; Alt. 659 m), and shorter at Aflou (Tb) (8.72 cm) (IA: 0.15; Alt. 1427 m) and Guetaya (Gt) (9.82 cm) (IA: 0.18; Alt. 1380 m). Therefore, the reduction in leaf size could be explained by the effect of aridity and altitude on plants, leading to a truly adaptive limitation of transpiring surfaces (leaves being the first organs exposed to water deficit) [45]. Altitude also impacts plant leaf surface, with climate changes as it rises. The air becomes colder and drier, affecting plant life accordingly. Many plants have adapted strategies to survive at high altitudes by reducing leaf surface to minimize water loss through transpiration and maintain a stable internal temperature. Our results align with [14], indicating that the smallest P. atlantica leaves were reported at higher stations rather than at the most arid ones.

We also found a significant difference between male and female individuals for the majority of quantitative leaf traits. Indeed, leaf and terminal leaflet dimensions are higher in female trees than in males for all P. atlantica populations.

However, no significant difference between the sexes in P. atlantica was observed for the number of leaflets and petiole length. This sex-related variability was also ob-served within P. atlantica populations. This sex-related variability was demonstrated in our study on the effect of individual sex on the morphological diversity of P. lentiscus leaves in a previous work [9]. This aligns with [46] results, who noted that leaf and terminal leaflet dimensions of female trees are slightly higher than those of males in P. atlantica. However, opposite trends are reported by [47] for P. lentiscus. These differences between male and female trees may be due to the energy needs of reproduction, as during fruit formation season, females require more energy than males [48]. According to [49], females develop adaptive strategies by keeping older leaves longer, as seen in P. lentiscus, or by increasing leaf surface area as in Siparuna grandiflora [50].

Regarding the macromorphological qualitative leaf traits, our study shows significant intraspecific heterogeneity for most studied variables. Leaf wings are a crucial character in identifying species of the Pistacia genus [2]. In this study, we observed that nearly half of P. atlantica leaves have green wings (narrow) along the rachis to the petiole (55.12%), and the other half only on the rachis (44.88%), with variable rates among populations (narrowly winged rachis and petiole) 62.45% (Tb), 63.63% (Oz), 79.33% (Dr), and 79.74 (Bg). However, the lowest percentage (19%) was found at station (Bt) with a lower aridity index (0.06). Our results for this variable are consistent with those of [10,39,40,43] for P. atlantica. According to [51], trees known as P. terebinthus in California could be a variety of P. atlantica, due to their high vigor and presence of a winged rachis.

Regarding leaf color, it ranges from green to dark green in P. atlantica, as confirmed by previous studies [14,42,52]. Depending on the origins, Oz (61.51%) and Bg (80.38%) with aridity indices (0.35, 0.32) and altitudes (779, 659 m) close to each other, have darker-colored leaves compared to other stations, while green color dominates in Djelfa Dr (76%), Gt (60%), and Bt (71.66%). Light green color was observed at very low rates. In terms of sex, the percentage of green color is nearly identical between male and female individuals, while the percentage of dark green color is higher in female leaves. The literature [14,42,52] reports a color range from green to dark green for this species, consistent with our results. Reference [53] demonstrated that under optimal conditions, the photosynthetic capacity of female Pistacia lentiscus is similar, if not higher, than that of males. However, under stress conditions, their photosynthetic capacity decreases compared to males. Therefore, Reference [54] noted that a high content of essential oils is present in female plants.

The majority of populations have petioles with a rounded and flattened shape on the adaxial surface (92.49%). Flat (5.89%) and round (1.61%) shapes were also observed. We found that the (Bt) origin had the lowest rate (79.66%) compared to other P. atlantica populations. These results are similar to those obtained by [41] in P. atlantica. Contrary to the flattened form reported by other authors [2,42,52,55], no difference was recorded between males and females for this species.

In our study, the size of the terminal leaflet compared to basal leaflets shows significant intraspecific variability. Indeed, its size relative to basal leaflets is either smaller, the same size, or larger. According to [2,42,52], the terminal leaflet is larger than basal leaflets in P. atlantica. However, Reference [43] found variable results for the same species, which align with our findings. As for its shape, it remains quite variable both between sexes and depending on the origin. Various shapes were observed, including narrow elliptical and lanceolate. These results are consistent with those recorded by [43] and [44], who observed a dominance of the lanceolate and narrow elliptical shapes in P. atlantica.

The apex of the leaf remains an important character for distinguishing species within the genus Pistacia [2]. In P. atlantica, it varies from acute, obtuse, mucronulate to Emarginate form (observed only in the Oz station). Eight forms of leaf apex in P. atlantica were observed in the study by [44]; for our study, six apex forms were recorded, among which (obtuse, acute, mucronulate, and acuminate) have been previously described [14,42,56]. The other forms (emarginate, retuse) have not been observed before, which constitutes new information for this species.

On the micromorphological level, all populations of P. atlantica present hairs with a remarkable density at the leaf margin (a line of microscopic cilia curved toward the apex); on the midrib, they are less dense on both leaf surfaces. We noticed a slight difference between males and females, with females having a higher adaxial hairiness. Additionally, observation using SEM allowed us to distinguish the presence of glandular capitate hairs across the leaf surface, as well as non-glandular trichomes of the ciliate type. Systematically speaking, the morphology and distribution of trichomes can provide important clues in the classification of species within the genus Pistacia [2,40,43,57]. According to [38], trichome distribution differs among varieties in P. atlantica, for example, they are present on both surfaces at the main vein level for the atlantica variety and on the entire lower surface for the cabulica variety, while they are absent in the kurdica variety [42,57]. According to [57], leaflets have a ciliate margin in the mutica variety and rarely ciliate in the kurdica variety.

Trichome density is also a characteristic influenced by ecological conditions. Altitude and minimum temperatures can play an important role in their distribution and density on leaves [43]. In arid environments, xeromorphic plants often have leaves covered with trichomes and wax. In this regard, [58] indicate that trichomes and waxes reduce the sedimentation rate of aphids and thus discourage herbivores in the presence of attractive food sources. Some hairs may also play a role in water absorption in semi-desert habitats [59–61]. This confirms our results regarding the presence of trichomes with very high intensity on the leaves of P. atlantica, which plays an important role in Algerian steppe and Saharan ecosystems.

Observation using SEM of the leaves allowed us to observe wax deposits of varied structure from one species to another [12]. In P. atlantica, a fine structure with flake deposits is observed (granular film with a powdery structure). In many xeromorphic plants, wax particles cover stomata to prevent water loss due to high radiation [62].

Stomata can also be an important character in species identification and taxonomy [63]. Our results showed that P. atlantica leaves are hypostomatic. They are more abundant on the abaxial surface; however, on the adaxial surface, we recorded rare stomata along the main and secondary veins. Reference [64] suggested that hypostomy is an evolutionary trait of Pistacia species and is considered a strongly xeromorphic trait.

Our results show a significant difference among the 7 populations studied for stomatal density on both surfaces, with the Oz (IA = 0.35), Bg (IA = 0.32), and Tb (Alt. 1427 m) stations having the highest abaxial stomatal density (408.83 st/mm²), (391 st/mm²), and (90.5 st/mm²) on the adaxial surface. On the other hand, the lowest abaxial and adaxial densities were observed at Msi (IA = 0.12) and Bt (IA = 0.06) with 311.06 and 332.16 st/mm² for the abaxial surface and 43.70 and 70 st/mm² for the adaxial surface, respectively. Our results also show that high-altitude stations have a high stomatal density on both surfaces for this species. Our data are in line with those of [65], who stated that increased stomatal density and reduced stomatal size correspond to better adaptation to water economy. Reference [64] suggested that this variation could be related to the ecological plasticity of Pistacia species across a wide range of environmental conditions. Indeed, our samples were collected from different sites with different climatic conditions (aridity and altitude). This demonstrates remarkable plasticity of this species, enabling it to survive in a very wide geographic distribution range. Similarly, significant variation was observed among the 10 provenances of P. lentiscus regarding this variable [13].

Regarding stomatal size on the abaxial surface, intra-specific and inter-sex variability was recorded. Indeed, the values of stomatal length and width on the abaxial surface vary from one population to another, with the Oz population (IA = 0.35) having the smallest length value and Tb (Alt. 1427 m and IA = 0.15) having the largest. Reference [66] indicated that adaptation to drought involves a decrease in stomatal size.

In this study, a significant difference between males and females was recorded for abaxial and adaxial stomatal size and density. We noticed that stomatal size is slightly higher in males, while females have the highest densities.

Studying the presence of Pistacia atlantica under different bioclimates in Algeria has led us to investigate its behavior and adaptive processes developed by this species in response to environmental constraints, particularly altitude, and aridity, through various macromorphological and micromorphological leaf traits.

This study has allowed us to highlight new characteristics in the studied species, such as the occurrence of waxes and glandular hairs on leaflets, sunken stomata in the epidermis, and stomatal shape. These criteria provide an important taxonomic tool for identifying species within the Pistacia genus. Additionally, highlighting certain xerophytic characteristics is an important asset considering the potential use of these species in new plantations to combat desertification, given their ability to withstand particularly harsh water-related conditions. Therefore, their rehabilitation and conservation are necessary to contribute to the sustainable development of arid areas.

Our study reveals a very significant divergence in the studied traits among different ecotypes of Pistacia atlantica. Genetic factors undoubtedly play a crucial role in this variability, but we have observed that environmental factors also have a remarkable impact on the heterogeneity of most measured leaf traits. Gender also plays a crucial role in this variability. We can summarize the main characteristics revealed for this species as follows:

- Leaf dimensions: 10.1 cm × 6.95 cm; 10.01 cm × 6.89 cm ♂; 10.18 cm × 7.01 cm ♀;

- Higher number of leaflets (9.04); imparipinnate leaves (83.87%);

- Terminal leaflet size: 3.66 cm × 1.09 cm and length/width ratio: 3.49;

- Petiole length: 2.24 cm; rounded and flattened on the upper surface;

- Leaf wings on the rachis only (44.88%), up to the petiole (55.12%);

- Entire leaf margin;

- Leaf color: green (53%) (53.70% ♂; 52.20% ♀), dark green (41.66%) (38.88% ♂; 44.81% ♀); narrow elliptical shape (48.89%) (51.99% ♂; 43.65% ♀) and lanceolate (32.26%) (28.5% ♂; 36.52% ♀);

- Terminal leaflet apex: acute shape (32.72%) (52.33% ♂; 50.77% ♀), obtuse (32.42%) (19.72% ♂; 21.63% ♀), or mucronulate (10.60%);

- Leaf pilosity concentrated at the margin and midrib of the adaxial surface, less concentrated on the abaxial and adaxial surfaces;

- Presence of 02 types of trichomes: glandular and non-glandular (ciliate type);

- Wax deposits with a finer structure;

- Hypostomatic leaves, with rare stomata on the adaxial surface located along the main and secondary veins;

- Abaxial stomatal density: (368.91 st/mm²); (356.72 st/mm² ♂ and 400.38 st/mm² ♀);

- Adaxial stomatal density: (74.74 st/mm²); (74.77 st/mm² ♂ and 85.05 st/mm² ♀);

- Stomatal size: (24.58 µm × 15.48 µm); length/width ratio (1.6); elliptical shape; slightly sunken in the epidermis.