Integrated ecological, anatomical, phytochemical and genetic characterization of Peganum harmala L. along a soil salinity gradient in the Taif Region, Saudi Arabia

Peganum harmala L. is a medicinal shrub that tolerates harsh, dry conditions, but the links between its habitat, structure, chemistry, and genetics are still not well understood. In this study, we investigated five geo‑referenced natural populations from the Taif region in Saudi Arabia, using an integrated ecological, anatomical, phytochemical, and molecular approach. All soils samples were slightly alkaline (pH 7.61–7.79), and fell within the non-saline to slightly saline range based on electrical conductivity (EC) classifications, but they differed in salinity. Sites P1 (Al Ahlam Park) and P2 (C36) had lower EC values, while P3 (Ashayra), P4 (As Sayl al Kabir), and P5 (Al Sir) showed relatively higher EC values within this overall low salinity range. In all locations, P. harmala L. formed well-established, low, spreading to dense dome-shaped shrubs on sandy–stony substrates. Multivariate analysis of soil variables (pH, EC, CaCO₃, major anions and cations) set P3 apart from the other sites, and both clustering and principal component analysis (PCA) grouped P1, P2, P4, and P5 together, with P3 forming its own distinct cluster. Anatomically, most populations had stem cross sections with well-organized vascular cylinders. By contrast, stems from P3 displayed an irregular outline, a compact multilayered cortex, and scattered vascular strands with visibly reduced secondary xylem. Total alkaloid content ranged from 3.208 to 4.106%, with the highest values at P1 and P2 and the lowest at P3. Molecular analyses using SCoT markers (10 primers; 121 bands, 54 polymorphic) and IRAP markers (10 primers; 130 bands, 76 polymorphic) revealed moderate to high nuclear genetic diversity (expected heterozygosity up to 0.50; Shannon’s index up to 0.69). Overall genetic similarity among populations was high, but their structure was non-random: populations 2, 4, and 5 consistently clustered together, whereas populations 1 and especially 3 were more divergent. Chloroplast barcodes (matK: PX089557; rpoC1: PV942406) from a representative voucher from Population 1 were consistent with its identification as P. harmala L. and clearly separated this sequence from those of related species in phylogenetic trees. In combination with morphological identification, this supports assignment of all sampled populations to P. harmala L. Among the studied populations, P3 emerges as an ecologically and genetically distinct unit within P. harmala L. in the Taif region, characterized by its unique nuclear genetic profile, modified stem anatomy, and reduced total alkaloid content associated with higher soil salinity and altered major-ion composition. This integrative framework—combining soil characterization, quantitative anatomy, total alkaloids, SCoT/IRAP markers, and plastid barcodes illustrate how fine-scale edaphic heterogeneity is associated with anatomical, chemical, and genetic variation in this key desert medicinal shrub.