Influence of sulphate and other anions on leaf apoplastic pH and stomatal closure in Vicia faba under salinity stress

Salinity-induced stress perceived at plant roots elicits complex systemically orchestrated physiological responses in plants, with ionic imbalances and osmotic pressures playing pivotal roles in modulating stomatal regulation. This study investigates how different anions (Cl^–, NO^3–, SO₄^2–, and PO₄^3–) influence root and leaf apoplastic pH (pH_apo) and stomatal closure in Vicia faba under saline conditions. Using a microscopy-based real-time pH_apo imaging approach, we demonstrate that transient alkalinisation in the leaf apoplast is anion-specific and varies with leaf maturity, with Cl^– and NO^3– eliciting stronger pH_apo increases than SO₄^2– and PO₄^3–. Notably, the root apoplast pH remained unaffected by short-term salinity, highlighting that the alkalinisation is localised to the shoot and likely linked to xylem-translocated ionic effects. Findings reveal that stomatal aperture reductions occurred across treatments regardless of the magnitude of apoplastic alkalinisation. Among the anions studied, SO₄^2– displayed unique regulatory potential, suggesting a role in stomatal behaviour. These results provide new insights into the interplay between ionic stress, apoplastic pH regulation, and stomatal responses, offering potential targets for improving salinity tolerance in crops.