Arbuscular mycorrhizal fungi and exogenous Ca2+ application synergistically enhance salt and alkali resistance in perennial ryegrass through diverse adaptive strategies

The challenge of soil salinization and alkalization, with its significant impact on crop productivity, has raised growing concerns with global population growth and enhanced environmental degradation. Although arbuscular mycorrhizal fungi (AMF) and calcium ions (Ca²⁺) are known to enhance plant resistance to stress, their combined effects on perennial ryegrass’ tolerance to salt and alkali stress and the underlying mechanisms remain poorly understood. This study aimed to elucidate the roles of Arbuscular mycorrhizal (AM) fungus Rhizophagus irregularis and exogenous Ca²⁺ application in molecular and physiological responses to salt-alkali stress. AM symbiosis and exogenous Ca²⁺ application enhanced antioxidant enzyme activity and non-enzymatic components, promoting reactive oxygen species (ROS) scavenging and reducing lipid peroxidation while alleviating oxidative damage induced by salt-alkali stress. Furthermore, they enhanced osmotic balance by increasing soluble sugar content (Proportion of contribution of the osmotic adjustment were 34∼38 % in shoots and 30∼37 % in roots) under salt stress and organic acid content (Proportion of contribution of the osmotic adjustment were 32∼36 % in shoots and 37∼42 % in roots) under alkali stress. Changes in organic solute and inorganic cation-anion contents contributed to ion balance, while hormonal regulation played a role in these protective mechanisms. Moreover, the protective mechanisms involved activation of Ca²⁺-mediated signaling pathways, regulation of salt-alkali stress-related genes (including LpNHX1 and LpSOS1), increased ATPase activity, elevated ATP levels, enhanced Na⁺ extrusion, improved K⁺ absorption capacity, and a reduced Na⁺/K⁺ ratio, all contributing to the protection of photosynthetic pigments and the enhancement of photosynthetic efficiency. Ultimately, the combined application of exogenous Ca²⁺ and AMF synergistically alleviated the inhibitory effects of salt-alkali stress on perennial ryegrass growth. This finding suggested that exogenous Ca²⁺ may participate in the colonization of perennial ryegrass plants by R. irregularis, while AM symbiosis may activate Ca²⁺ pathways. Consequently, the combined treatment of AM and Ca²⁺ is beneficial for enhancing plant regulatory mechanisms and increasing crop yield under salt-alkali stress.