Abstract: Soil salinization is a global problem constraining agricultural and forestry development. Utilizing micro-bial symbiosis to enhance the salt tolerance of woody plants is a sustainable and effective strategy. We conducted a pot experiment to investigate the regulatory mechanisms of single and combined inoculation with Funneliformis mosseae (Fm) and Piriformospora indica (Pi) on two-month-old ‘Red Kernel Walnut’ (Juglans regia) seedlings under salt stress. There were five treatments, including 1) non-stress control (CK), 2) salt stress (0.8% NaCl, S), 3) salt stress + Fm inoculation (S+Fm), 4) salt stress + Pi inoculation (S+Pi), and 5) salt stress + combined Fm and Pi inoculation (S+FmPi). We measured the growth parameters, chlorophyll content, antioxidant enzyme activities, osmotic regulatory substances, and endogenous hormone levels at 10, 20, and 30 days after the initiation of treatments. The results showed that the inhibitory effect of salt stress on plants intensified over time. Inoculation treatments effectively alleviated stress damage at all time points, with combined inoculation (FmPi) demonstrating superior efficacy compared to single inoculations. At 10, 20, and 30 days after treatment, compared to the salt stress group (S), the S+FmPi treatment increased seedling biomass by 6.6%, 18.4%, and 24.2%, respectively; leaf chlorophyll a content by 43.3%, 84.9%, and 56.5%, chlorophyll b content by 19.6%, 107.6%, and 98.6%; root superoxide dismutase activity by 40.6%, 10.8%, and 9.7%, and ascorbate peroxidase activity by 44.7%, 57.3%, and 22.6%; while decreased root malondialdehyde content by 26.0%, 28.3%, and 28.9%. Hormonally, compared with the salt stress group (S), combined inoculation (S+FmPi) resulted in 1.0% decrease, 4.0% increase, and 3.3% reduction in leaf indole-3-acetic acid content at 10 d, 20 d, and 30 d, respectively. Moreover, abscisic acid content was decreased by 15.9% at 10 d, increased by 2.0% at 20 d, and decreased by 7.9% at 30 d. Comprehensive evaluation using principal component analysis and membership function values ranked the alleviating effects of inoculation treatments as S+FmPi >S+Fm>S+Pi. Combined inoculation of Fm and Pi significantly enhanced salt tolerance of walnut seedlings through synergistic multi-pathway regulation. These findings would provide a theoretical foundation for applying mycorrhizal technology in walnut cultivation on saline soils.

Key words: Juglans regia ‘Robert Livermore’, salt stress, mycorrhizal fungi, physiological and biochemical cha-racteristic