Plasma-activated water (PAW) is a recently developed cutting-edge technology that is increasingly gaining interest for its applications in medicine, food industry and agriculture. In plant biology, PAW has been shown to promote seed germination, plant growth, and plant resistance to biotic and abiotic stresses. Despite increasing knowledge of the beneficial effects exerted by PAW on plants, little information is currently available about how this emerging technology may affect the mutualistic plant-microbe interactions in the rhizosphere. In this work we have investigated the impact of irrigation with PAW, generated by a plasma torch, on arbuscular mycorrhizal (AM) symbiosis between the model legumeLotus japonicusand the AM fungusRhizophagus irregularis. Since PAW sensing by plants has recently been demonstrated to occur through calcium-mediated signalling, we monitored early cellular responses to different doses of PAW inL. japonicusroots expressing the Ca2+-sensitive photoprotein aequorin targeted to either the cytosol or nucleus. Quantitative analyses of AM fungal accommodation in host roots along with phosphate accumulation in leaves, as well as chemical analysis of N, C, S in shoots, showed that treatments with PAW play a modulatory role on plant AM symbiotic performance, in a manner dependent on the time interval of water exposure to the plasma and on the duration of plant irrigation treatment with PAW. Establishing a solid scientific ground for plasma-related technology may provide key elements to develop tools and treatments aimed to increase crop plant yield in a sustainable manner.