Optimal Nitrogen and Phosphorus Supply Drives Synergistic Effects of Arbuscular Mycorrhizal Fungi and Rhizobia on Alfalfa (Medicago sativa L.) Growth

Abstract The dual symbiotic relationship between leguminous plants and arbuscular mycorrhizal fungi (AMF) and rhizobia (Rh) can enhance plant yield and stress resistance, but the efficiency of this symbiosis under different nitrogen (N) and phosphorus (P) supply conditions remains unclear. This study examined how N and P availability affect alfalfa (Medicago sativa L.) growth, symbiotic efficiency, nutrient accumulation, and enzyme activities under AMF (Rhizophagus clarus HR1) and Rh (Ensifer medicae) inoculation, using a three-factor randomized factorial design. The results showed that co-inoculation consistently enhanced plant performance compared with single inoculation and controls, with the best results under low N and low or moderate P. Obviously, Low P promoted AMF colonization and high N reduced nodulation. And co-inoculation increased nodule traits and activities of key enzymes, supporting a C–N–P cycle. This synergy peaked under 3 mM N and 0.05–0.5 mM P, but was inhibited by high N or P. This study provides a theoretical basis for precise regulation of legume-AMF-Rh tripartite symbiosis and practical guidance for microbial fertilizer application in sustainable agriculture.