Abstract: A field experiment was carried out to study the effects on alfalfa yield, soil microbial characteristics and soil enzyme activities, of partially replacing phosphate fertilizers with microbial fertilizers, and comprehensive evaluation were conducted using grey relational analysis. The aim of the research was to explore the effect of the biological fertilizer and to scientifically determine the optimum optimal application rate for alfalfa. The experiment had seven treatments: P₁₀₀, 100% phosphate fertilizer (80 kg/667 m²); M₁₀₀, phosphate solubilizing bacteria+nitrogen-fixing bacteria (0.5 kg/667 m²); P₇₅M₂₅,75% P+25% M; P₅₀M₅₀, 50% P+50% M; P₂₅M₇₅, 25% P+75% M; R₁₀₀, 100% nitrogen-fixing bacteria; CK (no fertilizer). P₁₀₀ increased the alfalfa yield significantly at the first cut, while the treatment P₇₅M₂₅ had produced the highest yield (P<0.05) at the second cut. Comparing the total yield from two cuts with CK, P₅₀M₅₀ was highest yielding and increased the alfalfa yield by 60.45%, while P₁₀₀ had the second highest yield, which increased about 58.40% than CK. Partially replacing phosphorus fertilizers with microbial fertilizers increased soil microbial population. P₅₀M₅₀ significantly increased the number of bacteria, fungi and actinomyces in the 0-10 cm soil layer at the first cutting by 204.6%, 174.5% and 33.2%, respectively, compared to CK. P₅₀M₅₀ and P₂₅M₇₅ improved soil enzyme activity significantly in the 10-20 cm soil layer at the first cut. Grey correlation analysis indicated that sucrase had the highest grey correlation value (0.842), followed by yield (0.814), while microbial biomass of phosphorus and carbon had lower grey correlation value (0.512 and 0.513), respectively. The treatment P₅₀M₅₀ demonstrated soil fertility improvement while reducing the application rate of chemical fertilizers, and was therefore agronomically beneficial.