Biochar and arbuscular mycorrhizal fungi (AMF) can promote plant growth, improve soil properties, and maintain microbial activity. The effects of biochar and AMF on plant growth, root morphological traits, physiological properties, and soil enzymatic activities were studied in spinach (Spinacia oleracea L.). A pot experiment was conducted to evaluate the effect of biochar and AMF on the growth of spinach. Four treatments, a T1 control (soil without biochar), T2 biochar alone, T3 AMF alone, and T4 biochar and AMF together, were arranged in a randomized complete block design with five replications. The biochar alone had a positive effect on the growth of spinach, root morphological traits, physiological properties, and soil enzymatic activities. It significantly increased the plant growth parameters, such as the shoot length, leaf number, leaf length, leaf width, shoot fresh weight, and shoot dry weight. The root morphological traits, plant physiological attributes, and soil enzymatic activities were significantly enhanced with the biochar alone compared with the control. However, the combination of biochar and AMF had a greater impact on the increase in plant growth, root morphological traits, physiological properties, and soil enzymatic activities compared with the other treatments. The results suggested that the combined biochar and AMF led to the highest levels of spinach plant growth, microbial biomass, and soil enzymatic activity.