Abstract <p>Biochar and cover crops have been extensively studied individually for their potential to improve soil health. However, the synergistic effects of co-applying these two sustainable practices remain relatively unexplored. This study aimed to investigate the impact of co-applying biochar and cover crops on soil microbial communities and enzyme activity. A Japanese apricot orchard was used as an experimental site, cultivated in a Fluvisol. Four treatments were established: control, biochar only, cover crop only, and biochar and cover crop co-application. Two types of biochar (rice husk and bamboo) and two cover crops (<i>Paspalum notatum</i> and <i>Trifolium repens</i>) were used. The soil bacterial community was analyzed using the 16s gene, and enzyme activities were measured. The co-application of biochar and cover crops significantly altered the soil microbial community compared to the control treatment. Specifically, the co-application treatments enriched complex compound-degrading bacteria, indicating improved soil health. The control treatment, on the other hand, exhibited potential pathogen infestation and limited soil pore structure. While enzyme activities did not show significant changes over the two-year study period, the observed shifts in the microbial community suggest a positive long-term impact on soil health. The increased abundance of beneficial microorganisms is likely to enhance nutrient cycling and improve soil resilience. The synergistic effects of co-applying biochar and cover crops were evident in the enhanced soil microbial diversity and abundance. Bacterial diversity varied significantly among treatments, with co-applications showing higher diversity than single applications, particularly in BbBg and BbWc. The combination of both cover crops (BbBgWc) resulted in the highest diversity. Linear Discriminant Analysis identified key bacterial families associated with specific treatments, highlighting distinct community compositions. Although enzyme activities remained stable, the results highlight the positive impact of the studied practices on soil microbial ecology. Further research is needed to elucidate the long-term implications for soil health and crop productivity.</p>