This study investigated the effects of combining bio-organic fertilizer with chemical fertilizers on wheat yield and soil microbial communities in different regions，aiming to provide a scientific basis for promoting the application of bio-organic fertilizer in Hebei province and enhancing wheat production capacity.Field experiments were conducted in Suning county (SN) and Ningjin county (NJ) of Hebei province to compare and analyze the effects of two treatments，namely chemical fertilizer alone (CK) and bio-organic fertilizer combined with chemical fertilizer (BOF)，on soil physicochemical properties，enzyme activities，and soil microbial communities.The results showed that compared with CK，the BOF treatment significantly increased wheat yield by 11.3% and 7.4% in SN and NJ，significantly increased root surface area by 75.8% and 22.4%，and increased aboveground fresh weight by 81.6% and 29.3%，respectively，demonstrating stronger bio-organic fertilizer effects in SN.Concurrently，BOF application improved soil nutrient contents across both sites.in SN，total nitrogen，total phosphorus，available phosphorus，and available potassium increased by 4.8%，8.7%，139.3%，and 27.1%，respectively，whereas in NJ，the increases were 13.6%，8.9%，47.8%，and 7.8%.Microbial community analysis showed that the effect of BOF treatment on the bacterial community in SN soil was significantly greater than that in NJ，while the effect on the fungal community was not significant.Notably，BOF treatment significantly increased the abundance of beneficial bacteria Sphingomonas and Massilia in SN soil，while reducing the abundance of pathogenic fungi such as Alternaria and Fusarium.Functional prediction revealed that the BOF treatment significantly enhanced the functional activity of ureolysis in SN soil and nitrate reduction in NJ soil.Correlation analysis showed that compared to NJ，the soil microbial communities and functions in SN had a stronger correlation with soil properties，and the increase in beneficial bacteria abundance was closely linked to improvements in soil nutrient indicators.The results demonstrated that bio-organic fertilizer (BOF) application could consistently enhance wheat yield and soil fertility while optimizing microbial community composition and structure across regions with varying initial soil conditions，with more pronounced effects observed in areas with higher baseline soil fertility levels.