Biocontrol agents applied after fumigation play an important role to the soil microenvironment. We studied the effect of Trichoderma applied after dimethyl disulfide (DMDS) plus chloropicrin (PIC) fumigation on the cucumber growth, soil physicochemical properties, enzyme activity, taxonomic diversity, and yield through laboratory and field experiments. The results confirmed that Trichoderma applied after fumigation significantly improved soil physicochemical properties, cucumber growth, soil-borne pathogens, and soil enzyme activity. Genetic analysis indicated that Trichoderma applied after fumigation significantly increased the relative abundance of Pseudomonas, Humicola and Chaetomium, and significantly decreased the relative abundance of the pathogens Fusarium spp. and Gibberella spp., which may help to control pathogens and enhanced the ecological functions of the soil. Moreover, Trichoderma applied after fumigation obviously improved cucumber yield (up to 35.6%), and increased relative efficacy of soil-borne pathogens (up to 99%) and root-knot nematodes (up to 96%). Especially, we found that Trichoderma applied after fumigation increased the relative abundance of some beneficial microorganisms (such as Sodiomyces and Rhizophlyctis) that can optimize soil microbiome. It is worth noting that with the decline in the impact of the fumigant, these beneficial microorganisms still maintain a higher abundance when the cucumber plants were uprooted. Importantly, we found one tested biocontrol agent Trichoderma 267 identified and stored in our laboratory not only improved cucumber growth, reduced soil-borne diseases in late cucumber growth stages but also optimized micro-ecological environment which may have good application prospect and help to keep environmental healthy and sustainable development.