In order to study the response changes of soil microbial community structure after the application of triazole fungicide difenoconazole（dif），the physical and chemical properties of soil after 35 days were monitored by simulating the application dose in production，and the changes of soil bacterial and fungal community composition were analyzed by high-throughput sequencing technology. The relationship between environmental factors and microbial community response was established by db-RDA analysis. The results showed that with the increase of dif application concentration，organic matter（SOM），total nitrogen（TN）and alkali-hydrolyzed nitrogen（AN）content in soil were increased，while available phosphorus（AP）content showed a decreasing trend，and bacterial Chao1，ACE and Shannon indices in Alpha diversity index were lower than those in untreated soil. The Shannon index of fungi was higher than that of untreated soil. A total of 31 phyla，91 classes，206 orders，315 families and 553 genera of bacteria were detected in all samples，and 11 phyla， 34 classes，79 orders，174 families and 370 genera of fungi were detected. When dif was applied at a concentration of 50 mg/kg，the relative abundances of Proteobacteria and Bacteroidetes in soil bacteria were increased by 14.2% and 1.81%， respectively，the relative abundances of Acidobacteria and chlorocurvula were decreased by 8.28% and 3.44%，respectively， and the relative abundances of Arthrobacter，the absolute dominant bacterium，was decreased by 7.85%. Gibberellus and Mortierella gradually became the dominant group of fungi. Among the dominant bacterial groups，Proteobacteria showed significant positive correlation with TN content，ascomycetes showed significant positive correlation with AN，TN content and dif application concentration，and mortierella showed significant negative correlation with SOM content. TN and AN content had great influence on the Shannon index of bacteria. The above results indicated that the application of high concentration dif had effects on soil physicochemical properties，microbial richness and community structure.