[Objective] Allylthiourea (ATU) is a promising nitrification inhibitor. However, its effects on soil nitrification and greenhouse gas (GHG) mitigation are still unclear.[Method] In this study, a 21-day microcosm incubation was established with the application of nitrogen (N) and different doses of ATU (1%, 5%, 10%, 15% and 20% of N applied) to a yellow-brown upland soil. Also, dicyandiamide (DCD, 10% of N applied) was applied to compare the inhibition efficiency on nitrification and GHG emission with ATU. The dynamics of inorganic nitrogen and N₂O/CO₂ emission during the incubation were detected, and changes in the different microbial population were analyzed by real-time PCR and 16S rRNA gene-based high through-put sequencing.[Result] N application greatly stimulated soil nitrification activity and promoted N₂O emission. DCD had a strong inhibitory effect on soil nitrification (68.6%) and N₂O emission (93.3%). ATU did not influence soil nitrification at low doses (<5%), but inhibited the nitrate accumulation at high doses (>10%) with the highest inhibition efficiency of 14.7%. All treatments with ATU decreased N₂O emission by 60.3%~68.2% but was still much higher than when DCD was applied. In general, the global warming potential (GWP) of N₂O and CO₂ were the highest in N treatment and seconded by ATU+N treatment. There were no significant differences in GWP between DCD+N and CK treatment, or among different doses of ATU with N treatments. The quantitative real-time PCR of amoA genes suggested that ammonia-oxidizing bacteria (AOB) rather than ammonia-oxidizing archaea (AOA) and complete ammonia-oxidizing bacteria (Comammox), had a positive relationship with soil nitrate accumulation and N₂O emission，but a negative correlation with pH. Microbial community analysis by high through-put sequencing revealed Nitrosovibrio tenuis-like AOB dominated in soil nitrification process which was greatly stimulated by nitrogen. Besides, ATU and DCD significantly increased the relative abundance of cupriavidus but reduced the relative abundance of Patulibacter, Aeromicrobium, Actinomycetospora, Defluviicoccus and Acidipila.[Conclusion] This study reveals the exact mechanisms of ATU on soil microbial guilds and GHG emission and plays an important role in the future implementation of agricultural management strategies and the evaluation of global climate change.