In order to assess the changes in soil physio-chemistry, the abundances and community structures of the N-cycling microorganisms, as well as total bacteria, after repeated application of dicyandiamide (DCD) and 3,4-dimethylpyrazolphosphate (DMPP), samples of soil were obtained from a field experiment where DCD and DMPP had been applied along with fertilizer annually for over 13 years. A randomized complete block design was used that included four treatments: no fertilization (CK), fertilizer application (NPK), NPK + DCD, and NPK + DMPP; each treatment with three replicates. Results showed that repeated chemical fertilizer application led to lower soil pH, significantly higher potential nitrification rates (PNRs) and increased population sizes of ammonia oxidizers compared with the control. However, soil pH was unaffected by the application DMPP or DCD over 13 years. Application of DMPP decreased the PNR and the abundance of ammonia-oxidizing bacteria (AOB) and nirK genes, compared to NPK, but DCD addition exhibited inhibitory effects only on PNR. There were no inhibitor- or fertilizer-induced changes in the abundances of nirS, nosZ and 16S rRNA genes. In addition, the results demonstrated that the community compositions of total bacteria and ammonia oxidizers were not altered by N addition and use of inhibitors at the phylum level. The DCD and DMPP inhibitors, therefore, appear to be effective mitigation technologies targeting specific microorganisms with no long-term adverse impact on the soil ecosystem.