Tillage practices and fertilizer application play important roles in nitrogen transformation and utilization and grain yield. The objective of this study was to determine the effects of urea types and tillage modes on soil nitrate and ammonium N content, urease activity, abundance of denitrifying bacteria and nitrite bacteria, and grain yield and nitrogen use efficiency in a maize (Zea mays L.) field. The treatments included two urea types (conventional urea and polycoated urea) applied at a rate of 225 kg N/hm² as a basal fertilizer; additionally, zero N treatments were set as the control under subsoiling and rotary tillage modes. Throughout the experiments in 2011 and 2012, the results showed that both the numbers of nitrite bacteria and denitrifying bacteria increased with the application of urea but then later would decrease to even fewer than in the zero N treatment. Compared with conventional urea, polycoated urea could stabilize soil urease activity and increase upland soil nitrite bacteria numbers but decrease the abundance of denitrifying bacteria. Also, polycoated urea was beneficial for accelerating nitrogen transformation into nitrate and reducing nitrogen losses from denitrification while keeping soil ammonium and nitrate N concentrations in the 0-40 cm layers at a high level for a long time, especially in the middle and later periods of maize growing season. Polycoated urea generally increased grain yield and agronomic efficiency of applied N by 7.25%-10.82% and 56.37%-84.54%, respectively, compared to those with conventional urea regardless of tillage practices. Rotary and subsoiling tillage could temporarily stimulate soil nitrate N concentration. Compared with rotary tillage, subsoiling tillage increased the numbers of denitrifying bacteria. Moreover, the interaction of polycoated urea and subsoiling tillage could not only increase the urease activity but also the abundance of nitrite bacteria. It would keep the nitrate N concentration at a high level in the later maize growing stage. However, nitrogen loss through denitrification and leaching in the early stage of the maize growing season may increase. Polycoated urea in rotary soils could significantly increase soil nitrate nitrogen concentration while reducing the number of denitrifying bacteria within a period of time. The application of polycoated urea in subsoil produced higher grain yield, with increases of approximately 1.41% and 10.62%, compared to those in rotary soil, in 2011 and 2012, respectively. So in a drought year, the effect of subsoiling tillage combined with polycoated urea may be more significant in increasing grain yield. In conclusion, subsoiling tillage coupled with polycoated urea increases urease activity and nitrous bacteria abundance, and stimulates nitrogen transformation, grain yield, and nitrogen use efficiency.