In order to elucidate the importance of biogeochemical interactions between NH4+ and aquifer media in groundwater runoff paths, a dynamic monitoring section in the riverbank zone, which is most sensitive to environmental characteristics and perpendicular to the flow direction of the Songhua River in northeastern China, was selected for field experiments in this study. The results indicated that the NH4+ concentration decreased gradually along the groundwater runoff path under exploitation conditions. The NH4+ concentrations of J1, J2, and J3 decreased by 8%, 18%, and 22%, respectively, as compared to the starting concentration of 1.3 mg/L. Adsorption of NH4+ by aquifer media at different depths is a monolayer adsorption process in accordance with pseudo-second-order kinetic equation. The maximum reduction of NH4+ from the aquifer media from top to bottom was 76%, 67%, 56%, and 42%, respectively. The function and activity of dominant functional bacteria have characteristics of coevolution with the NH4+ transformation process. The main genera in the fluctuation zone are Pseudomonas (8.83%) and Acinetobacter (4.37%), which mainly transform NH4+ by heterotrophic nitrification–aerobic denitrification (HN–AD). The main genera in the saturated zone are Flavobacterium (32.60%) and Sphingobium (3.54%), which mainly transform NH4+ by anaerobic denitrification. The spatial variations of species and abundance for NH4+ transformation functional bacteria decrease by 2.74% and 3.47%, respectively, along groundwater runoff paths. In the vertical and horizontal directions of groundwater runoff, the percentage of adsorption in NH4+ transformation gradually decreased and the percentage of biotransformation gradually increased. The adsorption processes in the O2/NO3− reduction, Fe/Mn reduction, and SO42− reduction zones were 20.7%, 3.6%, and 1.0%, respectively. The corresponding proportions of the biotransformation process were 79.3%, 96.4%, and 99.0%. This research is critical for elucidating the bio-geochemical interaction between NH4+ and aquifer media along the course of groundwater runoff in order to offer a scientific basis for the prevention and management of groundwater nitrogen pollution.