Agricultural systems that receive high amounts of inorganic nitrogen (N) fertilizer in the form of either ammonium (NH4þ), nitrate (NO3 ) or a combination thereof are expected to differ in soil N transformation rates and fates of NH4þ and NO3 . Using 15N tracer techniques this study examines how crop plants and soil microbes vary in their ability to take up and compete for fertilizer N on a short time scale (hours to days). Single plants of barley (Hordeum vulgare L. cv. Morex) were grown on two agricultural soils in microcosms which received either NH4þ, NO3 or NH4NO3. Within each fertilizer treatment traces of 15NH4þ and 15NO3 were added separately. During 8 days of fertilization the fate of fertilizer 15N into plants, microbial biomass and inorganic soil N pools as well as changes in gross N transformation rates were investigated. One week after fertilization 45e80% of initially applied 15N was recovered in crop plants compared to only 1e10% in soil microbes, proving that plants were the strongest competitors for fertilizer N. In terms of N uptake soil microbes out-competed plants only during the first 4 h of N application independent of soil and fertilizer N form. Within one day microbial N uptake declined substantially, probably due to carbon limitation. In both soils, plants and soil microbes took up more NO3 than NH4þ independent of initially applied N form. Surprisingly, no inhibitory effect of NH4þ on the uptake and assimilation of nitrate in both, plants and microbes,was observed, probably because fast nitrification rates led to a swift depletion of the ammonium pool. Compared to plant and microbial NH4þ uptake rates, gross nitrification rates were 3e75-fold higher, indicating that nitrifiers were the strongest competitors for NH4þ in both soils. The rapid conversion of NH4þ to NO3 and preferential use of NO3 by soil microbes suggest that in agricultural systems with high inorganic N fertilizer inputs the soil microbial community could adapt to high concentrations of NO3 and shift towards enhanced reliance on NO3 for their N supply.