Impact of short-interval, repeat application of dicyandiamide on soil N transformation in urine patches

In pastoral farming systems, bovine urine patches are ‘hot-spots’ of elevated soil nitrogen (N). Nitrification in urine patches is linked to deleterious environmental outcomes, such as formation of the greenhouse gas nitrous oxide (N2O). The nitrification inhibitor dicyandiamide (DCD) reduces the rate of nitrification in soils subsequently limiting N-losses. Dicyandiamide's bacteriostatic mode of inhibiting ammonia oxidising bacteria (AOB), has raised concerns about the efficacy of frequent DCD use. For example, frequent use could result in selection for a DCD-tolerant AOB community. Furthermore, the impacts of DCD on other aspects of microbial N transformation in soil are largely unknown. To test the influence of short-term repeat application of DCD on soil N cycling, we established a replicated field-trial in which ±urine and ±DCD were added to pasture soils (fully crossed, 2×2 factorial design). After 57 d, treatments were re-applied. Mineral N pools, pH, DCD concentration, moisture and temperature were measured, along with N2O fluxes. Microbial communities involved in ammonia oxidation (bacteria and archaea), nitrite reduction, and N2O reduction were quantified using real-time PCR targeting functional genes (amoA, nirS, and nosZ). Overall, the addition of DCD significantly (P<0.05) reduced both the formation of nitrate (∼64%), and N2O loss from urine-treated soils (∼44%). The effect of repeating the urine and DCD applications produced similar results with no decline in DCD efficacy. This was despite a doubling in the size of the AOB community, stimulated by the initial urine application. The application of urine+DCD had a significant but minor impact on denitrifying bacteria (nirS), rather the population size of these bacteria correlated with increasing soil temperature over time (rho=0.7; P=0.002). In contrast, copies of nosZ increased with urine addition (P<0.05) but were unaffected by sampling date (increasing soil temperature). Our results demonstrate that an initial DCD application does not affect its efficacy to inhibit nitrification and reduce N2O emissions following a subsequent DCD application 57 days later.