Nitrogen (N) input from fertilizers modifies the properties of agricultural soils as well as bacterial community diversity, composition and relationships. This can lead to negative impacts such as the deterioration of system multifunctionality, whose maintenance is critical to normal nutrient cycling. Synthetic nitrification inhibitors (NIs) can be combined with fertilizers to improve the efficiency of N use by reducing N losses. However, analysis of their effects on non-target bacteria are scarce. This study aimed to analyze the effect of applying the NIs DMPP and DMPSA on the whole bacterial community. Through 16S rRNA amplicon sequencing we determined the differences between samples in terms of microbial diversity, composition and co-occurrence networks. The application of DMPP and DMPSA exerted little impact on the abundance of the dominant phyla. Nevertheless, several significant shifts were detected in bacterial diversity, co-occurrence networks, and the abundance of particular taxa, where soil water content played a key role. For instance, the application of NIs intensified the negative impact of N fertilization on bacterial diversity under high water-filled pore spaces (WFPS) (>64%), reducing community diversity, whereas alpha-diversity was not affected at low WFPS (<55%). Interestingly, despite NIs are known to inhibit ammonia monooxygenase (AMO) enzyme, both NIs almost exclusively inhibited Nitrosomonas genera among AMO holding nitrifiers. Thus, Nitrosomonas showed abundance reductions of up to 47% (DMPP) and 66% (DMPSA). Nonetheless, non-target bacterial abundances also shifted with NI application. Notably, DMPSA application partially alleviated the negative effect of fertilization on soil multifunctionality. A remarkable increase in populations related to system multifunctionality, such as Armatimonadetes (up to +21%), Cyanobacteria (up to +30%) and Fibrobacteres (up to +25%) was observed when DMPSA was applied. NI application substantially influenced microbial associations by decreasing the complexity of co-occurrence networks, decreasing the total edges and node connectivity, and increasing path distances.

Agricultural sustainability is compromised by nitrogen (N) losses caused by soil microbial activity. Nitrous oxide (N2O) is a potent greenhouse gas (GHG) produced as consequence of nitrification and denitrification processes in soils. Nitrification inhibitors (NI) as 3,4-dimethylpyrazole-succinic acid (DMPSA) are useful tools to reduce these N losses from fertilization. The objective of this work was to test the efficiency of DMPSA in two different tillage management systems, conventional tillage (CT) and no-tillage (NT), in a winter wheat crop under Humid Mediterranean conditions. N fertilizer was applied as ammonium sulphate (AS) with or without DMPSA in a single or split application, including an unfertilized treatment. GHG fluxes (N2O, CO2 and CH4) were measured by the closed chamber method. amoA and nosZI genes were quantified by qPCR as indicators of nitrifying and denitrifying populations. Nitrification was inhibited by DMPSA in both CT and NT, while the higher water filled pore space (WFPS) in NT promoted a better efficiency of DMPSA in this system. This higher efficiency might be due to a greater N2O reduction to N2 as result of the nosZI gene induction. Consequently, DMPSA was able to reduce N2O emissions down to the unfertilized levels in NT. Provided that NT reduced CO2 emissions and maintained crop yield compared to CT, the application DMPSA under NT management is a promising strategy to increase agro-systems sustainability under Humid Mediterranean conditions.

Agriculture has increased the release of reactive nitrogen to the environment due to crops' low nitrogen-use efficiency (NUE) after the application of nitrogen-fertilisers. Practices like the use of stabilized-fertilisers with nitrification inhibitors such as DMPP (3,4-dimethylpyrazole phosphate) have been adopted to reduce nitrogen losses. Otherwise, cover crops can be used in crop-rotation-strategies to reduce soil nitrogen pollution and benefit the following culture. Sorghum (Sorghum bicolor) could be a good candidate as it is drought tolerant and its culture can reduce nitrogen losses derived from nitrification because it exudates biological nitrification inhibitors (BNIs). This work aimed to evaluate the effect of fallow-wheat and sorghum cover crop-wheat rotations on N₂O emissions and the grain yield of winter wheat crop. In addition, the suitability of DMPP addition was also analyzed. The use of sorghum as a cover crop might not be a suitable option to mitigate nitrogen losses in the subsequent crop. Although sorghum-wheat rotation was able to reduce 22% the abundance of amoA, it presented an increment of 77% in cumulative N₂O emissions compared to fallow-wheat rotation, which was probably related to a greater abundance of heterotrophic-denitrification genes. On the other hand, the application of DMPP avoided the growth of ammonia-oxidizing bacteria and maintained the N₂O emissions at the levels of unfertilized-soils in both rotations. As a conclusion, the use of DMPP would be recommendable regardless of the rotation since it maintains NH₄⁺ in the soil for longer and mitigates the impact of the crop residues on nitrogen soil dynamics.