A dataset of 87 020 nifH reads and 16 782 unique nifH protein sequences obtained over 2 years from four locations across a gradient of agricultural soil types in Argentina were analysed to provide a detailed and comprehensive picture of the diversity, abundance and responses of the N2 -fixing community in relation to differences in soil chemistry and agricultural practices. Phylogenetic analysis revealed an expected high proportion of Alphaproteobacteria, Betaproteobacteria and Deltaproteobacteria, mainly relatives to Bradyrhizobium and Methylosinus/Methylocystis, but a surprising paucity of Gammaproteobacteria. Analysis of variance and stepwise regression modelling suggested location and treatment-specific influences of soil type on diazotrophic community composition and organic carbon concentrations on nifH diversity. nifH gene abundance, determined by quantitative real-time polymerase chain reaction, was higher in agricultural soils than in non-agricultural soils, and was influenced by soil chemistry under intensive crop rotation but not under monoculture. At some locations, sustainable increased crop yields might be possible through the management of soil chemistry to improve the abundance and diversity of N2 -fixing bacteria.

In this survey, a total of 80 787 reads and 28 171 unique NifH protein sequences were retrieved from soil RNA. This dataset extends our knowledge about the structure and diversity of the functional diazotrophic communities in agricultural soils from Argentinean Pampas. Operational taxonomic unit (OTU)‐based analyses showed that nifH phylotypes related to Geobacter and Anaeromyxobacter (44.8%), Rhizobiales (29%), Cyanobacteria (16.7%), and Verrucomicrobiales (8%) are key microbial components of N2 fixation in soils associated with no‐till management and soil depth. In addition, quantification of nifH gene copies related to Geobacter and Cyanobacteria revealed that these groups are abundant in soils under maize–soybean rotation and soybean monoculture, respectively. The correlation of physicochemical soil parameters with the diazotrophic diversity and composition showed that soil stability and organic carbon might contribute to the functional signatures of particular nifH phylotypes in fields under no‐till management. Because crop production relies on soil‐borne microorganism's activities, such as free N2 fixation, the information provided by our study on the diazotrophic population dynamics, associated with the edaphic properties and land‐use practices, represents a major contribution to gain insight into soil biology, in which functionally active components are identified.