LAUSR

Abstract Biological nitrogen (N) fixation is one of the most significant parts of the N cycle in terrestrial ecosystems, and this process is carried out by soil diazotrophs. However, knowledge of diazotroph assembly processes and activity in response to diverse fertilization strategies in agroecosystems across a large geographical scale is still lacking. Here, we selected nine agro-ecological experimental sites that covered a wide geographical distance (~3500 km) at a continental scale, and investigated the diazotrophic communities, assembly processes, distance-decay patterns and N2 fixation activity in response to geographical factors and long-term fertilization strategies. The results showed that the dominant genera were Bradyrhizobium (~30.5%) and Azospirillum (~26.8%) in all samples, and RDA analysis showed that the relative abundance of Bradyrhizobium was positive correlated with MAP while specific negatively correlated with soil pH and the relative abundance of Azospirillum. Geographical factors (location and climate) and fertilization collectively drive diazotroph assembly processes and determine diazotroph activity. Diazotroph assembly processes were influenced by both stochastic (~36.2%) and deterministic (~63.8%) processes simultaneously at large geographical scales and under various fertilization strategies. Moreover, fertilization increased the proportion of deterministic processes under various fertilization strategies. The N fixation rate was determined by local soil properties. Fertilization changes but does not always suppress nitrogen fixation activity. Both geographical factors and fertilization through the shift of diazotroph community composition and the changes in soil properties, indirectly affected the assembly process and N fixation rate. Among soil properties, pH was the dominant factor and linearly related to diazotrophs assembly process, while N fixation rate reached peak at near-neutral pH. These results elucidate the mechanism of soil diazotroph assembly process and activity shaped by both geographical factors and fertilization; thus, expand the current understanding of the diazotroph community affected by fertilization strategies across a large geographical scale.