Abrupt drought-flood alternation is a frequent meteorological disaster that occurs during the summer in southern China. This study sought to clarify the composition of the bacterial community in the rhizosphere soil of rice under nitrogen (N) application after abrupt drought-flood alternation and to provide more basic data for understanding N absorption by the roots system under N application after abrupt drought-flood alternation. The activity of N metabolism enzymes and the N content and N accumulation in various organs improved under N application after abrupt drought-flood alternation. Between the T1_N and T1 comparison groups, 1758 differentially expressed genes were identified, including 1027 upregulated and 731 downregulated genes. Transcriptomic GO and KEGG analyses indicated that nitrate assimilation, response to bacterium, defence response to bacterium, and N compound metabolic processes were enriched. At the phylum level, Proteobacteria, Acidobacteria, Nitrospirota, and Gemmatimonadota were the dominant flora. The root glutamine synthetase activity was significantly negatively correlated with Nitrospirota. Nitrospirota is a key taxon regulating rice root N absorption under N application after abrupt drought-flood alternation. N application after abrupt drought-flood alternation promoted the rapid recovery ability of plants and improved the microbial environment in rice. This study preliminarily clarified the role of microorganisms in the root uptake of rice under N application after abrupt drought-flood alternation. • This study was subjected to RNA-seq and 16S rRNA gene technology. • Abrupt drought-flood alternation changed rice rhizosphere soil microbial environment. • The nitrospirota of T1_N is the dominant flora at the phylum level. • Response to bacterium, nitrogen compound metabolic processes were enriched between T1_N and T1.