Integrated 16S and metabolomics revealed the mechanism of drought resistance and nitrogen uptake in rice at the heading stage under different nitrogen levels.

The normal methods of agricultural production worldwide have been strongly affected by the frequent occurrence of drought. Rice rhizosphere microorganisms have been significantly affected by drought stress. To provide a hypothetical basis for improving the drought resistance and N utilization efficiency of rice, the study adopted a barrel planting method at the heading stage, treating rice with no drought or drought stress and three different nitrogen (N) levels. Untargeted metabolomics and 16S rRNA gene sequencing technology were used to study the changes in microorganisms in roots and the differential metabolites (DMs) in rhizosphere soil. The results showed that under the same N application rate, the dry matter mass, N content and N accumulation in rice plants increased to different degrees under drought stress. The root soluble protein, nitrate reductase and soil urease activities were improved over those of the no-drought treatment. Proteobacteria, Bacteroidota, Nitrospirota and Zixibacteria were the dominant flora related to N absorption. A total of 184 DMs (98 upregulated and 86 downregulated) were identified between low N with no drought (LN) and normal N with no drought (NN); 139 DMs (83 upregulated and 56 downregulated) were identified between high N with no drought (HN) and NN; 166 DMs (103 upregulated and 63 downregulated) were identified between low N with drought stress (LND) and normal N with drought stress (NND); and 124 DMs (71 upregulated and 53 downregulated) were identified between high N with drought stress (HND) and NND. Fatty acyl was the metabolite with the highest proportion. KEGG analysis showed that energy metabolism pathways, such as D-alanine metabolism and the phosphotransferase system (PTS), were enriched. We conclude that N-metabolism enzymes with higher activity and higher bacterial diversity have a significant effect on drought tolerance and nitrogen uptake in rice.