Table_5_Integrated 16S and metabolomics revealed the mechanism of drought resistance and nitrogen uptake in rice at the heading stage under different nitrogen levels.docx

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.

全球范围内的常规农业生产模式，均因干旱事件的频发而受到显著负面影响。水稻根际微生物同样会受到干旱胁迫的显著影响。为给提升水稻抗旱性与氮素利用效率提供假说支撑，本研究于抽穗期采用桶栽种植模式，设置无干旱与干旱胁迫两种水分处理，并搭配三个不同施氮梯度。本研究采用非靶向代谢组学与16S rRNA基因测序技术，分别解析水稻根系微生物群落与根际土壤的差异代谢物（DMs）变化特征。
结果显示，在相同施氮量条件下，干旱胁迫下水稻植株的干物质质量、氮素含量与氮素积累量均呈现不同程度的提升。水稻根系可溶性蛋白含量、硝酸还原酶活性与土壤脲酶活性，均显著高于无干旱处理组。变形菌门（Proteobacteria）、拟杆菌门（Bacteroidota）、硝化螺旋菌门（Nitrospirota）与齐西菌门（Zixibacteria）为与氮素吸收相关的优势菌群。
在无干旱的低氮组（LN）与正常氮组（NN）之间，共鉴定出184种差异代谢物（上调98种、下调86种）；无干旱的高氮组（HN）与NN组之间，鉴定出139种差异代谢物（上调83种、下调56种）；干旱胁迫下的低氮组（LND）与正常氮组（NND）之间，鉴定出166种差异代谢物（上调103种、下调63种）；干旱胁迫下的高氮组（HND）与NND组之间，鉴定出124种差异代谢物（上调71种、下调53种）。脂肪酰类代谢物为占比最高的代谢物类别。KEGG富集分析结果显示，D-丙氨酸代谢、磷酸转移酶系统（PTS）等能量代谢通路发生显著富集。
本研究认为，高活性氮代谢酶与较高的细菌多样性，对水稻的抗旱性与氮素吸收具有显著影响。