The use of nitrogen-fixing bacteria in agriculture is increasingly recognized as a sustainable method to boost crop yields, reduce chemical fertilizer use, and improve soil health. However, the microbial mechanisms by which inoculation with nitrogen-fixing bacteria enhance rice production remain unclear. In this study, rice seedlings were inoculated with the nitrogen-fixing bacterium R3 (Herbaspirillum) at the rhizosphere during the seedling stage in a pot experiment using paddy soil. We investigated the effects of such inoculation on nutrient content in the rhizosphere soil, plant growth, and the nitrogen-fixing microbial communities within the rhizosphere and endorhizosphere. The findings showed that inoculation with the R3 strain considerably increased the amounts of nitrate nitrogen, ammonium nitrogen, and available phosphorus in the rhizosphere by 14.77%, 27.83%, and 22.67%, respectively, in comparison to the control (CK). Additionally, the theoretical yield of rice was enhanced by 8.81% due to this inoculation, primarily through a 10.24% increase in the effective number of rice panicles and a 4.14% increase in the seed setting rate. Further analysis revealed that the structure of the native nitrogen-fixing microbial communities within the rhizosphere and endorhizosphere were altered by inoculation with the R3 strain, significantly increasing the α-diversity of the communities. The relative abundance of key nitrogen-fixing genera such asRalstonia,Azotobacter,Geobacter,Streptomyces, andPseudomonaswere increased, enhancing the quantity and community stability of the nitrogen-fixing community. Consequently, the nitrogen-fixing capacity and sustained activity of the microbial community in the rhizosphere soil were strengthened. Additionally, the expression levels of the nitrogen absorption and transport-related genesOsNRT1andOsPTR9in rice roots were upregulated by inoculation with the R3 strain, potentially contributing to the increased rice yield. Our study has revealed the potential microbial mechanisms through which inoculation with nitrogen-fixing bacteria enhances rice yield. This finding provides a scientific basis for subsequent agricultural practices and is of critical importance for increasing rice production and enhancing the ecosystem services of rice fields.

Submergence stress is a major constraint in direct-seeded rice production. This study investigated the effect and biochemical mechanism of sprouting, a traditional agronomic practice, on improving submergence tolerance in rice. Our findings demonstrate that sprouting is an effective seed treatment that significantly enhances the plant’s ability to withstand flooding. Specifically, 48 h sprouting increased shoot height and root length by 163% and 423%, respectively, in the YLYJ48 variety under 6-day submergence. Sprouting upregulated the activity of glutathione reductase (GR) and the expression of its related genes, thereby significantly promoting the biosynthesis of glutathione (GSH). GSH content in seeds increased from 64.86 µg g⁻¹ FW (0 h) to 83.00 µg g⁻¹ FW (48 h) in HZ, and from 82.14 to 92.52 µg g⁻¹ FW in YLYJ48. This process provides critical antioxidant protection for seedlings to implement a rapid “escape strategy,” ultimately enhancing their submergence tolerance. Functional verification showed that seed soaking with exogenous GSH (0.1%) effectively improved submergence tolerance by increasing antioxidant reserves. Exogenous GSH treatment elevated shoot height by approximately 50% in both HZ and YLYJ48 varieties under submergence. Field trials further demonstrated that exogenous GSH application significantly enhanced seedling establishment rates by 30–35% and improved seedling growth traits under submergence tolerance stress across multiple rice varieties. This study reveals part of the mechanism by which sprouting enhances submergence tolerance by influencing glutathione metabolism, offering practical strategies for flood-resilient direct-seeded rice cultivation.

S-metolachlor is a selective pre-emergence herbicide used in dryland. However, it is challenging to employ in paddy fields due to its phytotoxic effects on rice. As a common phytohormone, Gibberellin-3 (GA₃) is inferred to have the ability to alleviate herbicide phytotoxicity. This study first quantitatively verified the phytotoxicity of s-metolachlor to rice and then demonstrated the mitigative effect of GA₃ on these adverse reactions. Furthermore, a transcriptome of rice seedlings subjected to different treatments was constructed to assemble the reference genes, followed by comparative metabolomics and proteomics analyses. Metabolomics revealed an enrichment of flavonoid metabolites in the group of adding GA₃, and these flavonoids can eliminate ROS in plants. Proteomics analysis indicated that differential proteins were enriched in the phenylpropanoid biosynthesis pathway responsible for the synthesis of flavonoids and that the functions of most differential proteins are associated with peroxidase. The proteome, combined with the transcriptome, revealed that the expressions of proteins and genes was related to the POD activity in the group of adding GA₃. It was speculated that the elimination of ROS is key to alleviating the stress of s-metolachlor on rice growth. It was inferred that the mechanism of GA₃ in alleviating the phytotoxicity of the substance s-metolachlor is by increasing the activity of the POD and influencing the growth of rice seedlings through the restoration of flavonoid synthesis. In this study, we screened GA₃ as a safener to alleviate the phytotoxicity of s-metolachlor on rice. On this basis, the mechanism of alleviating phytotoxicity was studied. The application range of s-metolachlor might be expanded, providing a new supplementary method for weed control and herbicide resistance management.