Abstract
INTRODUCTION: Host genotype is a key driver in shaping plant microbiome in response to dynamic changes in soil nitrogen (N) availability. However, the effects of rapeseed (Brassica napus) genotypes with different N use efficiency characteristics on microbiome assembly, as well as the underlying plant-microbe interaction mechanisms, remain poorly understood. OBJECTIVES: This study aims to: (1) assess microbial assembly differences between N-use efficient and inefficient genotypes; (2) identify specific microbiota associated with plant N acquisition; and (3) reveal the molecular mechanisms driving plant-microbe interactions. METHODS: We conducted comparative microbiome profiling of N-use efficient and inefficient genotypes, followed by functional validation of microbial roles in plant N uptake. Multi-omics approaches, including RNA-seq and metabolomics, were used to uncover the regulatory interactions between the host and rhizosphere microbiota. RESULTS: The N-use efficient genotype constructed more diverse root-associated microbes than the inefficient genotype, with Bacillaceae emerging as the most enriched taxon. A representative isolate, Bacillus sp. 41S2 from the N-use efficient genotype, markedly enhanced root biomass and N uptake in the N-use inefficient genotype, as confirmed by (15)N tracer assays. RNA-Seq analysis further demonstrated that genes involved in jasmonic acid and ethylene signaling pathways were upregulated in strain 41S2-inoculated plants, likely contributing to enhanced root development. Metabolomic profiling identified kaempferol, a flavonol with the highest fold-change between genotypes, as a key root exudate promoting the growth and biofilm formation of strain 41S2. Furthermore, the fls1 mutant (deficient in kaempferol biosynthesis) failed to recruit Bacillaceae, confirming the role of kaempferol in mediating genotype-specific microbial enrichment. CONCLUSIONS: Our findings reveal a novel, microbe-dependent N acquisition pathway in N-use efficient rapeseed genotypes, driven by kaempferol-mediated recruitment of Bacillaceae. This work highlights the potential of host genotype and metabolite signaling to shape beneficial microbiota for improved nutrient efficiency and sustainable crop production.