Background and aims <p>Understanding how maize genotype and nitrogen (N) availability interact to shape root-associated microbiomes is key to improving nitrogen use efficiency (NUE) for sustainable agriculture. This study investigates how N levels and maize genotypes co-regulate rhizosphere and root endophytic bacterial communities and their contribution to N efficiency.</p> Methods <p>Field experiments evaluated four maize hybrids with contrasting NUE (N-efficient: Zhengdan 958, Yudan 132; N-inefficient: Xundan 1618, Yudan 922) under high N (HN, 210&#xa0;kg ha⁻¹) and low N (LN, no fertilizer) treatments. Rhizosphere and root bacterial communities were characterized via 16S rRNA sequencing. Plant biomass, yield, N accumulation, and plant growth-promoting bacteria (PGPB) abundances were analyzed.</p> Results <p>Under LN stress, N-efficient genotypes maintained higher biomass and yield, with a smaller reduction in N accumulation (20–21%) than N-inefficient lines (30–52%). Genotype was the dominant factor influencing microbial assembly (PERMANOVA: rhizosphere R² = 20.6%, root endophytic R² = 33.8%, <i>P</i> &lt; 0.001). N-efficient lines specifically enriched beneficial taxa such as <i>Sphingobium</i> and <i>Mesorhizobium</i> in the root endosphere. The abundance of PGPB in roots increased by 70% under LN in efficient genotypes and was 125% higher than in inefficient lines. Critically, root endophytic diversity correlated positively with yield and N accumulation, whereas rhizosphere diversity showed negative correlations, underscoring the functional importance of endophytes in NUE.</p> Conclusions <p>Maize genotypes modulate root-associated microbiomes in response to N availability. N-efficient lines recruit key beneficial endophytes and enhance PGPB under LN, highlighting root endophytes as critical targets for microbiome-assisted nitrogen management.</p>