The development and productivity of plants are governed by their genetic background, nutrient input, and the microbial communities they host, i.e. the holobiont. Accordingly, engineering beneficial root microbiomes has emerged as a novel and sustainable approach to crop production with reduced nutrient input. Here, we tested the effects of six bacterial strains isolated from sugarcane stalks on sugarcane growth and physiology as well as the dynamics of prokaryote community assembly in the rhizosphere and root endosphere under two N fertilization regimes. All six strains, Paraburkholderia caribensis IAC/BECa 88, Kosakonia oryzae IAC/BECa 90, Kosakonia radicincitans IAC/BECa 95, Paraburkholderia tropica IAC/BECa 135, Pseudomonas fluorescens IAC/BECa 141 and Herbaspirillum frisingense IAC/BECa 152, increased in shoot and root dry mass, and influenced the concentration and accumulation of important macro- and micronutrients. However, N input reduced the impact of inoculation by shifting the sugarcane microbiome (rhizosphere and root endosphere) and weakening the co-dependence between soil microbes and sugarcane biomass and nutrients. The results show that these beneficial microbes improved plant nutrient uptake conditioned to a reduced N nutrient input. Therefore, reduced fertilization is not only desirable consequence of bacterial inoculation but essential for higher impact of these beneficial bacteria on the sugarcane microbiome. Display omitted •Inoculation shifted the sugarcane microbiome (rhizosphere and endosphere).•N input decreased the importance of plant growth promoting microbes.•Rhizosphere prokaryotes are associated with nutrient uptake in sugarcane.•N input reduced the co-dependence between soil microbes and plant nutrients.•Leifsonia abundance increased with N input as potential risk of ratoon stunting disease.