Carbon nanoparticles (CNPs) can potentially promote plant biomass and root growth by modulating the rhizosphere microbial community. However, the mechanism between CNPs and rhizosphere microorganism remains largely elusive. The purpose of this study was to systematically explore the effects of CNPs on the diversity and structure of rhizosphere soil bacterial and fungal communities, as well as soil enzyme activities and nutrients. In this study, we applied 16S and ITS sequencing techniques to analyze the changes of bacterial and fungal communities for tobacco under CNPs (0.6 and 1.2 g.L^-1 by concentration) treatment at three different developmental stages. Our results demonstrated there were significant reductions in pH and available phosphorus contents after the treatment of CNPs, while the total nitrogen and total phosphorus were increased by CNPs application. Also, less sucrase, ß-1,4-glucosidase, polyphenoloxidase and acid phosphatase were observed in CNPs treatment soils. This study indicated that CNPs affected the diversity, composition and structure of the rhizosphere bacterial community. Fungal and bacterial communities had different response pattern for CNPs treatment, with phased and dose-effect effects. Compared with control, CNPs significantly increased the relative abundances of some potential beneficial bacteria, including Burkholderia genus, Sphingomonas genus, Lactobacillus genus. Furthermore, 278 culturable bacteria and 25 culturable fungi were isolated from soil and roots, and the strains enriched under CNPs treatment were tested for their ability to promote plant growth. Finally, five of them were validated for their microbial-mediated growth effects on tobacco. Metagenomic analysis revealed that CNPs increased functional diversity of root endosphere, with genes related to nutrient metabolism and plant hormone biosynthesis enriched after CNPs treatment. In general, our results have demonstrated the key role of rhizosphere microorganism in the interaction between CNPs and plants, and provide evidence and strategies for promoting tobacco growth with CNPs.