Many microorganisms are found in the roots of plants, and are important for plant growth, nutrition uptake, and disease prevention. While the microorganisms of many crops such as soybean, rice, and maize have been studied, the root-associated microbial diversity of horticultural plants such as Paeonia lactiflora has been little studied. In this study, we investigated the composition, abundance, diversity and variability of microorganisms associated with different rhizospheric compartments (bulk soil, rhizosphere, rhizoplane, root endosphere) of different cultivars of P. lactiflora. We used the root and root-associated soil of a medicinal cultivar P. lactiflora cv ‘Hang Baishao’ (HS, 2n = 10) and an ornamental cultivar P. lactiflora cv ‘Zi Fengyu’ (ZFY, 2n = 10) as our study objectives. Based on Illumina Novaseq 6000 high-throughput sequencing, the 16S rRNA V4 amplicon of two cultivars of P. lactiflora root-associated bacterial communities and the ITS1-spanning amplicon of the fungal communities were sequenced, and the microbial communities were analyzed for relevant bioinformatics. Observed Species, Simpson, Shannon, and Chao1 indices showed that the microbial diversity of the rhizosphere was greater than that of the root, with the with the diversity decreasing from the bulk soil to the root endosphere. In addition, Cyanobacteria and Proteobacteria were the most abundant of the root-associated bacterial communities, accounting for 19.59 % and 31.23 %, respectively. On the other hand, Ascomycota (37.74 %) and Basidiomycota (11.15 %) were the most dominant fungal communities in the four root-associated compartments. Compared to ZFY, the root-associated compartments of HS were able to enrich more functional bacteria associated with bacterial inhibition, such as Kribbella, Actinophytocola and Actinoallomurus. This study explored the complex root microflora structure of P. lactiflora and provided a theoretical basis for future studies on the synthetic microbial community of soil.