The tree fruit industry in Nova Scotia, Canada, is dominated by the apple (Malus domestica) sector. However, the sector is faced with numerous challenges, including apple replant disease (ARD), which is a well-known problem in areas with intensive apple cultivation. A study was performed using 16S rRNA/18S rRNA and 16S rRNA/ITS2 amplicon sequencing to assess soil- and root-associated microbiomes, respectively, from mature apple orchards and soil microbiomes alone from uncultivated soil. The results indicated significant (p< 0.05) differences in soil microbial community structure and composition between uncultivated soil and cultivated apple orchard soil. We identified an increase in the number of potential pathogens in the orchard soil compared to uncultivated soil. At the same time, we detected a significant (p< 0.05) increase in relative abundances of several potential plant-growth-promoting or biocontrol microorganisms and non-fungal eukaryotes capable of promoting the proliferation of bacterial biocontrol agents in orchard soils. Additionally, the apple roots accumulated several potential PGP bacteria from Proteobacteria and Actinobacteria phyla, while the relative abundances of fungal taxa with the potential to contribute to ARD, such as Nectriaceae and plant pathogenicFusariumspp., were decreased in the apple root microbiome compared to the soil microbiome. The results suggest that the health of a mature apple tree can be ascribed to a complex interaction between potential pathogenic and plant growth-promoting microorganisms in the soil and on apple roots.