Microbial communities living in nine vineyards distributed over three altitudinal transects were studied over 2 years. Fungal and bacterial community dynamics were explored using automated ribosomal intergenic spacer analysis (ARISA) and by determining bacterial cells and fungal colony-forming units (CFUs). Moreover, extensive chemical and physical analyses of the soils were carried out. Multivariate analyses demonstrated that bacterial and fungal communities are affected by altitude, which acts as a complex physicochemical gradient. In fact, soil moisture, Al, Mg, Mn and clay content are changing with altitude and influencing the bacterial genetic structure, while in the case of fungi, soil moisture, B and clay content are found to be the main drivers of the community. Moreover, other exchangeable cations and heavy metals, not correlating with altitude, are involved in the ordination of the sites, especially Cu. Qualitative ARISA revealed the presence of a stable core microbiome of operational taxonomic units (OTUs) within each transect, which ranged between 57% and 68% of total OTUs in the case of fungi and between 63% and 72% for bacteria. No seasonal effect on the composition of microbial communities was found, demonstrating that bacterial and fungal communities in vineyards are mostly stable over the considered seasons.