Aims: The aim of this work was to investigate the effects of biodynamic management with and without the addition of green manure, in comparison with organic management, on the microbiota in vineyards soil. Methods and Results: High throughput sequencing was used to compare the taxonomic structure of the soil bacterial and fungal communities from vineyards managed with different methods (organic, biodynamic or biodynamic with green manure). Our results showed that microbial communities associated with biodynamic and organic farming systems were very similar, while green manure was the greatest source of soil microbial biodiversity and significantly changed microbial richness and community composition compared with other soils. Green manure also significantly enriched bacterial taxa involved in the soil nitrogen cycle (e.g. Microvirga sp., Pontibacter sp. and Nitrospira sp.). Conclusions: Our results showed that the diversity and composition of the microbial communities associated with biodynamic and organic farming systems were similar, indicating that the use of biodynamic preparations 500 and 501 did not cause any significant detectable changes to the soil microbial community in the short term, while the effects of green manure were significant in soil microbiota. Significance and Impact of the Study: The microbiological richness and structure of soil are used as a sensitive indicator of soil quality. The extension of organic/biodynamic farming, associated with green manure application, could contribute to increase the abundance of functional groups of biological and agronomical relevance and maintaining microbial biodiversity in vineyard soils.