We developed a new protocol to study arbuscular mycorrhizal fungal (AMF) communities in Bordeaux vineyards in a standardized way, in order to compare results obtained over years and between locations. To this end, we first used grapevine root samples instead of soil samples to avoid AMF spores or hyphae interacting with cover plants. We next increased the number of grapevine samples to obtain more representative coverage of AMF communities while decreasing variability intra-vineyard, especially for the larger parcels. In addition, we adapted the DNA extraction protocol dedicated to soil samples to grapevine roots, as a way to increase the yield and the purity of samples. These features, coupled to the choice of the LSU-D2 region as the molecular marker for highthroughput sequencing (MiSeq® technology), further allowed us to assess the AMF populations of Bordeaux vineyards.