The aim of this study was to determine the abundance and diversity of root endophytic fungi in organic versus conventional vineyards on the eastern Adriatic coast (Croatia). Samples were collected from four locations in the karst region along the coast. Each of these locations included both organically and conventionally managed vineyards of cv. ‘Plavac mali’ on Vitis berlandieri × Vitis rupestris rootstock. The total root colonization ranged from 39% to 78% for the grapevines, and from 9% to 99% for the dominant weeds, with generally significantly higher fungal colonization in the organic than the conventional vineyards. Increased abundance of the smallest class of fungal spore (diameter, 40–100 μm) was observed in organic vineyards. There was a similar pattern for fungal diversity, with the overall dominance of Ascomycota, and an average of 8.5% of all sequences belonging to Glomeromycota. Our study confirms the initial hypothesis that the change in vineyard soil management toward more sustainable practices improves the abundance and diversity of endophytic fungi, thus suggesting their greater ecological importance in environments with significantly reduced inputs.