The present study investigates the functionalecological status of typical light clay soils in vineyards in the southern region of Crimea, using both traditional (including fallow soils) and organic land-use systems. This analysis was carried out by examining agrochemical indicators, microbial respiratory activity, microbial biomass, and the ecological status of the microbial community. In organic vineyard soils, the mean substrate-induced respiration, microbial biomass carbon and the ratio of microbial biomass to organic carbon were found to be 2.8, 4.0, and 4.1 times higher, respectively, compared to conventional farm soils. On the contrary, the microbial metabolic coefficient was 1.4 times lower, signifying more favorable conditions for the functioning of the soil microbiota. The increased mobile sulfur content in organic vineyard soils (18.3 mg kg−1 vs. 8.0 mg kg−1 in traditional farms) and inadequate mobile phosphorus supply in some farms present potential risks. The suboptimal functional state of the microbiome in fallow soils previously under traditional plant protection necessitates comprehensive ecotoxicological analyses before development. Assessing the soil functional ecological status through an ecophysiological evaluation of the microbiome is vital for understanding ampelocenosis soils and making informed decisions on vineyard management practices.