The widespread application of elemental sulfur (S0) to vineyards may have ecosystem effects at multiple scales. We evaluated the short-term fates of applied S0 in a Napa Valley vineyard; we determined changes in soil sulfur (S) speciation (measured by X-ray absorption near-edge structure (XANES) spectroscopy), soil pH, extractable sulfate (SO4 2−), and total S to evaluate changes in acidity and soil S within the vineyard over time. Surface soil samples were collected immediately prior to and following two applications of S0 (6.7 kg S0 ha−1), with weekly collections in the 2 weeks between applications and following the last application. XANES spectra indicated that the majority of soil S persists in the +6 oxidation state and that S0 oxidizes within 7 days following application. Soil pH and extractable SO4 2− measurements taken at 30 min after S0 application revealed generation of acidity and an increase in extractable SO4 2−, but by 12 days after application, soil pH increased to approximately pre-application levels. These data suggest that the major consequence of reactive S applications in vineyards may be the accumulation of soil SO4 2− and organic S during the growing season, which can be mobilized during storm events during the dormant (wet) season. In spatially-extensive winegrowing regions where these applications are made by hundreds of individual farmers each year, it will be important to understand the long-term implications of this perturbation to the regional S cycle.