Particle films are widely used in viticulture to protect vines from environmental stresses, but their effects on vineyard microbial communities, especially fermentation yeasts, are not well understood. This study assessed how pre-harvest applications of kaolin (5%) and potassium silicate (Si) at 0.05% and 0.10% influence yeast dynamics during spontaneous fermentation of Vitis vinifera “Touriga Nacional” and “Touriga Franca” musts. Grapes were harvested from a commercial vineyard in the Douro Region, and microvinifications were conducted under controlled conditions. Fermentations occurred in custom-designed fermenters, with daily monitoring. Yeast populations were analyzed at three stages (beginning, middle, and end) using culture-dependent methods and 26S rDNA sequencing. Particle film treatments affected yeast density, diversity, and composition, particularly in early and mid-fermentation. Non-Saccharomyces yeasts dominated initial stages, with marked differences in Saccharomyces cerevisiae presence across treatments. In “Touriga Nacional”, the 0.05% Si treatment promoted early S. cerevisiae dominance, unlike in “Touriga Franca”, where it was absent at the same stage. Kaolin-treated musts showed the lowest yeast diversity, while 0.05% Si-treated musts showed the highest. These results indicate that particle films significantly influence yeast communities during fermentation, with effects dependent on grape variety and treatment, reinforcing the need to consider microbial impacts in vineyard management.