Soil microbial communities significantly contribute to global fluxes of nutrients and carbon. Their response to climate change, including winter warming, is expected to modify these processes through direct effects on microbial functions through osmotic stress, and changing temperature regimes. Using four European peatlands reflecting different frequencies of frost events, we show that peatland testate amoeba communities diverge among sites with different winter climates, and that this is reflected through contrasting functions. We found that exposure to harder soil frost promoted species β-diversity (species turnover) thus shifting the community composition of testate amoebae. In particular, we found that harder soil frost, and lower water-soluble phenolic compounds, induced functional turnover through the decrease of large species (-68%, > 80 μm) and the increase of small-bodied mixotrophic species (i.e. Archerella flavum; +79%). These results suggest that increased exposure to soil frost could be highly limiting for large species while smaller species are more resistant. Furthermore, we found that β-glucosidase enzymatic activity, in addition to soil temperature, strongly depended (R2 = 0.95, ANOVA) of the functional diversity of testate amoebae. Changing winter conditions can therefore strongly impact peatland decomposition process, though it remains unclear if these changes are carried–over to the growing season.