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Microbiota are essential components of the soil, driving biogeochemical cycles. Fungi affect decomposition and biotic interactions with plants across scales. Climate projections suggest that extended dry seasons may transform sensitive rain forests into savanna-like vegetation, with consequent changes in biogeochemistry. Here we compare the impacts of natural seasonality with 14 years of partial throughfall exclusion in an Amazonian rain forest, focussing on soil fungal functional diversity, extracellular soil enzyme activities (EEA) and their implications for nutrient dynamics. Large changes in fungal diversity and functional group composition occur in response to drought, with a conspicuous increase in the abundance of dark-septate fungi and a decrease in fungal pathogens. The high seasonality of EEA in the control (non droughted) and suppression of seasonality in the drought treatment, together with an increased implied nitrogen demand in the dry season induced by experimental drought, suggest that the changed soil microbiota activity may signal a pending shift in the biogeochemical functioning of the forest.