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Extended drought periods followed by heavy rainfall may increase in many regions of the Earth, but the consequences for the quality of soil organic matter and soil microbial communities are poorly understood. Here, we investigated the effect of repeated drying and re-wetting on microbial communities and the quality of particulate and dissolved organic matter in a Haplic Podzol from a Norway spruce stand. After air-drying, undisturbed soil columns were re-wetted at different intensities (8, 20 and 50 mm per day) and time intervals, so that all treatments received the same amount of water per cycle (100 mm). After the third cycle, SOM pools of the treatments were compared with those of non-dried control columns. Lignin phenols were not systematically affected in the O horizons by the treatments whereas fewer lignin phenols were found in the A horizon of the 20- and 50-mm treatments. Microbial biomass and the ratio of fungi to bacteria were generally not altered, suggesting that most soil microorganisms were well adapted to drying and re-wetting in this soil. However, gram-positive bacteria and actinomycetes were reduced whereas gram-negative bacteria and protozoa were stimulated by the treatments. The increase in the (cy 17: 0 + cy 19: 0)/(16:1ω7c + 18:1ω7c) ratio indicates physiological or nutritional stress for the bacterial communities in the O, A and B horizons with increasing re-wetting intensity. Drying and re-wetting reduced the amount of hydrolysable plant and microbial sugars in all soil horizons. However, CO2 and dissolved organic carbon fluxes could not explain these losses. We postulate that drying and re-wetting triggered chemical alterations of hydrolysable sugar molecules in organic and mineral soil horizons.