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Extreme hydrological events cause stress in a soil environment, which directly affects the activity of the soil microbiome. The objective of this research is to determine the functional and structural biodiversity changes of soil bacteria in three different fluvisols from river floodplains under the condition of simulated floods. The nine soil sods (three for each fluvisol), with living vegetation, were collected in August 2018, and fitted into a separate transparent, polypropylene container. At the same time, water was taken from the Vistula River. The sods were flooded with water from the river to a level of 5 cm above the soil. The mesocosm experiment was conducted over 14 days under controlled conditions. During the experiment, the pH values, dehydrogenase activity, metabolic potential on EcoPlate™ and structure of the soil bacterial community by next generation sequencing (NGS) were analysed. The results obtained show that simulated flooding affects soil pH, dehydrogenase activity as well as metabolic potential and structural diversity of soil bacteria communities. The proportions of individual bacteria change, and new species appear in the soil, both those introduced by the water and woken up from inactive forms, while some groups of microorganisms die out. It can be unequivocally stated that extreme conditions caused by simulated flooding have a significant impact on soil microorganisms.