The water regime is often the primary force driving the evolution of freshwater lakes, but how soil microbes responded to this process is far from understood. This study sampled wetland soils from a shallow lake that experienced water regime changes, Poyang Lake of China, to explore the features of bacterial and fungal community in response to water level changes. The soil physicochemical properties, T-RFLP based community structures and soil activities (including basal respiration, microbial biomass and enzymes) were all determined. Soil microbial eco-function was captured by testing the carbon metabolism with Biolog-Ecoplate. The results showed remarkable influence of the water level gradients on the soil physicochemical properties, microbial community structures and soil activities. However, the carbon utilization profile exhibited weak connections with the environmental variables and microbial community structures (p > 0.05). The microbial activities were significantly correlated with both bacterial and fungal community structures. Our results also emphasized the ascendant role of the deterministic process in the assemblages of microbial community structures and functions in wetland. In conclusion, this study revealed the discrepancy between community structures and eco-functions in response to water level gradients, and a relatively stable eco-function helped to maintain the ecosystem function of wetland from a long-term perspective.