Incipient motion of sediment, which is important in fluvial processes, can be affected by biofilm growth. In this paper, a series of flume experiments are carried out using non-uniform sediments, to investigate the entrainment of sediment with and without biofilm. Differences in entrainment and the velocity at incipient motion are measured over an eight week period, as well as the biomass dynamics. Sediment with biofilm is more stable, i.e., a greater incipient velocity, and the incipient velocity increases to a threshold level over time and then declines. Biofilm development is clearly an important control on the stability of sediments, especially in eutrophic water bodies. A theoretical expression of incipient velocity that includes the cohesive and adhesive forces is derived, where the biomass dynamics are directly incorporated. Such analyses can help to predict sediment transport changes due to biofilm presence in nutrient-rich water bodies.