Sediment particles are often colonized by biofilm in a natural aquatic ecological system, especially in eutrophic water body. A series of laboratory experiments on particle size gradation were conducted after natural sediment was colonized by biofilm for 5, 10, 15 and 20 days. Particle image acquisition and particle tracking techniques were utilized to analyze the changes of these properties. The experimental results indicate that the size gradation of bio-particles underwent significant change due to the growth of biofilm onto the sediment surface. The bio-particle diameter will become 1 to 2 orders of magnitude larger than the initial size. Because of the drag force, in the flowing water, the diameter will decrease with the increasing velocity. These results may be specifically used in the low energy reservoir or lake environment.