Eleven test runs (including two replicates) were carried out to explore the interaction of shear stress and chlorine concentration on the growth of heterotrophic microorganisms. Experimental results revealed that influent chlorine concentration and shear stress had no interaction on biofilm formation. Biofilm bacterial numbers decreased with the increase of influent chlorine concentration. Increasing the shear stress up to a specific level could significantly reduce the potential of biofilm formation. A strong interaction on bacterial quality or chlorine consumption rate of bulk water existed. With non-chlorinated and lower chlorinated conditions, the specific growth rate of biofilm increased with the increase of shear stress. However, an inverse relation occurred at higher chlorine conditions. No significant interaction of chlorine concentration and shear stress existed for particle numbers with 2–5, 5–15, 50–100 and >100 μm diameters. However, a significant interaction existed on particle numbers of 15–25 and 25–50 μm diameters.