This study focused on the effect of predated sludge recycle on the contribution of extracellular polymeric substances (EPSs) and their subfractions to sludge aggregation in combined MBR system. It was observed that aggregation abilities of sludge samples were decreased by worm predation. Furthermore, worm predation enhanced the energy barriers and weakened the secondary energy minimum in the interaction energy profiles of slime, loosely bound EPS (LB-EPS) and tightly bound EPS (TB-EPS). Further investigations demonstrated that the content decrease and structural change of different EPS fractions induced by worm predation may be the reason for the decreased aggregation of sludge. Concomitantly, the adsorption tests and atomic force microscopy observation confirmed that the worm predation decreased the adsorption of slime, LB-EPS and TB-EPS on membrane. This would indicate the worm predation could keep an Optimum EPS level for which floc structure was maintained and the fouling propensity of mixed liquid was reduced. (C) 2013 Elsevier Ltd. All rights reserved.