The safety of recycled water for potable water reuse can be enhanced by improving the removal of pathogenic microorganisms through reverse osmosis (RO) membrane. The present study aimed to identify the causes of the passage of particles through intact RO membrane elements using stable 0.5 mu m fluorescent (FL) microspheres as bacteria-sized surrogates. Pilot-scale tests revealed that the removal of FL particles varied between 1.6 and 6.1-log among seven commercial RO membrane modules, and a less permeable RO membrane had a higher efficiency for removing bacteria-sized particles. To clarify the causes of particles passing through the tight RO membrane, further investigations were performed focusing on the following three types of membranes based on the RO membrane manufacturing process: an ultrafiltration membrane (the supporting layer of RO membrane), and RO membranes obtained before assembly and after disassembly of their membrane elements. The highest concentrations of FL particles in permeate were obtained in the disassembled RO membranes, suggesting that the commercial spiral wound membrane elements may have lesser integrity than the intact RO membrane sheets for removal of bacteria-sized particles. However, the conductivity removal of the two types of RO membranes was equivalent regardless of the assembly process; thus, typical performance tests could not identify the membrane integrity breach for particle removal. To the best of our knowledge, this is the first study to report that the integrity of the RO membrane for particle removal may be compromised after assembling its spiral wound membrane element.