[Objective] We identified predominant strains causing membrane biofouling in membrane bioreactors (MBR). [Methods] 454 high-throughput pyrosequencing was applied to analyze the genes of bacteria originated from the sludge mixed liquor (S1) and membrane foulant (S2). Chao richness index and Shannon diversity index of two samples were calculated and the sequencing results were analyzed by phylogenetic analysis. [Results] In total 9 353 and 7 504 effective reads were obtained from S1 and S2, respectively, indicating that the microbial richness and diversity of S2 were higher than those of S1. By performing the phylogenetic spectra, we found that the microbial community structure of S2 was changed during the membrane surface colonization, and in S2, the abundance of β-Proteobacteria was significantly reduced while α-Proteobacteria、γ-Proteobacteria and Phycisphaerae were enriched. [Conclusion] 454 high-throughput pyrosequencing revealed that Xanthomonadaceae, Thermoanaerobacter, Phycisphaera and 2 strains of uncultured bacteria (Candidate_division_TM7 and Candidate_division_OD1) were predominant taxons involved in membrane fouling. These strains included those with high surface viscosity and hydrophobicity (e.g., γ-Proteobacteria) which were prone to adhesion onto the membrane surfaces, and the proficient species (e.g., Candidate_division_OD1) that facilitated the interspecies hydrogen transfer.