The diversity of buoyant Microcystis colony-associated, settling particle-associated and free-living freshwater bacteria in the eutrophic Lake Taihu in China was compared by the polymerase chain reaction–denaturing gradient gel electrophoresis (PCR–DGGE) of 16S rRNA genes and by clone library analysis. The cluster analysis of DGGE profiles revealed that the buoyant Microcystis colony associates collected in summer and winter were clustered together and were distinct from settling particle-associated and free-living bacteria. In contrast, the bacterial communities of the latter two populations collected in summer were clustered together and varied from those collected in winter. The diversity indices of the Microcystis-associated bacterial population were significantly lower than those of the other two bulk bacterial communities (P < 0.05). Clone library analysis revealed that no shared operational taxonomic units were found in the three populations. Phylogenetic analysis confirmed that buoyant Microcystis-associated bacteria were distinct from the other communities and that they were dominated by Alphaproteobacteria, Betaproteobacteria and Gammaproteobacteria. Proteobacteria, Bacteroidetes and Actinobacteria were prevalent in all three habitats. Other bacterial taxa such as Planctomycetes, Verrucomicrobia, Gemmatimonadetes and Acidobacteria were associated with settling particles and with the water column. Buoyant Microcystis colonies, settling particles and the water column in the eutrophic lake are thus inhabited by different bacterial flora. In addition, specific bacterial communities are associated with buoyant Microcystis colonies.