Aims: We determined if the intra-specific genetic diversity of Microcystis aeruginosa correlates with phenotypic characteristics. Methods and Results: Microcystis aeruginosa isolates from various Japanese waters were characterized using genetic analyses based on the 16S-23S rDNA internal transcribed spacer (ITS) region and DNA-independent RNA polymerase (rpoC1) gene sequences. In addition, morphological and biochemical properties, and the toxicity of M. aeruginosa strains were determined. We found a correlation in phylogenetic clusters of the ITS region and rpoC1 gene sequences. Using a polyphasic approach, genotypic and phenotypic variations in M. aeruginosa showed that the three genetic lineage groups are comprised of a particular phenotype or subgroup of closely related phenotypes. However, some strains had high phenotypic and genotypic diversity compared to the three lineage groups and did not show distinct lineages; therefore, these strains were designated as the 'complex group'. Conclusions: The 'complex group' consisted of genetically and phenotypically incoherent and high diverse populations in M. aeruginosa, although some genotypes or lineages displayed consistent phenotypes. Significance and Impact of the Study: The polyphasic approach combining phenotypic and genetic characterization was effective for comprehending distinct lineages and discriminating the potential complexity of M. aeruginosa populations at the intra-species level.