The delineation of species boundaries in the potentially harmful cyanobacterium Planktothrix Anagnostidis et Komárek 1988 is particularly tangled. Genetic recombination has been invoked to explain the occurrence of overlapping biological traits among recognized species. Although horizontal gene transfer is shown as a driver of diversification in this genus, clear evidence for homologous recombination at the single gene level is still lacking. Several Planktothrix strains (n = 244) sampled in eight fresh water lakes in north Italy were characterized by sequencing the rpoC1 gene, a molecular marker previously proposed to discriminate between species. Six haplotypes were detected, four of which are newly described. A relevant number of rpoC1 sequences (n = 54) showed evidence of homologous recombination. By comparing the sequences produced in the work presented here to those available on GenBank for the genus, multiple recombination events were tracked between haplotypes associated to P. rubescens, P. suspensa and P. agardhii, the latter being a species not found in our survey. Recombination signals were in form of (i) a vast mosaic structure present in the alignment of rpoC1 haplotypes, (ii) multiple and statistically significant paths in the split decomposition network connecting these haplotypes and (iii) many individual crossing-over events detected by means of recombination detection tests. Data suggest that the molecular evolution of the rpoC1 gene in the genus Planktothrix appears as strongly influenced by homologous recombination. In addition, rpoC1 diversity effectively tracks recombinational processes among species in the complex made up by P. rubescens, P. agardhii and P. suspensa, which are not isolated in terms of gene-flow.