This paper adds new insight to a molecular phylogeny of Gyrodactylus, based on a complete sequence of the ITS rDNA region of 4 subgenera and a more detailed molecular analysis. We propose a hierarchical approach in elucidating the phylogeny of this species-rich genus. A total of 37 sequences (915-1239 bp) from 10 representative species from 4 out of 6 subgenera, as defined by Malmberg (1970), are included in the analysis. Genetic differences observed at the 5.8S locus provide objective criteria to separate (sub)genera, while deep genetic differences of the spacers form a sound basis for species-specific identification. We demonstrate that each Gyrodactylus subgenus possesses a unique sequence of the 5.8S gene. Thus, there is concordance between the 5.8S gene and the excretory system used by Malmberg (1970) as a diagnostic character of subgenus status. At the species level, there is a discrepancy between morphological and molecular variation. Whereas the morphological variation, expressed in the shape and size of the attachment apparatus, is very low, the molecular variation, expressed at the I