Bitterling fishes lay their eggs on the gills of living freshwater mussels and are valuable models in behavioural and evolutionary ecology. To develop a fuller understanding of the coevolutionary relationship between European bitterling and mussels an aim of this study was to conduct a phylogenetic reconstruction of the colonisation of Europe by bitterling, and to resolve the taxonomic question of whether eastern and western populations ofbitterling represent the same species using genetic and traditional morphological/meristic methods. A second aim was to obtain a better understanding of the life-history parameters of R. amarus in Europe, including larval growth parameters. Previous studies have suggested the European bitterling is either a single species with a wide but discontinuous geographic distribution, or a complex of species. Morphometric and meristic data identified differences between three putative species; with a clear distinction between the eastern Asian Rhodeus sericeus sensu stricto, western European bitterling R. amarus and colchian bitterling, R. colchicus. Polymorphism. in the mitochondrial DNA control region was predominantly due to insertion/deletion events, making phylogenetic inference difficult. Cytochrome b data produced a phylogeny with strongly-supported differentiation between a clade of two R. sericeus haplotypes and a clade of six R. amarus/colchicus haplotypes. The starlike topology of the R. amarus/colchicus haplotypes in a minimum spanning network suggested a rapid radiation in this clade. The results are consistent with an hypothesis of relatively ancient divergence of R. sericeus from R. a11larus/colchicus and more recent and rapid differentiation between R. a11larus and R. colchicus. Analysis of otolith microstructure demonstrated a relationship between mussel availability for spawning and length of spawning season. In addition, these data strongly suggest the onset of exogenous feeding by larval R. amarus while still in their mussel host.