Two freshwater armoured dinoflagellates, Peridiniopsis cf. kevei from Japan and Peridiniopsis penardii from Japan and Italy, were examined by means of light, scanning and transmission electron microscopy. Morphological studies indicated that the two dinoflagellates had similar type of cellular structure and possessed an endosymbiotic diatom. The diatom endosymbiont, which contained a eukaryotic nucleus, chloroplasts and mitochondria, was separated from the dinoflagellate cytoplasm by a single unit membrane. The dinoflagellate cytoplasm contained a triple-membrane-bound eyespot, in addition to typical dinoflagellate organelles. Molecular phylogenetic analyses based on small subunit ribosomal RNA gene (SSU rDNA) revealed a close relationship between these two dinoflagellates. They formed a clade with other dinoflagellates possessing a diatom endosymbiont, suggesting a single origin of diatom-harbouring dinoflagellates. On the contrary, the phylogenetic analyses based on plastid-encoded rbcL and nuclear-encoded SSU rDNA of the endosymbionts included the endosymbiont of these two freshwater dinoflagellates in the Thalassiosira/Skeletonema-clade (Centrales), whereas the endosymbionts of other diatom-containing dinoflagellates, except for Peridinium quinquecorne, were closely related to members of the Bacillariaceae (Pennales), most likely a Nitzschia-like diatom. The discrepancy between the host phylogeny and the endosymbiont phylogeny suggested, as in the case of Peridinium quinquecorne, that there was a serial replacement of endosymbionts from original a pennate Nitzschia-like diatom to a centric diatom, such as Thalassiosira (or possibly Skeletonema).