The family Kareniaceae is mostly known in France for recurrent blooms of Karenia mikimotoi in the Atlantic, English Channel, and Mediterranean Sea and for the unusual green discoloration in the saltwater lagoon of Diana (Corsica) caused by Karlodinium corsicum in April 1994. In terms of diversity, this taxonomic group was long overlooked owing to the difficult identification of these small unarmored dinoflagellates. In this study, thanks to the molecular characterization performed on single cells from field samples and cultures, twelve taxonomic units were assigned to the known genera Karenia, Karlodinium and Takayama, whereas one could not be affiliated to any described genus. The molecular phylogeny inferred from the D1–D2 region of the LSU rDNA showed that five of them formed a sister taxon of a known species, and could not be identified at species-level, on the basis of molecular analysis only. Among these latter taxa, one Karlodinium which was successfully cultured was investigated by studying the external morphological features (using two procedures for cells fixation), ultrastructure, pigment composition, and haemolytic activity. The results of our analyses corroborate the genetic results in favour of the erection of Karlodinium gentienii sp. nov., which possesses an internal complex system of trichocysts connected to external micro-processes particularly abundant in the epicone, and a peculiar pigment composition. In addition, preliminary assays showed a haemolytic activity.