Diarrhoeic shellfish poisoning has increasingly become a problem throughout the world. Because the causative organisms Dinophysis spp. cannot be cultured in the laboratory, new approaches are needed to obtain ecological and physiological information. In this study, D. acuminata. D. norvegica and D. acuta were collected directly from field samples and used in polymerase chain reactions. The D1–D2 region of the large-subunit ribosomal RNA gene was amplified, cloned and sequenced. Sequence analyses showed that D. acuminata and D. norvegica were nearly identical (> 99%), and that D. acuminata showed an intraspecies variation of 0.8%. The D. acuta sequence was 98.7% similar to that of D. acuminata. The slight differences between D. norvegica and D. acuminala suggest that they may have evolved into separate species rather recently. Phylogenetic analyses show that species within the Dinophysiales order should be included in the ‘GPP complex’, a lineage associated with a diverse array of taxa within the orders Gymnodiniales, Prorocentrales and Peridiniales. The Prorocentrales and Dinophysiales would be sister groups within the GPP complex. Amplification of Swedish D. acuminata isolates always resulted in a single LSU rDNA fragment. In contrast, amplification of the North American D. acuminata always produced two distinct fragments. The longer (735 bp) fragment showed 99.3–1 00% homology among all sequenced clones of different D. acuminata field isolates. The shorter gene fragment had a 70 bp deletion, but it was otherwise highly homologous to the larger gene fragment. This fragment is possibly a pseudogene and might be an important genetic marker. A variable region that is suitable as a target for a probe to identify Dinophysis was also identified. Dinophysis specificity was confirmed for the probe, in that hybridization to cultured representatives of dinoflagellates and environmental samples containing mixed phytoplankton assemblages resulted in specific labelling of D. acuminata. D. norvegica and D. acuta, but not other dinoflagellates. No labelling of D. rotundata was observed.