The genus Amphidinium Clap. & J. Lachm. comprises a high diversity of planktonic and benthic (epiphytic and sand-dwelling) dinoflagellates from marine and freshwater ecosystems. High morphological plasticity and vaguely defined genus characteristics (e.g., a small epicone size) have complicated the clear delineation of species boundaries. Although six Amphidinium morphospecies have been reported from Mexican coastal waters, species identifications are uncertain and not generally supported by molecular phylogenetic data. In this study, seven isolates of Amphidinium from diverse benthic coastal locations on the NE Pacific, Gulf of California, and southern Gulf of Mexico were subjected to critical morphological analysis using photonic and scanning electron microscopy. The phylogenetic reconstruction was based on nuclear-encoded, partial large-subunit (LSU) rDNA and internal transcribed spacer I and II (ITS1 and ITS2) sequences. The revised phylogenetic analysis was consistent with the traditional subdivision of the genus Amphidinium into two sister groups: Herdmanii and Operculatum clades. This study provided the first confirmed records of A. theodorei and A. massartii from coastal waters of Mexico. The molecular phylogenetic evidence indicated that the morphologically described A. cf. carterae from Baja California was in fact more closely allied with A. eilatiensis sequences. A few Amphidinium species are known to form toxigenic (i.e., fish-killing) harmful algal blooms worldwide, and therefore knowledge on species diversity and biogeography is critical in developing effective strategies for evaluating the potential emerging threat in Mexican coastal waters.