96 J. Phycol. 38, 96–106 (2002) SINGAPORE ISOLATES OF THE DINOFLAGELLATE GYMNODINIUM CATENATUM (DINOPHYCEAE) PRODUCE A UNIQUE PROFILE OF PARALYTIC SHELLFISH POISONING TOXINS 1 Michael James Holmes 2 Department of Biological Sciences, National University of Singapore, Singapore 119260 Christopher J. S. Bolch, David H. Green Scottish Association for Marine Science, Dunstaffnage Marine Laboratory, Oban, Argyll, Scotland, PA34 4AD, United Kingdom Allan D. Cembella Institute of Marine Biosciences, National Research Council of Canada, 1411 Oxford Street, Halifax, Nova Scotia, Canada B3H 3Z1 and Serena Lay Ming Teo Tropical Marine Science Institute, National University of Singapore, Singapore 119260 We investigated the cell morphology, toxicity and toxin composition, and rDNA sequences of clonal cultures of the chain-forming dinoflagellate Gymno- dinium catenatum H.W.Graham isolated from the port of Singapore. The cell morphology was consis- tent with most descriptions of this species except for sparsely distributed putative trichocyst pores visible on some cells under SEM. Nucleotide sequences (697 base pair) of the D1–D2 conserved regions and intervening variable domain of the large subunit rDNA were identical among isolates from Singapore and those of all other global populations examined so far (from Australia, China, Japan, Korea, New Zealand, Spain, and Uruguay), and this is consistent with the morphological conservatism of the species. Among isolates of G. catenatum that produce toxins associated with paralytic shellfish poisoning, the cel- lular toxicity of Singapore clones, as determined by intraperitoneal mouse bioassay (30–50 pg saxitoxin equivalentscell -1 ) and immunoassay (24  8 saxitoxin equivalentscell -1 ) was relatively high. The mouse bio- assay toxicity was comparable with that of Spanish and Philippine isolates that have undergone acid hydrolysis. However, analysis of toxin composition of Singapore clones by HPLC with fluorescence detection or HPLC- mass spectrometry/mass spectrometry revealed a unique toxin profile that was dominated by the highly potent carbamate toxins, primarily gonyautoxin (GTX) 1 and 4 with lesser amounts of GTX2, GTX3, neosax- itoxin, and saxitoxin. No N - sulfocarbamoyl, decarba- moyl, or deoxy-decarbamoyl toxins were detected. In contrast, less potent N - sulfocarbamoyl toxins domi- nate the toxin profiles of all other global populations examined to date (from Australia, China, Japan, New Zealand, the Philippines, Portugal, Spain, and Uru- guay). The lack of genetic diversity found among broadly distributed populations of G. catenatum is consistent with the hypothesis of a relatively recent global spread of this species. Yet the unique toxin profile of Singapore strains indicates that it is un- likely that this strain has been recently translocated from any of the populations with characterized toxin profiles. In any case, the unique carbamate-dominated toxin profile may be a useful signature to identify the potential spread of this strain from the port of Sin- gapore, one of the world’s busiest. Key index words: ballast water; dinoflagellate; Gymno- dinium catenatum; mass spectrometry; paralytic shell- fish poisoning; rDNA; ribosomal RNA gene; toxin Abbreviations: dc, decarbamoyl; FD, fluorescence de- tection; GTX, gonyautoxin; HILIC, hydrophilic inter- action liquid chromatography; LSU, large subunit; STX, saxitoxin; MS, mass spectrometry; NEO, neosax- itoxin; PSP, paralytic shellfish poisoning; SRM, se- lected reaction monitoring Gymnodinium catenatum H.W.Graham is a chain-form- ing, toxic, unarmored dinoflagellate reported with in- creasing frequency from temperate to tropical marine waters around the world (Hallegraeff and Fraga 1998). In nature, G. catenatum is commonly observed as chains of 8–16 cells and occasionally up to 64 cells. As part of its sexual life cycle, G. catenatum produces benthic resting cysts (hypnozygotes) (Bravo 1986, Anderson et al. 1988). These microreticulate cysts were originally believed to be unique, but morphologically similar cysts are now known to be produced by two nontoxic non-chain forming spe- cies, G. nolleri Ellegaard et Moestrup and G. microreticu- latum Bolch et Hallegr. (Ellegaard and Oshima 1998, Bolch et al. 1999a). Resting cysts of G. catenatum are highly resistant to environmental stress and are thought 1 Received 14 August 2001. Accepted 7 November 2001. 2 Author for correspondence: e-mail dbshmj@nus.edu.sg.