Abstract
Saxitoxin (STX) is a secondary metabolite and potent neurotoxin produced by several genera of harmful algal bloom (HAB) marine dinoflagellates. The basis for variability in STX production within natural bloom populations is undefined as both toxic and non-toxic strains (of the same species) have been isolated from the same geographic locations. Pyrodinium bahamense is a STX-producing bioluminescent dinoflagellate that blooms along the east coast of Florida as well as the bioluminescent bays in Puerto Rico (PR), though no toxicity reports exist for PR populations. The core genes in the dinoflagellate STX biosynthetic pathway have been identified, and the sxtA4 gene is essential for toxin production. Using sxtA4 as a molecular proxy for the genetic capacity of STX production, we examined sxtA4+ and sxtA4- genotype frequency at the single cell level in P. bahamense populations from different locations in the Indian River Lagoon (IRL), FL, and Mosquito Bay (MB), a bioluminescent bay in PR. Multiplex PCR was performed on individual cells with Pyrodinium-specific primers targeting the 18S rRNA gene and sxtA4. The results reveal that within discrete natural populations of P. bahamense, both sxtA4+ and sxtA4- genotypes occur, and the sxtA4+ genotype dominates. In the IRL, the frequency of the sxtA4+ genotype ranged from ca. 80–100%. In MB, sxtA4+ genotype frequency ranged from ca 40–66%. To assess the extent of sxtA4 variation within individual cells, sxtA4 amplicons from single cells representative of the different sampling sites were cloned and sequenced. Overall, two variants were consistently obtained, one of which is likely a pseudogene based on alignment with cDNA sequences. These are the first data demonstrating the existence of both genotypes in natural P. bahamense sub-populations, as well as sxtA4 presence in P. bahamense from PR. These results provide insights on underlying genetic factors influencing the potential for toxin variability among natural sub-populations of HAB species and highlight the need to study the genetic diversity within HAB sub-populations at a fine level in order to identify the molecular mechanisms driving HAB evolution.
Highlights
These results provide insights on underlying genetic factors influencing the potential for toxin variability among natural sub-populations of harmful algal bloom (HAB) species and highlight the need to study the genetic diversity within HAB sub-populations at a fine level in order to identify the molecular mechanisms driving HAB evolution
Saxitoxin is a secondary metabolite produced by several genera of dinoflagellates and cyanobacteria that is better known as a potent neurotoxin due to its detrimental effects on human health
This is in keeping with the trend recorded for Alexandrium, in that the overall percent similarity was higher for cDNA copies than gDNA copies, indicating that only certain copies are transcribed [14,19]
Summary
Saxitoxin is a secondary metabolite produced by several genera of dinoflagellates and cyanobacteria that is better known as a potent neurotoxin due to its detrimental effects on human health. The collective findings with microsatellite markers that monospecific dinoflagellate blooms are not monoclonal events coupled with the isolation of both toxic and non-toxic strains of the same species from similar geographic regions underscores the need to examine the genetic capacity for toxin production as a contributor to toxin variability within bloom sub-populations. A large gap in our understanding is the factors influencing toxin variability among natural dinoflagellate bloom populations, as both toxic and non-toxic strains of the same species have been isolated from the same geographic locations [31,40]. While the core genes in the dinoflagellate STX biosynthetic pathway are known [14,15], how this relates to variability in STX production within natural populations from the same geographic area remains unknown. Our data reveal that both sxtA4+ and sxtA4- genotypes occur within the same sub-population and that frequency of the sxtA4+ genotype dominates in Florida populations
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