UC San Diego

Domoic acid (DA), the causative agent of amnesic shellfish poisoning in humans, is a potent neurotoxin produced by algal blooms of the diatom genus Pseudo-nitzschia. Due to its bioaccumulation in shellfish, DA production must be closely monitored in the marine environment. Despite extensive research on Pseudo-nitzschia toxicity, the basic biology underlying DA production has remained unclear. Specifically, a genetically encoded biosynthesis for DA has eluded chemists and algal bloom researchers. Such genetic information would provide a genetic basis for monitoring toxin production in the environment. This dissertation describes the discovery of the DA biosynthesis (dab) genes and subsequent application to environmental monitoring. Chapter 2 details the initial discovery of the dab genes in Pseudo-nitzschia multiseries using comparative transcriptomics to identify candidate genes upregulated under DA-inducing culturing conditions. Subsequent validation of enzyme activity in vitro establishes the chemical transformations required to build the key structural features of DA. Chapter 3 describes the identification of dab gene homologues in the red macroalgae Chondria armata, a DA-producing seaweed from which the toxin was first isolated. The red algal DA (rad) biosynthesis genes uncovered here suggest a biosynthetic pathway consistent with the Pseudo-nitzschia pathway with only slight modifications to enzyme activity in vitro. Similarities and differences between the rad genes and diatom dab genes suggest a complex evolutionary history for toxin production in the two distantly related organisms. Finally, Chapter 4 describes a molecular approach to Pseudo-nitzschia and DA monitoring, focusing on the 2015 North American West Coast Pseudo-nitzschia australis bloom. In this study, RNA barcoding and metatranscriptomics datasets were generated from weekly phytoplankton net tow samples taken from Monterey Bay, California, a hotspot for DA production during the 2015 bloom. Sequence barcoding improved description of Pseudo-nitzschia species composition in the context of the larger microbial community. Meanwhile, metatranscriptomics enabled the identification of dab transcripts simultaneous with detection of DA and toxic species, and also provided insight into the shifting physiology of the P. australis bloom throughout its lifespan. The combination of efforts describes herein establishes a paradigm for the genetic monitoring of DA producing algal blooms.