2003 Progress Report: Linking Population and Physiological Diversity in a Toxin-producing DinoflagellateEPA Grant Number: R830413
Title: Linking Population and Physiological Diversity in a Toxin-producing Dinoflagellate
Investigators: Campbell, Lisa , Gold, John R.
Institution: Texas A & M University
EPA Project Officer: Hiscock, Michael
Project Period: September 1, 2002 through August 31, 2005 (Extended to February 28, 2007)
Project Period Covered by this Report: September 1, 2002 through August 31, 2003
Project Amount: $464,880
RFA: Ecology and Oceanography of Harmful Algal Blooms (2002) RFA Text | Recipients Lists
Research Category: Water Quality , Ecosystems , Water , Aquatic Ecosystems
The main objective of this research project is to gain understanding of the dynamics of blooms of Karenia brevis, a toxic dinoflagellate, in the Gulf of Mexico. Preliminary results have demonstrated that clonal cultures of K. brevis can have significantly different rates of growth and levels of toxin production when grown under identical conditions. Our aim is to develop new hypervariable molecular markers (microsatellites) for K. brevis to link population and physiological diversity in K. brevis. Ultimately, information will be used to predict the response of K. brevis at the population level to environmental changes, and to assess how these responses affect and influence bloom formation and dynamics. The specific objectives of this project are to: (1) optimize a suite of hypervariable, nuclear-encoded DNA markers (microsatellites) to characterize genetic diversity among strains of the toxic dinoflagellate K. brevis; (2) isolate and establish clonal cultures of K. brevis sampled during the onset, bloom, and decline of a toxic algal bloom event; and (3) utilize the highly polymorphic molecular markers to elucidate population structure.
A library of 480 microsatellite-enriched clones was constructed from DNA derived from the SP2 isolate of K. brevis from a bloom that occurred in the Fall of 1999, near Brownsville, Texas. Twenty-one clones had sufficient numbers of repeats to potentially produce sufficiently variable microsatellite markers. Six clones did not have sufficient flanking regions for primer development. Primer pairs were created and optimized for the remaining 15 loci using the OLIGO software package. Of these 15 potential microsatellite loci, 2 primer pairs produced banding patterns concordant with single-copy microsatellites using the SP2 isolate DNA sample. Four additional loci appear to have characteristics of Random Amplifications of Polymorphic DNA (RAPDs), and are able to positively differentiate between and among samples of K. brevis and Karenia mikimotoi. Four loci have not yet been optimized, and five remaining potential microsatellite loci did not meet the requirements for optimization. As a result, they were discarded. Six additional potential microsatellites are in the process of rescreening, sequence analysis, and primer design. Twenty-two new clonal cultures of K. mikimotoi cultures were established from a bloom in Corpus Christi Bay, January 2002. Preliminary results have shown that a number of markers appear to be useful for fingerprinting clones of both K. brevis and K. mikimotoi. The capability to positively differentiate both between and among individual K. brevis and K. mikimotoi clones is significant, and represents a potentially extremely useful tool. Because of the difficulty for routine identification of these strains, along with the apparent variation in toxicity among strains/clones within each of these species, the markers we have identified may be applicable for the development of a simple fingerprinting assay for routine use by managers.
We will isolate and establish clonal cultures of K. brevis sampled during the onset, bloom, and decline of a toxic algal bloom event, and we will assess genetic and physiological variability within a bloom population. We will determine allele distributions at 10-15 (independent) microsatellites among spatial and temporal isolates of K. brevis, including clones currently in culture. We also will determine growth rates for genetically distinct clones to assess physiological variability within and among K. brevis haplotypes.