Florida’s red tide, a harmful algal bloom caused by the dinoflagellate Karenia brevis, produces brevetoxins that can accumulate in shellfish and lead to neurotoxic shellfish poisoning in humans. The increasing frequency and duration of shellfish farm closures in the region due to toxic red tides have severely damaged southwest Florida’s shellfish industry and reduced the availability of shellfish for consumers. We aim to advance two innovative methods for measuring brevetoxins in seafood that will facilitate early detection, which can help mitigate the impacts of red tide on the shellfish industry.
Why We Care
Florida has long benefited from shellfish farming, with revenue to the state economy estimated at over $37 million. Shellfish harvest closures due to toxic blooms of K. brevis have severely impacted southwest Florida’s shellfish industry and reduced the availability of safe shellfish for consumers.
Shellfish growers need rapid, cost-effective end-product testing to mitigate the consequences of K. brevis blooms on the shellfish industry and reduce the financial and administrative burden carried by state and federal agencies that manage shellfish safety.
Current methods to test brevetoxins in shellfish include the time-consuming and labor-intensive mouse bioassay (MBA) and an enzyme-linked immunosorbent assay (ELISA) that has limited application for end-product testing. Preliminary data suggest that a new fluorescence aptamer assay (FAA) and enzyme-linked aptasorbent assay (ELASA) are potential alternatives to the ELISA for testing brevetoxins in shellfish. The FAA and ELASA have more attractive qualities for the end user, including assay cost, reduced time required to complete the assay, improved sample throughput, and ease of replenishing aptamers (single-stranded nucleic acid molecules that can bind to specific targets, such as proteins, with high affinity and specificity).
What We Are Doing
This project will evaluate FAA and ELASA method prototypes using shellfish tissue extracts from hard clams, sunray venus clams, and oysters provided by the Florida Fish and Wildlife Conservation Commission (FWC). Studies will demonstrate that the FAA and ELASA methods can detect both A- and B-type brevetoxins and can be used as alternatives to current methods used to determine brevetoxins in shellfish.
The objectives of the project are to:
- Evaluate the FAA and ELASA detection capabilities and their potential as screening methods for brevetoxins in shellfish.
- Determine assay cross-reactivity to 14 available brevetoxin reference standards.
- Validate the FAA and ELASA through spike-recovery and linearity of dilution testing.
- Rigorously test the two aptamer-based BTX detection methods to examine assay stability and ruggedness under various environmental and laboratory conditions.
- Analyze shellfish extracts using the FAA, ELASA, and National Shellfish Sanitation Program (NSSP) ELISA methods and correlate results to MBA data (provided by FWC).
- Analyze subsamples using an analytical method (LC-MS) in support of future screening methods that may be developed and compare and correlate all brevetoxin analysis data.
- Communicate study results to FWC for input to determine the suitability of the proposed assays as alternatives for the NSSP ELISA method within the U.S. shellfish sanitation regulatory framework.
Detection of brevetoxins using an indirect competitive enzyme-linked aptasorbent assay (ELASA).
Benefits of Our Work
Florida shellfish growers need rapid, improved end-product testing to measure brevetoxins in shellfish that correlate well with established regulatory protocols. The innovative technologies for brevetoxin detection tested here will help farmers make strategic harvesting and processing decisions that promote sustainable U.S. seafood production. High throughput, accurate, and less expensive brevetoxin testing with reduced analysis time will alleviate state testing burdens and reduce economic losses to the shellfish industry.
Dr. Dana Wetzel of the Mote Marine Laboratory leads this project, which is funded through the NCCOS Prevention, Control, and Mitigation of Harmful Algal Blooms (PCMHAB) Program.