Development of Real-Time Instrumentation for the Robotic Detection of Paralytic Shellfish Poisoning (PSP) Toxins in Massachusetts Coastal Waters

In support of MIT Sea Grant's goal of a safe and sustainable seafood supply, the PI proposes to adapt an instrument known as the Environmental Sample Processor (ESP) to robotically detect and quantify the presence of Paralytic Shellfish Poisoning (PSP) toxins, in situ and in real time. The instrument will be field-tested in the Nauset Marsh area on Cape Cod. Historically, PSP has posed a significant health threat, and the economic impact of shellfish bed closures during red tide is ongoing. This new technology will provide much more accurate assessment and prediction capabilities to regulatory agencies, shellfishermen, aquaculturists, and the general public.

Objectives: To aid in the elucidation of bloom dynamics of harmful algal bloom (HAB) species, including the toxic dinoflagellate Alexandrium fundyense, a novel instrument known as the Environmental Sample Processor (ESP) was developed to quantitate real-time in situ cell abundances. While this was a necessary first step, the instrumentation needs to be adapted to quantify Paralytic Shellfish Poisoning (PSP) toxins in phytoplankton if the goal of a safe and sustainable seafood supply is to be realized in MA coastal waters. This project, therefore, proposes to (1) develop an ESP module that will robotically detect in situ concentrations of PSP toxins in phytoplankton in real time; and (2) field test the instrument in the Nauset Marsh System (NMS), Cape Cod, MA. During field testing of the ESP, we propose to conduct a field experiment that will utilize current shellfish monitoring programs to (3) determine the relationship between shellfish toxicity and ambient A. fundyense cell abundances and toxin content in the NMS.

Methodology: A competitive enzyme-linked immunosorbent assay (cELISA) for the quantitation of PSP toxins will be adapted for the ESP according to a protocol developed by Co-PI Doucette for another algal toxin, domoic acid. A variety of solvents and temperatures will be used to extract PSP toxins from field material until an acceptable recovery (≥ 90%) is reached. Arrays and assays will be developed using currently available antibodies, or new immunoreagents with improved cross-reactivities as they become available. Validation of the cELISA module will be done in the laboratories of PI Anderson and Co-PI Doucette using A. fundyense cultures and field material; paired samples will be quantified by the ESP and standard analytical methods (e.g., HPLC-FLD). New operational script will be written and tested for the enhanced ESP. The ESP will be field tested for three weeks in Mill Pond, (Orleans MA) using a modification of a deep-water mooring system. Discrete water samples will be processed daily by the ESP for A. fundyense cell abundance and toxin content and compared to independent water samples collected twice a week from the mooring site. Samples will be analyzed for toxin content (HPLC-FLD and cELISA) and cell abundance (whole cell and/or sandwich hybridization assay). The ESP will then be transported to Salt Pond (Eastham,MA) where it will be housed in a shore-based shed, collecting samples daily for six weeks. Shellfish (i.e., Mytilus edulis) will be collected weekly, except for one week, when shellfish will be collected daily. Shellfish toxicity will be analyzed by the MA Division of Marine Fisheries (DMF). During the one-week intensive study, shellfish will be sampled from both the natural population and nylon bags of clean shellfish which will be deployed for one day or one week, depending on the treatment. Weekly and daily shellfish toxicity data and daily ESP values will be statistically analyzed to determine relationships and if shellfish toxicity reflects short-term fluctuations in ambient toxin content.

Rationale: The coastal waters of New England are subject to recurrent outbreaks of Paralytic Shellfish Poisonings (PSP) caused by the toxic dinoflagellate A. fundyense. PSP is the most widespread of all HAB poisoning syndromes. Economic impacts are significant – i.e., the losses from a single red tide in 2005 cost the Massachusetts Shellfish Industry $50M. Here we propose a study that will develop instrumentation to be incorporated into the ESP to provide real-time, in situ data on A. fundyense toxicity within the NMS, a critical part of the Cape Cod National Seashore that experiences annual closures of its recreational and commercial shellfishing due to PSP toxicity. Resources will also be allocated to determining the relationship between ESP data (i.e., A. fundyense cell abundance and toxin content) and shellfish toxicity on weekly and daily time scales. This information is of value to managers and public health officials, as more accurate, frequent data regarding bloom dynamics and toxicity within the system will lead to quicker response time regarding regulatory actions (e.g., closing and opening shellfish beds), as well as improved early warning and forecasting capabilities. This study fits within the strategic goals of MIT Sea Grant for the management of a safe and sustainable seafood supply by conducting research that will assist managers to formulate science-based regulations and providing them with new technology that improves assessment and prediction accuracy. Other beneficiaries would include shellfishermen, aquaculturists, tourists, journalists, and the general public, all of whom are eager for up-to-date information during regional red tide outbreaks and related shellfish bed closures.