By: Katherine Hubbard
Water samples for harmful algal bloom (HAB) monitoring are collected throughout the state by numerous partners and shipped to FWRI in St. Petersburg, where light microscopy is used to enumerate toxic species including the Florida red tide alga, Karenia brevis . These counts require expertise to discriminate K. brevis from morphologically similar, non-toxic species, and to provide critical information about the timing, extent and severity of the bloom for partners, managers and stakeholders.
Through funding from the NOAA Prevention Control and Mitigation of HABs (PCMHAB) program, researchers at FWC , the University of South Florida (USF), Mote Marine Laboratory and the Gulf of Mexico Coastal Ocean Observing System are working together to develop and test new hand-held genetic sensor technology to permit more timely and sensitive enumeration of K. brevis. The project team at USF developed this tool initially for the seafood industry – to differentiate grouper from other fish – and adapted it for HAB detection.
For the K. brevis assay, a seawater sample is collected, cells are concentrated and nucleic acids are extracted using a simplified,
field-friendly approach. The extracted sample is then added to a cocktail of reagents including an enzyme and a fluorescent probe specific to K. brevis. These assays are placed in a battery-operated sensor, dubbed the “tricorder,” that conducts isothermal amplification of a targeted gene. The sensor measures the increases in fluorescence signal as the K. brevis-specific fluorescent probe binds to target K. brevis genes in the extracted sample. The assay has a limit of detection of approximately 800 cells L-1, is completed within an hour and the instrument displays changes in signal as the assay is running allowing confirmation of cells in as little as 5-10 minutes.
An ongoing bloom of K. brevis has provided the project team with several opportunities to conduct field trials, which have been successfully completed in Sarasota Bay (shown in the images), at Fort de Soto State Park and during a 24-hour cruise onboard the R/V Bellows. Samples collected at shore sites were processed rapidly while still on site, where microscopic and genetic counts detected similar concentrations of K. brevis. Rough seas prevented sample analysis while on the R/V Bellows and samples were instead processed in the lab after the cruise; however, future efforts aim to successfully analyze samples while onboard large or small vessels to provide timely information about bloom distributions and inform adaptive sampling.
The project team is currently focused on simplifying and validating the K. brevis method further, and determining how variable gene copy number is across different strains of K. brevis and under different environmental conditions. This is a critical step in relating genetic and microscopic enumeration methods and automating genetic quantification, and in preparation for integrating this sensor technology into state and citizen monitoring programs in Florida and other states.