A DNA hybridization assay to identify toxic dinoflagellates in coastal waters: detection of Karenia brevis in the Rookery Bay National Estuarine Research Reserve |
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| Authors: | KD Goodwin SA Cotton G Scorzetti JW Fell |
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| Institution: | aNational Oceanographic and Atmospheric Administration, Atlantic Oceanographic and Meteorological Laboratories, Ocean Chemistry Division, 4301 Rickenbacker Causeway, Miami, FL 33019, USA;bCooperative Institute of Marine and Atmospheric Studies, Rosenstiel School of Marine and Atmospheric Science, University of Miami, 4600 Rickenbacker Causeway, Miami, FL 33019, USA;cMarine Biology and Fisheries, Rosenstiel School of Marine and Atmospheric Science, University of Miami, 4600 Rickenbacker Causeway, Miami, FL 33019, USA |
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| Abstract: | A DNA hybridization assay was developed in microtiter plate format to detect the presence of toxic dinoflagellates in coastal waters. Simultaneous detection of multiple species was demonstrated using Karenia brevis, Karenia mikimotoi, and Amphidinium carterae. Molecular probes were designed to detect both K. brevis and K. mikimotoi and to distinguish between these two closely related species. The assay was used to detect K. brevis in coastal waters collected from the Rookery Bay National Estuarine Research Reserve. Assay results were verified by species-specific PCR and sequence analysis. The presence/absence of K. brevis was consistent with microscopic observation. Assay sensitivity was sufficient to detect K. brevis in amounts defined by a regional monitoring program as “present” (≤1000 cells/L). The assay yielded quick colorimetric results, used a single hybridization temperature, and conserved the amount of genomic DNA utilized by employing one set of PCR primers. The microplate assay provides a useful tool to quickly screen large sample sets for multiple target organisms. |
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| Keywords: | Amphidinium carterae DNA hybridization Karenia brevis Karenia mikimotoi Molecular probes Rookery Bay NERR |
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