Monitoring for Heterosigma akashiwo using a sandwich hybridization assay

• Published in: Harmful algae 2002-06, Vol.1 (2), p.205-214
• Main Authors: Tyrrell, John V, Connell, Laurie B, Scholin, Chris A
• Format: Article
• Language: English
• Subjects: Heterosigma akashiwo; Marine; Molecular probes; Puget Sound; Raphidophyceae; rRNA; Sandwich hybridization assay
• ISSN: 1568-9883; 1878-1470
• DOI: 10.1016/S1568-9883(02)00012-4

Field testing of a ribosomal RNA (rRNA)-targeted sandwich hybridization assay (SHA) for Heterosigma akashiwo (Raphidophyceae) in Puget Sound, WA, USA, has showed that the lower limit of detection is well below the level at which cells pose a danger to fish. Moreover, the assay has proven to be both rapid and easy-to-use. Isolates of H. akashiwo from Australia, Japan, New Zealand, South Korea, Spain and USA were correctly identified using the SHA, indicating that this diagnostic tool could be deployed globally. Samples containing H. akashiwo can be preserved for subsequent SHA analysis using several methods: fixation with acidic Lugol’s iodine followed by room temperature storage, collection onto Durapore filters followed by storage at −70 °C or, alternatively, the filters are mixed with a lysis solution buffer and the sample lysate stored at −70 °C. Additionally, we sought to determine whether the SHA could successfully detect H. akashiwo in the presence of clay that might some day be used to mitigate the impacts of natural H. akashiwo blooms. Results from preliminary laboratory trials indicate that clay at the maximum proposed dosage rate does not interfere with the assay. Thus, it may be possible to use the SHA as a simple means of following the fate of H. akashiwo cells during larger-scale clay mitigation trials.