Comparative Analysis of Quantitative Detection Methods for Enumeration of HAB Species: Applications for Resource Management
Project Status: This project began in January 2010 and was completed in December 2014
We are providing a thorough comparison of two different genetic methods used to quickly count the number of harmful algae present in a water sample. Our results will improve harmful algae monitoring and research, both in the laboratory and in the field.
Why We Care
Multiple research initiatives have contributed to the development and validation of harmful algal bloom (HAB) detection and quantification technologies in laboratory and natural settings. Despite these advances, fundamental questions still remain unanswered, such as:
Will each technique provide accurate and comparative results?
Do cell growth and nutrient conditions affect the tests?
Does calibration using laboratory cultures translate to accurate estimates of natural field populations?
From a resource management standpoint, which quantification technique is most suitable for a particular monitoring need and budget?
Traditionally, planktonic microscopic algae have been counted and identified using a light microscope. More recently, light microscopy has been supplemented with and often replaced by molecular approaches for quantifying (counting) algal species. Rapid detection and counting of HAB species is important for economic and public health reasons. Cell detection based on light microscopy can be tedious and time-consuming when large numbers of samples need to be processed. Also, identification of some HAB species may require highly trained personnel.
Newer genetic methods—such as quantitative real-time polymerase chain reaction (QPCR) and sandwich hybridization (SHA)—meet the need for faster sample processing that is also accurate and precise. Both methods have been validated for HAB species quantification. However, they have not been thoroughly compared, representing a key gap in the ability to provide recommendations to managers for a specific regulatory requirement. Through this project, we are providing a thorough comparison and assessment of QPCR and SHA for HAB monitoring and research.
What We Are Doing
Our results will be applicable to several HAB species, but the model species for this study is the globally-distributed, fish-killing alga Heterosigma akashiwo. The accurate enumeration of H. akashiwo in natural samples can be difficult due to its fragile structure and small size. Our objectives are to:
Directly compare QPCR and SHA for quantification of H. akashiwo isolates spanning a range of cell abundances, growth phases, and nutrient conditions;
Determine the extent to which quantification of H. akashiwo is comparable using QPCR and SHA for natural phytoplankton communities; and
Synthesize comparisons according to a suite of criteria to enhance HAB monitoring and research activities.
In this targeted study, QPCR and SHA will be compared (using light microscopy as the “gold standard”) to provide recommendations to managers for HAB monitoring strategies based upon multiple criteria, including:
Range and limit of detection.
Accuracy and specificity.
Cost per sample.
Initial investment and equipment maintenance cost.
Applicability to live and preserved samples.
Circumstances when one method would be preferable over another. Comparisons will focus on H. akashiwo representing a range of geographic regions (e.g., South Carolina, Delware, Washington, Alabama, and Rhode Island), cell growth phases, and nutritional conditions in laboratory and field studies.
This study is led by Dr. Dianne Greenfield (University of South Carolina's Algal Ecology Laboratory), with assistance from Dr. Kathryn Coyne (University of Delaware's Microbial Ecology Research Laboratory). The project is part of the NCCOS Monitoring and Event Response for Harmful Algal Blooms (MERHAB) program.
Benefits of Our Work
Resource managers will be provided with the necessary tools to make informed decisions about appropriate method(s) for individual HAB monitoring needs and budgets. We will convey this information through workshops, a website, publications, and scientific presentations. Expected outcomes include: improved knowledge for management decisions, and changes in management/monitoring approaches as method(s) are incorporated into HAB monitoring programs.
Regions of Study: Delaware, South Carolina
Primary Contact: Marc Suddleson
Harmful Algal Blooms (Monitoring and Event Response)
Related NCCOS Center: CSCOR
- Main, Christopher R., Cameron Doll, Colleen Bianco, Diane I. Greenfield, and Kathryn J. Coyne. 2014. Effects of growth phase, diel cycle and macronutrient stress on the quantification of Heterosigma akashiwo using qPCR and SHA. Harmful Algae 37: 92–99. DOI: 10.1016/j.hal.2014.05.014
- Doll, Cameron, Christopher R. Main, Colleen Bianco, Kathryn J. Coyne, and Diane I. Greenfield. 2014. Comparison of sandwich hybridization assay and quantitative PCR for the quantification of live and preserved cultures of Heterosigma akashiwo (Raphidophyceae). Limnology and Oceanography: Methods. 12: 232–245. DOI: 10.4319/lom.2014.12.232
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