Project Brief

E. Coli is a commonly used indicator of sewage contamination at public swimming beaches.

In 2005, closing and advisory days at U.S. ocean, bay, and Great Lakes beaches topped 20,000. This project resulted in an assay that can be adapted to test for a variety of disease-causing organisms.

How’s the Water?
New DNA hybridization assay provides sensitive, species-specific way to test beach water quality.

Challenge
Every year, thousands of beaches around the U.S. are closed due to contamination by microbes such as fecal bacteria or harmful algae. Current testing methods involving cell culture and microscopy tools can take days to show results. Because levels of disease-causing microbes change much more rapidly than that, contaminated beaches often remain open when they should be closed, and clean beaches are often closed after the threat of contamination has washed away. To balance the protection of human and economic health, water quality managers need accurate, quick, and cost-effective approaches to pinpoint the type, level, and source of microbial contamination.

Approach
With funding from CICEET, researchers from the University of Miami and NOAA’s National Atlantic Oceanic and Meteorologic Laboratories have developed a DNA hybridization microplate assay that quickly and accurately identifies multiple species of toxic plankton and fecal indicator bacteria.

Researchers tested two groups of organisms: toxic dinoflagellates that cause “red tide,” and bacteria that indicate the presence of sewage. They created molecular probes that would bind to the DNA of these microorganisms, and attached the probes to the wells of microplates. DNA from a water sample was then placed in each well. If there was a match, a chemical reaction caused a yellow color to indicate the presence of that organism in the water sample.

The red tide assay successfully detected “very low” levels of Karenia brevis, a red tide organism common in Florida coastal waters. The sewage-indicating assay detected Escherichia coli, total coliforms, the Bacteroides fragilis group (anaerobic bacteria found in humans and animals), and Bacteroides distasonis (associated with the human gut). The Bacteroides assays are a promising way to distinguish between human and nonhuman sewage contamination in coastal waters.

Impact
This project resulted in a DNA hybridization microplate assay that provides sensitive, species-specific molecular analysis in a convenient, adaptable, and relatively inexpensive format. Water quality managers can adapt this technique to the molecular probes of their choice, or it can be used as an introduction to molecular methods for educational purposes.

The assay allows for easy sample storage and helps differentiate between human and non-human fecal pollution sources. It provides immediate visual results, is more specific and convenient than a series of species-specific PCR reactions, and is faster, easier, and less expensive than cloning. It is simple to add and adapt probes to the assay. The technique conserves the amount of genomic DNA utilized, which can be critical to certain applications.

The assay process and a video tutorial are available online: http://ciceet.unh.edu/project_extras/microplate_assay

Learn More
Dr. Jack Fell
University of Miami
T: 305.361-4603
E: jfell@rsmas.miami.edu

Dr. Kelly Goodwin
National Atlantic Oceanic and Meteorologic Laboratories
T: 305.361.4384
E. kelly.goodwin@noaa.gov

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