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Project: Alpha-Testing of Rapid Microbiological Methods for Measuring Recreational Water Quality

Background and Objectives

In California, public health officials conduct regular beach monitoring to safeguard the public against contact with contaminated recreational waters. The methods approved by the US Environmental Protection Agency for enumerating indicator bacteria require the samples to be incubated for 18 to 96 hours, though, leaving a window when beach water quality is in question. Scientists are working toward developing new methods that would require less than four hours to obtain results, thus allowing officials to sample a beach and post any necessary warnings or closures on the same day the sample is taken. Reducing the window for time needed for sample processing not only improves public health protection but also permits tracking of contamination sources. Because several scientific research groups around the country are working on developing rapid methods for microbiological monitoring, the methods under development require rigorous independent testing. Among other research activities, SCCWRP conducted several objective trials to evaluate and advance the science of rapid microbiological methods for application to beach monitoring in Southern California.

The goal of this project was to evaluate the performance of several rapid microbiological methods as applied by the method developers to a standard set of natural and laboratory created samples exemplifying ambient shoreline conditions in Southern California. The objectives were to determine which rapid methods demonstrated the best equivalency with existing measurement methods, characterize the risk of false positives or false negatives for each of the methods, and identify factors that would recommend or impair their application to beach monitoring.


This study was completed in 2005.


Two trials were undertaken in 2004 and 2005 to evaluate performance of various rapid methods as applied by method developers. Identical samples were prepared by SCCWRP and shipped to each of the participating laboratories. Four methods were included in the 2004 trial: immunomagnetic separation coupled with ATP bioluminescence (IMS-ATP), flow cytometry, quantitative polymerase chain reaction (QPCR), and dual-wavelength fluorimetry (DWF). Two additional permutations of the QPCR method were added to the 2005 trial, as well as Transcription-mediated Amplification (TMA) and an immunological dipstick method . Most of the methods focused on measuring enterococci, though some were also used to measure E. coli. In both trials, five southern California laboratories processed the same samples using traditional methods (chromogenic substrate (CS) and membrane filtration (MF)) for comparison. In total, 18 blind samples were processed in triplicate over the course of three days, including both natural and laboratory-created samples across a range of concentrations, matrices, and interferences.


In the 2004 trial, none of the rapid methods performed well enough to replace existing methods, but two of them showed encouraging results. QPCR produced results within the range of the reference methods for two-thirds of the samples, but overestimated others, particularly those with complex matrices. DWF had the best precision among the new methods, but also produced results that were generally higher than existing methods, especially for samples that contained urban runoff. When beach warning decisions based on data from the rapid methods was compared to the decision based on the median result from the reference laboratories, QPCR showed the best agreement, though it tended to produce a false positive result.

Percentage of false positives and false negatives for each method tested in the 2004 trial with respect to California's standard beach warning threshold of 104 cfu/ 100mL.

In the 2005 trial, results from two of the QPCR methods and the TMA method were more than 80% accurate with respect to the State standard for enterococci. They also concurred with beach management decisions based on EPA-approved method results for more than 75% of samples. Results for one of the E. coli QPCR methods showed 90% agreement in terms of beach management decisions, which is comparable to the rate of agreement between the two approved traditional methods. Based on these trials, the California Beach Water Quality Workgroup the workgroup identified a need for additional beta-testing by personnel from local laboratories using a larger number of ambient samples.


This study was conducted in collaboration with EPA Region I, EPA National Exposure Research Laboratory, MLT/Gen-Probe, Silverlake Research, Rosewood Industries, the University of North Carolina at Chapel Hill Institute of Marine Sciences, Cepheid Inc, Orange County Sanitation District, the City of Los Angeles Environmental Monitoring Division, City of San Diego, Los Angeles County Sanitation Districts, Orange County Public Health Laboratory, and Weston Solutions, with funding from the State Water Resources Control Board and the Cooperative Institute for Coastal and Estuarine Environmental Technology (CICEET).

For more information on Alpha-Testing of Rapid Methods, contact John Griffith at (714) 755-3228.
This page was last updated on: 7/2/2014