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Quality Assurance/Quality Control Manual: Ohio Water Microbiology Laboratory

Research Methods

Rapid methods for indicator bacteria
Traditional microbiological methods for detecting fecal-indicator bacteria and pathogenic organisms can take at least 18 hours to obtain results. Because water quality can change significantly during this timeframe, the safety of the water may not be accurately assessed. The need for rapid detection methods that provide reliable results of the current day’s water-quality conditions is widely recognized. The USGS Ohio Water Microbiology Laboratory (OWML) is currently using two rapid detection methods for the enumeration of E. coli and enterococci.

The immunomagnetic separation/adenosine triphosphate (IMS/ATP) rapid method requires approximately 2 hours from sample collection to availability of results (Bushon and others, 2007; Bushon and others, 2009; Lee and Deininger, 2004). Magnetic beads that are coated with antibodies for either Escherichia coli (E. coli) or enterococci are added to a water sample. This mixture is then subjected to IMS, in which the bacteria-antibody-bead complex is separated from extraneous materials in the sample by use of a strong magnet. Following several wash/concentration steps, the bacterial cells are ruptured by an enzymatic process, releasing ATP, which is the energy molecule found in living cells. The amount of ATP in the sample is measured with a microluminometer and results are reported in relative light units (RLUs).

The quantitative polymerase chain reaction (qPCR) method enumerates targeted genetic sequences within microorganisms in less than three hours. First, water samples are concentrated by passing through a 0.4 m filter. The concentrated organisms are lysed by both physical and chemical disruption and the genetic material that was contained in these organisms is then isolated and purified. The qPCR is run and the genetic sequence unique to the organism of interest is quantified by detecting the accumulation of a fluorescent probe. The OWML is currently applying qPCR assays for enterococci (U.S. Environmental Protection Agency, 2012a) and E. coli (Noble and others, 2010).