Evaluation of Quantitative Real Time PCR (qPCR) for the Measurement of E. coli in Well Water Samples

Home / Research / Evaluation of Quantitative Real Time PCR (qPCR) for the Measurement of E. coli in Well Water Samples
Project Number:


Funding Year:


Contract Period:

7/1/15 - 6/30/16

Funding Source:


  • Greg Kleinheinz, UW-Oshkosh
  • Nilay Sheth, UW-Oshkosh

Culture-based microbiology methods have been used to determine potability of drinking water since the beginning of the 20th century. The detection of coliform bacteria in well water is an indicator of potential human fecal contamination and therefore the possible presence of disease causing organisms. These culture-based methods are widely accepted because of their relative ease of use, low cost, and demonstrated relationship to health risk. However, the time required for sample analysis range from 18 to 48 hours or even longer for confirmation steps. In cases where contamination occurs a more rapid method for analysis would be more protective of public health. Additionally, after wells have been disinfected a rapid method would allow for quicker follow-up testing results to save business owners money by decreasing the time bottled water is served or operations are suspended. A rapid method, such as quantitative real-time PCR (qPCR), is intended to shorten the period between sampling and publicly available results, with the goal of having same day water quality information. The US EPA recently developed several qPCR methods for a reliable real-time assessment tool for water analysis (US EPA 2014). These have received approval for use in surface water applications and this project proposes to evaluate one of these qPCR methods for monitoring well water. By using qPCR, the results can be obtained in 3-4 hours, which allows for same-day notification to public and private clients.

The overall goal of this project is to evaluate the relationship between US EPA qPCR Method C and EPA approved culture-based methods for detection of microbial contaminants in well water samples. Method C was chosen as this is approved by EPA for recreational water in some cases and tests for E.coli. Other approved qPCR methods for recreational water test for enterococci. There is currently no approved qPCR coliform test. Over a 16 week study period well water samples were collected from East Central Wisconsin (Kewaunee and Door County) and analyzed using qPCR and culture-based microbial enumeration methods. This study will evaluate the relationship between qPCR and traditional culture based methods for detecting coliforms and E. coli in well water. This project filled a key gap in knowledge by adapting existing rapid surface water microbial detection techniques to well water systems which will result in faster monitoring results that will increase our ability to protect public health, decrease economic impact of positive well results on business owners, and increase sensitivity of our monitoring techniques to match those available for surface waters.

Project Reports: