Validation of a cell-based assay for detection of active Shiga toxins produced by Escherichia coli in water

Authors: Hughes, Anna; PATFIELD, STEPHANIE - Former ARS Employee; Rasooly, Reuven; He, Xiaohua

Submitted to: International Journal of Environmental Research and Public Health
Publication Type: Peer Reviewed Journal
Publication Acceptance Date: 10/20/2020
Publication Date: 10/28/2020
Citation: Hughes, A.C., Patfield, S., Rasooly, R., He, X. 2020. Validation of a cell-based assay for detection of active Shiga toxins produced by Escherichia coli in water. International Journal of Environmental Research and Public Health. 17(21):7901. https://doi.org/10.3390/ijerph17217901.
DOI: https://doi.org/10.3390/ijerph17217901

Interpretive Summary: The current standards of water quality have eliminated massive outbreaks of waterborne disease, but cases of gastrointestinal illnesses were reported among individuals drinking tap water. Shiga toxin-producing E. coli (STEC) is one of the major bacteria that cause human intestinal diseases. There are many sources of STEC infection, including water pond contaminated with animal feces. The purpose of this study was to determine if a rapid assay could detect low dose of STEC present in environmental water. We proved that a HeLa cell assay is rapid, inexpensive, and sensitive. Less than 0.4 cfu/mL of STEC in water could be detected and common water additives, such as chlorine and fluoride, or background flora present in pond water, did not appear to interfere with the detection or cause any growth competition with the low dose of STEC inoculated in the water. This HeLa cell assay holds promise for identifying live STEC present in environmental water.

Technical Abstract: Shiga toxin-producing E. coli (STEC) causes a wide spectrum of disease, including hemorrhagic colitis and hemolytic uremic syndrome (HUS). Almost 5% of STEC infections result from Water-borne exposures, yet there is no test listed in the current EPA’s Selected Analytical Methods for the detection of active Shiga toxins (Stxs) in water. In this study, a HeLa cell-based assay was validated for the detection of metabolically active Stxs produced by STEC in water, including tap, bottled, and pond water. This assay was able to detect active Stxs from less than 0.4 CFU/mL of STEC in water and the assay performance was not affected by background flora or chlorine (up to 3 ppm) in water. This assay is not only as simple, and affordable as cell-free assays, but also detects active holotoxins without the use of live animals. In addition, the assay was designed for use in multi-well formats, making it ideal for high-throughput screening of water samples, thus useful for environmental public health surveillance programs to reduce human risk of infection with STEC.