Submitted to: American Society for Microbiology Annual Meeting
Publication Type: Abstract Only
Publication Acceptance Date: May 1, 2003
Publication Date: May 1, 2003
Citation: Shelton, D.R., Van Kessel, J.S., Karns, J.S. 2003. Estimating Escherichia coli O157 populations in water samples using immunomagnetic ectrochemiluminescence. American Society for Microbiology Annual Meeting. p.96 Interpretive Summary: There is increasing concern regarding the impact of enterohemorrhagic Escherichia coli (e.g., E. coli O157:H7) on human health. Enterohemorrhagic E. coli poses a serious health threat, particularly to children, in North America, Europe, and Australia. It causes bloody diarrhea and, if not treated promptly, can result in kidney failure and death. There are an estimated 11,000 cases of water-borne enterohemorrhagic E. coli infections in the U.S. annually. One of the largest water-borne outbreaks of E. coli O157:H7 recently occurred in Walkerton, Ontario, Canada, with 167 confirmed infections. At present, the risk from water-borne transmission of E. coli O157 is difficult to estimate because traditional cultural methods are prohibitively laborious and expensive. The goal is to develop detection methods which are relatively rapid, accurate, sensitive, and quantitative. We have previously described a novel method for the quantitative detection of E. coli O157 in water samples using antibodies coupled to magnetic beads and a light producing catalyst (IM-ECL). The sensitivity of this method has been optimized by including an enrichment step (growth of E. coli O157 in a semi-selective broth) prior to detection. This method can be used to estimate E. coli O157 levels in 100 mL of water with a limit of detection of one E. coli O157 cell.
Technical Abstract: Enterohemorrhagic Escherichia coli (e.g., E. coli O157:H7) has emerged as a serious gastrointestinal pathogen in developed countries. One of the largest water-borne outbreaks of E. coli O157:H7 recently occurred in Walkerton, Ontario, Canada, with 167 confirmed infections. Traditional methods for detection of viable enterohemorrhagic E. coli rely on enrichment and plating followed by biochemical and serological characterization. Such methods are not only time-consuming and laborious, they lack sensitivity and quantitation. Consequently, detection methods are needed which are relatively, rapid, accurate (few false positives), sensitive (few false negatives), and quantitative (to establish exposure levels). We report here on the use of immunomagnetic (IM) electrochemiluminescence (ECL), a sandwich antibody assay, for estimation of E. coli O157:H7 in water samples following enrichment using a minimal lactose broth (MLB). In-house IM beads for cell capture were prepared using commercial anti-O157 monoclonal antibodies, while a polyclonal anti-O157 antibody was used for detection. The dynamic linear range of IM-ECL was ca.102 to 105 cells mL-1 E. coli O157 in MLB. There was considerable variability in recovery of E. coli O157 cells from enriched water samples; net ECL signals ranged from 1 to 100% of expected values (i.e., percent inhibition from 0 to 99%). However, competitive binding could be quantified by including a spiked control for each enriched water sample. Turbidity (i.e., total cell numbers) were measured for each MLB-enriched water sample, allowing for the fraction E. coli O157 to be calculated. Overall growth rates in MLB were 0.20±0.06; E. coli O157:H7 Odwalla growth rates were ca. 0.23 in MLB. In conclusion, we propose that E. coli O157 populations in water samples can be estimated by multiplying the fraction of E. coli O157 in enriched water samples by the total coliform population in raw water samples. Data indicate that as few as one viable E. coli O157 cell per 100 mL raw water can be detected.