Microbial contamination of drinking water is widespread in many resource-limited environments, but existing microbial tests require 18-48 h of incubation to detect coliform indicator bacteria at the required WHO standard of no detectable culture-forming units in a 100 mL sample. Rapid yet sensitive tests would allow users to test their water quality to select the appropriate treatment, and also enable the incorporation of point-of-use water testing into awareness campaigns for sanitation and water treatment. In our field interviews and workshops with end users in rural India, we identified interest in same-day point-of-use water testing and a strong interest in sensors that would provide the level of contamination and recommend an action, but there were also concerns about toxicity and handling of any reagents that such tests may involve. Based on these considerations, we identified electrochemical sensing of enzymatic activity of E. coli as a potential inexpensive method to transduce the level of bacterial contamination into an electrical signal which can provide integrated reporting of contamination and recommended actions. To maximize sensitivity and specificity to the indicator bacteria, we selected Chlorophenol red-β-D-galactopyranoside (CPRG) as a substrate that is cleaved by β-Galactosidase to form redox-active Chlorophenol red (CPR), which is less toxic than other common β-Galactosidase substrate products and can be electrochemically detected at the nanomolar level using Copper(II) Phthalocyanine-functionalized carbon electrodes (Figs. 1a and 1b). To enable the rational design of a point-of-use bacterial sensor, we evaluated the kinetics of substrate turnover (Figs. 1c and 1d) and observed that an induction time of approximately 3 h is required for maximum production of CPR by unpermeabilized cells in the presence of the inducer Isopropyl β-D-1-thiogalactopyranoside (IPTG). This investigation of in vivo enzyme kinetics and electrochemical CPR detection informs the development of improved point-of-use bacterial sensors for drinking water.

Electrochemical detection of E. coli enzyme activity for improved point-of-use bacterial water testing in resource-limited environments

255th National Meeting (2018)

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