Two electrochemical methods based on engineered bacteriophages were developed for the detection of <i>Escherichia coli </i>(E. coli). Electrochemical methods for bacteria detection offer the advantages of instant quantification with minimal equipment. The E. coli in food samples was quantified through the electrochemical measurement of reporter enzymes expressed from E. coli. In order to increase the enzyme level to achieve a more sensitive detection, we engineered bacteriophages (phages) with gene encoding for reporter enzymes. During the infection, phages facilitated the overexpression and release of the reporter enzymes. In this study, T7 phages, specific to E. coli, were engineered to express two enzymes β- galactosidase (β-gal) and gold binding peptide fused alkaline phosphatase (GBP-ALP), respectively. β-gal is an intracellular enzyme and widely used as an indicator enzyme of E. coli. GBP-ALP provided a novel way to immobilize the enzyme on the gold electrode, enabling the reaction to happen directly on the electrode. These two enzymes can be used for the electrochemical detection using different substrates and electrochemical models. The substrate of each enzyme was catalyzed by the enzyme to produce an electroactive compound that can be monitored on the electrode. We demonstrated the application of these two engineered phages for the detection of E. coli in aqueous samples. We were able to detect E. coli at the concentration of approximately 105 CFU/mL in 3 hours, and 102 CFU/mL after 7 hours. This strategy has the potential to be extended to detect different bacteria using specific phages engineered to express appropriate enzymes.

Engineered bacteriophage-based electrochemical detection of <i>Escherichia coli</i> in food samples

255th National Meeting (2018)

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Engineered bacteriophage-based electrochemical detection of Escherichia coli in food samples