Electrochemical Detection of Escherichia coli in Aqueous Samples Using an Engineered Bacteriophage with β-galactosidase Gene

Introduction: Electrochemical detection of bacteria offers the advantages of instant quantification of pathogen in complex samples with minimal equipment. Bacteriophage has the high specificity and quick reproducibility which allows for the rapid and sensitive detection of bacterial pathogen. The use of the engineered phage with an enzyme gene provided opportunities to achieve a more sensitive detection: (1) release of the intracellular enzyme from Escherichia coli (E. coli); (2) overexpression of more enzyme in E.coli during the specific infection.

Purpose: An electrochemical method was developed for the detection of Escherichia coli (E. coli) using the engineered bacteriophage with β-galactosidase (β-gal) gene.

Methods: Here, the bacteriophage T7 which specifically targets E. coli BL21, was engineered to carry the β-gal gene. The bacteriophage after the bioengineering therefore has the ability to express β-gal during the infection of E. coli. The released and phage-induced β-gal was detected by electrochemical methods using 4-aminophenyl-β-galactopyranoside (PAPG) as a substrate. The β-gal catalyzed PAPG to an electroactive species p-aminophenol (PAP) which can be monitored on the electrode. The electrochemical signal is proportional to the concentration of E. coli.

Results: We demonstrated the application of our strategy in aqueous samples (drinking water and apple juice). Using this method, we were able to detect E. coli at the concentration of approximately 10⁵ CFU/ml in drinking water (or apple juice) in 3 hours, and 10² CFU/ml after 7 hours of incubation.

Significance: This method provides a specific and sensitive detection for E. coli in aqueous samples. It has the potential to be extended to detect other bacteria using different specific bacteriophage engineered with other enzyme genes.