Permeabilization-free β-galactosidase-induction-based electrochemical detection of Escherichia coli

Escherichia coli (E. coli) is recognized as an indicator of fecal bacterial contamination, and the induction of intracellular beta-galactosidase (Gal) is commonly used for sensitive and selective E. coli detection. However, this method requires a permeabilization process to ensure that the Gal substrate and product are highly permeable through the E. coli membrane, making it unsuitable for simple point-of-care detection. Here, we report a sensitive electrochemical method for E. coli detection that does not require a permeabilization process. Intracellular Gal expression is increased via induction by isopropyl-beta-D-thiogalactopyranoside (IPTG). 4-Methoxyphenyl-beta-D-galactopyranoside (MPGP) and 4-methoxyphenol (MP) are used as the highly cell-permeable Gal substrate and product, respectively. Externally-added tyrosinase (Tyr) converts MP into electrochemically active 4-methoxycatechol, which is then electrochemically oxidized via electrochemical-chemical redox cycling involving an electrode, 4-methoxycatechol, and tris(2-carboxyethyl)phosphine. The presence of MPGP during the induction of intracellular Gal led to amplified MP production. The results of four different detection methods using three Gal substrates (MPGP, phenyl-beta-D-galactopyranoside, and 4-aminophenyl-beta-D-galactopyranoside) were compared. The detection limit for both drinking water and tap water spiked with E. coli, obtained following a 2-h-long IPTG treatment and 5-min-long Tyr reaction, was similar to 2 x 10(3) colony-forming units (CFU)/mL, indicating that this method can be applied to monitoring E. coli contamination in water samples. Importantly, the present method does not require the electrode modification, affinity binding, washing, and filtration steps.

Automated summary

A new electrochemical method for detecting E. coli was developed in this study, which does not require a permeabilization process. The presence of MPGP during induction of Gal expression led to increased production of MP. The detection limit of this method in both drinking water and tap water was similar to 2 x 10(3) CFU/mL.