Ultrasensitive Impedimetric Electrochemical DNA Biosensor for Detection of Enterotoxigenic <i>Escherichia coli</i> (ETEC)

Enterotoxigenic <i>Escherichia coli</i> (ETEC) is a pathogenic strain of <i>E. coli</i>, which is transmitted via food and water, primarily causing diarrhea in children below 5. Due to similar biochemical properties, conventional culture and biochemical identification methods are unsuitable for distinguishing ETEC. Immunoassay and molecular detection are alternative techniques for detecting ETEC, which target heat-labile (LT) and heat-stable (ST) enterotoxins. However, they may require specialized equipment, and some methods may require a pre-enrichment process, making them unsuitable for rapid and on-site detection. This study designs a DNA-based electrochemical biosensor, specifically for detecting the <i>heat-labile toxin</i> (<i>LT</i>) gene of ETEC. The screen-printed electrode (SPE) was functionalized with diazonium salt and immobilized with an aminated-LT probe, which was confirmed by Fourier transform infrared spectroscopy (FTIR), cyclic voltammetry (CV), and electrochemical impedance spectroscopy (EIS). This biosensor showed high efficiency in determining ETEC's DNA in the range of 1.9 × 10^-8 μg/mL -1.9 μg/mL with a limit of detection (LOD) of 1.9 × 10^-8 μg/mL. The developed biosensor exhibited high specificity without cross-reaction with other foodborne pathogenic bacteria, including <i>Shigella dysenteriae</i>, <i>Salmonella</i> Typhi, and <i>E. coli</i> ATCC25922. Moreover, this biosensor was challenged with spiked samples, prepared by the standard addition method, which demonstrated good performance, represented by good accuracy (Recovery >90%) and precision (coefficient of variation or COV < 10%). This is a promising technology that has the potential to be further developed as a point-of-need for medical diagnosis and epidemiological surveillance.