Escherichia coli O157:H7 (E. coli O157:H7) is one of the top pathogens of interest for the development of rapid diagnostic systems for food and water samples. The objective of this research is to develop a rapid, novel electrochemical biosensor based on the use of polypropylene microfiber membranes coated with a conductive polypyrrole and antibody functionalized for the biological capture and detection of E. coli O157:H7 inthe field. Using glutaraldehyde, pathogen specific antibodies are covalently attached to conductive microfiber membranes which are then blocked using a 5% bovine serum albumin solution. The functionalized membranes are then exposed to E. coli O157:H7 cells washed in Butterfield’s phosphate buffer and added to a phosphate-buffer electrolyte solution. When a voltage is applied to the system, the presence of the captured pathogen on the fiber surface results in an increase in resistance at the electrotextile electrode surface, indicating a positive result. In this study, the initial resistance of the membrane in the electrochemical system was established and found to range between 5.8 and 13 . The resistance of the system not associated with the electrotextile fibers was calculated to contribute to only 2.8% of the total system resistance, and found not to be significant. A proof of concept experiment was conducted and determined that the electrotextile electrode was able to differentiate between small changes in a solution’s conductivity associated with the presence of E. coli O157:H7 cells over a concentration range of log 0 - 9 CFU/mL.