The selection of a suitable nucleic acid probe sequence for specific target recognition is the first step in the development of an electrochemical nucleic acid-based sensor, and have a huge influence on its specificity . This could also affect the affinity of redox molecules, often used in electrochemical detection. For example, methylene blue has a high affinity towards guanine, therefore a higher concentration of guanine bases on the probe DNA, correlates with a higher signal from methylene blue . The probe sequence could also influence the intensity of an electrochemical signal of the sensor. Table highlights the types of nucleic acid and target genes most used in nucleic acid-based sensors for the detection of general E. coli and STEC. The main challenge in nucleic acid-based specific detection of generic E. coli is its close genetic relationship with Shigella spp., another common pathogen that belongs to the family Enterobacteriaceae. It was estimated that their nucleotide similarity is between 80 and 90% and they are usually differentiated based on their biochemical characteristics. This makes the selection of truly unique and specific primer for E. coli detection a real challenge to avoid a non-specific detection of microorganisms not related to faecal contamination .