An antifouling coating that enables electrochemical biosensing of MecA gene in complex samples

As an important emerging pollutant, antibiotic resistance genes (ARGs) monitoring is crucial to protect the ecological environment and public health, but its accurate detection is still a major challenge in complex environment samples. In this study, a robust antifouling electrochemical DNA biosensor (E-DNA biosensor) was proposed for highly sensitive and selective detection of the mecA gene in complex water samples. The antifouling coating of electrode consisted of a branched-shaped zwitterionic peptide and gold nanoparticles (CPEK-AuNPs). And, the PPPP linker of peptide between the anchoring and antifouling domains adopted a polyproline helix conformation. The CPEK-AuNPs-modified electrode interface with stable hydration layer prevented non-specific interactions while enhancing electron transfer to the electrode surface, resulting in high current response and outstanding antifouling performance. Coupled with synergistic isothermal strand displacement polymerase reaction (ISDPR), the proposed antifouling sensing strategy exhibited superb sensitivity for the mecA gene. The limit of detection (LOD) was as low as 16.67 fM (S/N = 3) in a broad linear range of 0.05–5000 pM, and a high specificity to resolve from even single base mismatch towards the complex water samples was obtained. Benefiting from the stable hydration layer, the as-prepared E-DNA biosensor preserved 91.8 % of the initial signal after 15 days of exposure in unprocessed wastewater samples. The excellent specificity, stability and reproducibility (RSD<3%) of mecA gene detection indicated that the proposed sensing strategy with antifouling performance can be a versatile tool for accurate quantitative analysis of ARGs in complex environment samples.