An electrochemical aptasensor for quick detection and identification of vancomycin-sensitive bacteria
Novaković, Zorica; Khalife, Majd; Costache, Vlad; Gadjanski, Ivana; Radovic, Marko; Vidic, Jasmina
- Publisher
- Zenodo
- Language
- en
Abstract
In this research, we present a new portable electrochemical device with vancomycin and aptamer recognition capabilities for quick detection and identification of Gram-positive bacteria. This type of biosensor overcomes the constraints of conventional bacteria detection approaches, such as time-consuming multi-step procedures, expensive prices, and lengthy processing durations. A vancomycin-modified carbon electrode was utilized to selectively collect Gram-positive bacteria susceptible to this antibiotic. Because of its high affinity, as a biological recognition receptor with a role for binding-sensitive bacteria, vancomycin was deposited on an albumin-modified working carbon electrode. Vancomycin interacts with the peptidoglycan moieties of d-Ala-d-Ala that are completely obtainable in Gram-positive bacteria's cell walls, allowing for a very rapid detection technique. Electrochemical impedance spectroscopy and scanning electron microscopy revealed that capture took only 10 minutes and had a detection limit of a few CFU/mL. After electrode exposure to particular aptamers, differential pulse voltammetry and electrochemical impedance spectroscopy were used to identify the collected strains of Staphilococcus aureus and Bacillus cereus. When gram-positive bacteria were identified using particular aptamers, current intensity decreased and charge transfer resistance increased, whereas the signal remained the same when non-specific aptamers were used. The technique performed well in genuine samples, milk, and serum, detecting a low level of S. aureus and B. cereus in less than 45 minutes.
Full text
An electrochemical aptasensor for quick detection and identification of vancomycin-sensitive bacteria INTRODUCTION We developed an electrochemical sensor that can fully support rapid, sensitive, and strain-specific detection of Gram-positive bacteria directly in a biological matrix (milk and serum) at concentrations of only a few CFUs per mL using two recognition elements ─ vancomycin and aptamers. EXPERIMENT RESULTS Acknowledgments: This work was supported in part by the IPANEMA project, which received funding from the European Union’s Horizon 2020 research and innovation program under grant agreement No 872662 1 2 34 5 6 Vancomycin conjugated to amine groups of an albumin-modified SPCE captures the cells of Grampositive bacteria. Aptamer with a specific affinity toward different bacterial strains identifies the bacteria strain initially captured by vancomycin. Zorica Novakovic1, Majd Khalife2, Vlad Costache2, Ivana Gadjanski1, Marko Radovic1, Jasmina Vidic2 1BioSense, Novi Sad, Serbia. 2National Research Institute for Agriculture Food and Environment, France SEM images: (a) before and (b) after incubation with BSA (c) BSA + vancomycin. (d) CVs response shows the welldefined and reversible redox peaks for [Fe(CN)6] 3–/4– decreased in intensity after every step of functionalization protocol (BSA, BSA/vancomycin, and BSA/vancomycin/ethanolamine) confirming electrode functionalization (e) Current variations for each functionalization step. (f) Nyquist plots of impedance spectra obtained from bare SPCE and SPCE modified with BSA, BSA/vancomycin, and BSA/vancomycin/ethanolamine (g) Fitting values of the charge transfer resistance, Rct, after each step of SPCE functionalization. I Biosensor Fabrication II Electrode characterisation III Biosensor Responses - Systematic increase in Rct with increasing bacterial concentrations; - LOD 2.4 CFU/mL and 2.7 CFU/mL for B. cereus and S. aureus; - Vancomycin did not capture gram - bacteria (E.coli) -no variation in resistance. IV Identification of captured bacterial cells No change in peak current was observed when the electrode with captured B. cereus cells was incubated with Apt1, or when the electrode with captured S. aureus was incubated with Sp14. a b V Specifity CONCLUSIONS VI Real sample We used vancomycin-modified SPCEs to create the first electrochemical aptasensor capable of both detecting and identifying Gram-positive bacteria. The change in Rct varied linearly with the concentration of vancomycin-sensitive bacteria (S. aureus, B. cereus, B. subtilis, S. carnosus, and L. lactis), while no variation in Rct was observed with vancomycin-resistant E. coli. With the incubation of the captured bacteria with a strain-specific aptamer, it is possible to identifies the individual bacterial species captured by vancomycin.
