Impedimetric detection of pathogenic bacteria with bacteriophages using gold nanorod deposited graphite electrodes

Abstract

Electrochemical impedance spectroscopy (EIS) is applied for the detection of bacteria using bacteriophages as a bioprobe together with gold nanorods (GNRs). Escherichia coli -E. coli K12 was used as a model target bacteria and also for the propagation of its specific T4-phages. Gold nanorods (GNRs) were synthesized via a two-step protocol and characterized using different techniques. EIS measurements were conducted in an electrochemical cell consisting of a three electrode system. Single-use pencil graphite electrodes (PGE) were modified by the physical adsorption of GNRs to increase their interfacial conductivity and therefore sensitivity for impedimetric measurements. Therefore, interfacial charge-transfer resistance values (R ct ) sharply decreased after GNRs deposition. Phages were adsorbed on these electrodes via a simple incubation protocol at room temperature, which resulted in an increase in R ct values, which was concluded to be as a result of nonconductive phage layers. These phage-carrying GNRs-PGEs were used for impedimetric detection of the target bacteria, E. coli. Significant increases at the R ct values were observed which were attributed to the insulation effects of the adsorbed bacterial layers. This increase was even more when the bacterial concentrations were higher. In the case of the non-target bacteria Staphylococcus aureus (S. aureus), conductivity noticeable decreases (due to nonspecific adsorption).However, in the case of E. coli, the R ct value increase is time dependent and reaches maximum in about 25-30 min, then decreases gradually as a result of bacterial lysis due to phage invasion on the electrode surfaces. In contrast, there were no time dependent changes with the non-target bacteria S. aureus (no infection and no lytic activity). It is concluded that the target bacteria could be detected using this very simple and inexpensive detection protocol with a minimum detection limit of 10 3 CFU mL À1 in approximately 100 mL bacterial suspension.

Phage-based Electrochemical Sensors: A Review

“…The limit of detection could be reduced to 10 2 cfu/mL with this method in less than 1 h with a detection range of 10 2 -10 7 cfu/mL. The same trend of R ct in terms of bacteria concentration could also be found in the other research works with the impedance measurements [52,82]. Bhardwaj et al developed a screen-printed graphene electrode for sensitive detection of S. arlettae by covalently immobilizing the highly specific lytic bacteriophages against S. arlettae on the sensor surface [52].…”

“…Zhou et al developed a T2 bacteriophage-based carbon nanotube transducer for electrochemical detection of E. coli B. EIS measurements were used to monitor the variance in the interfacial impedance due to the specific capturing of target bacteria. When the bacteria concentration was increasing, the R ct was decreasing as shown in Figure 3, which was contrary to the previously introduced research works [52,81,82]. To understand this issue, the author also studied the interaction between phage and bacteria with a Backlight bacteria viability kit under fluorescence microscopy.…”

“…A linear increase in the values of R ct was observed in respect to the increasing bacteria concentration (2-2 × 10 6 cfu/mL), based on which the limit of detection can be obtained as around 2 cfu/mL with good stability over 3 months [52]. Moghtader et al developed a single-use pencil graphite electrode (PGE) physically coated with synthesized gold nanorods (GNRs) to increase the interfacial conductivity, which could enhance the sensitivity for impedance measurements [82]. T4 phages specific for E. coli K12 were then immobilized on the surface of GNRs-PGEs.…”

Fast Detection of Pathogenic Escherichia coli from Chicken Meats

“…These techniques need traditional sample preparation, though very efficient in extracting the target analyte, which is time-consuming and produces large amount of solvent wastes. Among the various techniques, electrochemical impedance spectroscopy technique and surface plasmon resonance imaging have previously been investigated to study the detection of Pathogenic Bacteria on gold [11]. These two techniques offer several advantages: First, they are label-free and direct detection method for biomolecular interactions because the measurements are based on small electric signal and very large range of frequency (100 mHz-100 kHz) and refractive index changes [12][13][14].…”

Impedance Detection of 3‐Phenoxybenzoic Acid with a Noncompetitive Two‐site Phage Anti‐immunocomplex Assay

“…Bacteriophage have numerous advantages over antibodies for biomolecular recognition, including their simplicity and reduced cost, improved stability with respect to temperature and harsh reagents, and easier modification by chemical conjugation or genetic engineering . The most common electroanalytical application is bacterial pathogen detection by electrochemical impedance spectroscopy (EIS), following surface immobilization of a bacteriophage that infects the pathogen of interest . Several studies report significantly reduced pathogen cross‐reactivity for bacteriophage‐based relative to antibody‐based assays .…”