Label‐free molecular detection of antibiotic susceptibility for Mycobacterium smegmatis using a low cost electrode format

Abstract

Today, the emergence of antibiotic resistance in pathogenic bacteria is considered an important problem for society. Excessive consumption of antibiotics, long‐term treatments, and inappropriate prescriptions continually increase the severity of the problem. Improving antibiotic stewardship requires improved diagnostic testing, and, therefore, in vitro antibiotic susceptibility testing is becoming increasingly important. This research details the development of an antibiotic susceptibility test for Mycobacterium smegmatis using streptomycin as antibiotics. This strain was selected because it is a member of the slow growing Mycobacterium genus and serves as a useful surrogate organism for M. tuberculosis. A commercially available and low‐cost screen‐printed gold electrode in combination with a specifically developed nucleic acid probe sequence for the 16SrRNA region of the mycobacterial genome was employed to monitor M. smegmatis nucleic acid sequences using the techniques of square‐wave voltammetry and electrochemical impedance spectroscopy. The results show that it was possible to detect M. smegmatis sequences and distinguish antibiotic‐treated cells from untreated cells with a label‐free molecular detection. As a result, the in vitro antibiotic susceptibility test revealed that M. smegmatis showed sensitivity to streptomycin after a 24‐H incubation, with the developed protocol representing a potential approach to determining antibiotic susceptibility more quickly and economically than current methods.

Highly Sensitive Label-Free Electrochemical Detection of Heat Shock Protein With Low-Cost Screen-Printed Electrodes

“…As a result of the hybridization, an increase in negative charge was observed at the electrode surface. With this increase of negative charge, the charge transfer of the negatively charged redox molecules was hindered in the solution, causing a change in the RCT, measurable by EIS as reported previously [29,31]. In EIS, impedance behaviour is modelled by solution resistance (Rs), constant phase element (CPE), RCT and Warburg impedance (W), and these are expressed by Randles circuit (Figure 3A) and Nyquist plot is illustrated by these elements (Figure 3B) [32].…”

On‐chip label‐free impedance‐based detection of antibiotic permeation

“…As for the permeability testing, an LCR metre was chosen to measure the impedance. Impedance testing has been previously reported to be sensitive to the cell type that could be stimulated by different chemicals or drugs [ 36 ,[ 37 ]. Likewise, we hypothesized that the uptake of antibiotics into the GUVs could be monitored by a simple impedance testing because the outer membrane of GUVs in solution comprises an RC circuit [ 38 , 39 , 40 ] (Figure 4 ).…”

Electrochemical Quantification of Tobramycin Retention in Pseudomonas aeruginosa as Antimicrobial Susceptibility Indicator

“…17,18 Because of advantages such as high sensitivity, cost-effectiveness, portable instrumentation, and short analysis time, electroanalytical methods emerged as attractive approaches to detect bacterial drug resistance over the last two decades. 19,20 These methods allow direct measurements in complex biological samples, such as urine and blood, making them promising tools for the development of sensors for the point-of-care detection of antibiotic resistance. 20 including those for Staphylococcus aureus, 21 Mycobacterium smegmatis, 19 Klebsiella pneumoniae, 22 Salmonella gallinarum, 23 and Escherichia coli 24 strains.…”

“…19,20 These methods allow direct measurements in complex biological samples, such as urine and blood, making them promising tools for the development of sensors for the point-of-care detection of antibiotic resistance. 20 including those for Staphylococcus aureus, 21 Mycobacterium smegmatis, 19 Klebsiella pneumoniae, 22 Salmonella gallinarum, 23 and Escherichia coli 24 strains. Although these studies report successful methodologies to detect antibiotic resistance, the described systems are not user-friendly enough to find broad application in clinical settings, as they involve complex electrode surface modification steps, which often limits sensor stability and storage in addition to increasing production costs.…”

“…Patients are commonly treated empirically, exposed to a broad spectrum of different antibiotics in severe cases, thus increasing the chances of antibiotic resistance development. , New methods and technologies for AST, aiming to overcome these significant drawbacks, are under constant development, including genome sequencing, , mass spectrometry, , surface enhanced Raman scattering (SERS), , Fourier transform infrared spectroscopy (FTIR), and infrared microscopy. , The aforementioned methods depend on laboratory-based analytical techniques that require expensive and bulky equipment, increasing the cost of the test. , In addition, the detection of a resistance gene or mutation often does not necessarily correlate with resistance, as the presence of specific resistance genes does not directly imply its active expression. , Because of advantages such as high sensitivity, cost-effectiveness, portable instrumentation, and short analysis time, electroanalytical methods emerged as attractive approaches to detect bacterial drug resistance over the last two decades. , These methods allow direct measurements in complex biological samples, such as urine and blood, making them promising tools for the development of sensors for the point-of-care detection of antibiotic resistance . Studies employing electrochemical techniques to detect bacterial antibiotic susceptibility have been reported, including those for Staphylococcus aureus, Mycobacterium smegmatis, Klebsiella pneumoniae, Salmonella gallinarum, and Escherichia coli strains. Although these studies report successful methodologies to detect antibiotic resistance, the described systems are not user-friendly enough to find broad application in clinical settings, as they involve complex electrode surface modification steps, which often limits sensor stability and storage in addition to increasing production costs.…”