A revolutionary technique that measures the metabolic activity of bacteria with an electric probe can now identify antibiotic resistance in under 90 minutes, a dramatic improvement from the one to two days required by current techniques.
This discovery means that doctors could quickly know which antibiotics will or won't work for a patient's life-threatening infection, a quandary faced by doctors on a daily basis in hospitals worldwide.
A Washington State University research team has reported on its work in the journal Biosensors and Bioelectronics.
“The idea is to give the doctors faster results so they can make clinically appropriate decisions within that timeframe that they're working, rather than having to wait,” said Douglas Call, Regents Professor in the Paul G. Allen School for Global Health and a co-author of the paper. “Instead of looking for growth of a culture, we look for metabolism, and that is basically what we’re detecting by the movement of these electrons so it can happen in much shorter time spans compared with a conventional culture-based assay.”
The prevalence of antibiotic resistance is increasing around the world and threatens the ability to treat many common infectious diseases. For example, millions of people in the US are infected annually with drug- resistant pathogens, and thousands of people die from pneumonia or bloodstream infections that become impossible to treat.
To determine definitively whether a particular infection is resistant to antibiotics requires separating and then growing the bacteria in a lab and watching the population grow in a process that can take up to two days or more. Doctors with sick patients often have to prescribe an antibiotic immediately without having complete information on how well it will work.
In their paper, the WSU team used a probe to directly measure the electrochemical signal of the bacteria, thereby measuring their metabolism and respiration and learning how they are faring long before they would be visible in culture. Looking at eight different strains of bacteria, they used the bacteria's electric signal to determine in less than 90 minutes which were susceptible or resistant to the antibiotics.
The bacteria that are still metabolising and “breathing” after antibiotic treatment are considered resistant.
Previous attempts to measure the electrochemical activity of bacteria had been limited because most bacterial species are not capable of transferring electrons directly to an electrode, said Abdelrhman Mohamed, a postdoctoral researcher in the Gene and Linda Voiland School of Chemical Engineering and Bioengineering, who was a lead author on the paper.
The researchers added a chemical mediator to their assay, which acted as a shuttle, taking the electrons from the surface proteins of the bacteria and moving them to the researchers’ electrode, where the electric signal could be measured.
“That allows us to have a universal mechanism that can test all types of pathogens,” said Mohamed.
The researchers tested four different bacterial species that cause hospital-acquired infections and tested a variety of antibiotics that work via different mechanisms. They also developed an antibiotic susceptibility index to categorise the results in a way that could help doctors decide which antibiotic to use.
They are now planning to engineer their probe to be convenient and standardised for clinicians to use, and hope to commercialise it.
“It’s really exciting to be involved in a project that not only is valuable from a scientific view but has commercial and industrial applications that could potentially someday actually improve people’s lives,” said Gretchen Tibbits, a lead author on the paper and graduate student in the Voiland School.
They are also working to better understand the fundamental mechanisms of the electrochemical process to further improve it.
“We are doing it in two hours, but if we understand mechanisms better, maybe we can do this in minutes,” said Haluk Beyenal, co-author on the paper and a professor in the Voiland School. “As long as the bacteria are alive, we can do this measurement.”
Study details
Rapid differentiation of antibiotic-susceptible and -resistant bacteria through mediated extracellular electron transfer.
Gretchen Tibbits, Abdelrhman Mohamed, Douglas R. Call, Haluk Beyenal.
Published in Biosensors and Bioelectronics on 2 November 2021
Highlights
• We developed an electrochemical assay to rapidly detect antibiotic susceptibility.
• Current response was used to define antibiotic susceptibility index (ASI).
• ASI along with an algorithm was used to classify antibiotic susceptible and resistant strains.
• We tested the assay with four important nosocomial pathogens and diverse antibiotics.
• The assay was validated using blind test and compared with conventional tests.
Abstract
Conventional methods for testing antibiotic susceptibility rely on bacterial growth on agar plates (diffusion assays) or in liquid culture (microdilution assays). These time-consuming assays use population growth as a proxy for cellular respiration. Herein we propose to use mediated extracellular electron transfer as a rapid and direct method to classify antibiotic-susceptible and -resistant bacteria.
We tested antibiotics with diverse mechanisms of action (ciprofloxacin, imipenem, oxacillin, or tobramycin) with four important nosocomial pathogens (Acinetobacter baumannii, Staphylococcus aureus, Escherichia coli, and Klebsiella pneumoniae) by adding the bacterial culture to a custom-designed electrochemical cell with a glassy-carbon electrode and growth media supplemented with a soluble electron transfer mediator, phenazine methosulfate (PMS). During cell respiration, liberated electrons reduce PMS, which is then oxidised on the electrode surface, and current is recorded.
Using this novel approach, we were able to consistently classify strains as antibiotic-resistant or -susceptible in <90 min for methodology development and <150 min for blinded tests.
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