Schematic of the antibiotic susceptibility testing device. The bacteria are cultured in miniature chambers, each of which contains a filter for bacterial capture and electrodes for readout of bacterial metabolism. [U of T Engineering]
Schematic of the antibiotic susceptibility testing device. The bacteria are cultured in miniature chambers, each of which contains a filter for bacterial capture and electrodes for readout of bacterial metabolism. [U of T Engineering]

Researchers at the University of Toronto say they have designed a small and simple chip to test for antibiotic resistance in just one hour, giving doctors the opportunity to choose the most effective antibiotic to treat potentially deadly infections. Many tests for antibiotic resistance currently take up to three days to come back from the lab.

Their Toronto team’s study (“Rapid electrochemical phenotypic profiling of antibiotic-resistant bacteria”) was published in Lab on a Chip.

A 2015 Health Canada report estimates that superbugs have already cost Canadians $1 billion, and are a “serious and growing issue.” Each year two million people in the U.S. contract antibiotic-resistant infections, and at least 23,000 people die as a direct result.

Resistant bacteria arise in part because of imprecise use of antibiotics. When a patient comes down with an infection, the doctor wants to treat it as quickly as possible and often prescribes a broad-spectrum antibiotic.

“Guessing can lead to resistance to these broad-spectrum antibiotics, and in the case of serious infections, to much worse outcomes for the patient,” says Justin Besant, Ph.D., a member of the scientific team. “We wanted to determine whether bacteria are susceptible to a particular antibiotic, on a timescale of hours, not days.”

The problem with most current tests is the time it takes for bacteria to reproduce to detectable levels. Dr. Besant and his colleagues drew on their collective expertise in electrical and biomedical engineering to design a chip that concentrates bacteria in a miniscule space, i.e., just two nanoliters in volume, to increase the effective concentration of the starting sample.

They achieve this high concentration by flowing the sample, containing the bacteria to be tested, through microfluidic wells patterned onto a glass chip. At the bottom of each well a filter, composed of a lattice of microbeads, catches bacteria as the sample flows through. The bacteria accumulate in the nano-sized well, where they're trapped with the antibiotic and a signal molecule called resazurin.

Living bacteria metabolize resazurin into a form called resorufin, changing its electrochemical signature. If the bacteria are effectively killed by the antibiotic, they stop metabolizing resazurin and the electrochemical signature in the sample stays the same. If they are antibiotic-resistant, they continue to metabolize resazurin into resorufin, altering its electrochemical signature. Electrodes built directly into the chip detect the change in current as resazurin changes to resorcin.

“This electrochemical phenotyping approach is effective with clinically-relevant levels of bacteria, and provides results comparable to culture-based analysis,” wrote the investigators.

“This gives us two advantages,” says Dr. Besant. “One, we have a lot of bacteria in a very small space, so our effective starting concentration is much higher. And two, as the bacteria multiply and convert the resazurin molecule, it's effectively stuck in this nanoliter droplet—it can't diffuse away into the solution, so it can accumulate more rapidly to detectable levels.”

Rapid alternatives to existing antibiotic resistance tests rely on fluorescence detection, requiring expensive and bulky fluorescence microscopes to see the result.

“The electronics for our electrochemical readout can easily fit in a very small benchtop instrument, and this is something you could see in a doctor's office, for example,” says Dr. Besant. “The next step would be to create a device that would allow you to test many different antibiotics at many different concentrations, but we're not there yet.”








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