Shining a Light on Antibiotic Heteroresistance

Interferometry To Detect Antibiotic Heteroresistance

In August 2016, a Nevada woman died from an incurable infection. After an extended trip to India, the woman, who was in her 70s and had been traveling with a femur fracture, contracted an infection in her hip. Her infection, deemed a "superbug," was resistant to all 26 antibiotics available in the U.S. – including colistin, a last-line antibiotic. The case made national headlines and alarmed public health officials. But in the midst of this tragedy, Emory scientists had been working on a way to treat antibiotic resistance, offering new insight into how these “invincible” bacteria operate.

David Weiss, PhD, a professor at Emory School of Medicine, and his research team used two isolates from the Nevada case, and found that a customized antibiotic combination could effectively kill antibiotic resistant bacteria.

David Weiss, PhD

Shifting from a binary view of bacteria
For years, scientists and medical professionals believed that all the cells from a single infecting strain of bacteria would respond uniformly to a given antibiotic. In diagnostic tests used in clinics and hospitals, you could have two outcomes: Either the antibiotic would defeat the bacteria, or the bacteria would resist the drug. But in cases like the Nevada woman’s, there can be a third, sneaky outcome called antibiotic heteroresistance.

Antibiotic heteroresistance is the idea that subpopulations of both resistant and susceptible bacteria can exist within one strain; they are not mutually exclusive. It was a case like the Nevada woman’s that led to the initial discovery of antibiotic heteroresistance – one originating not in the lab, but in the clinic.

The head of the clinical microbiology lab at Emory Hospital approached Weiss with a mysterious isolate and asked that he study it further. Weiss and his team first tried to figure out how the isolate worked. “It was just so weird and everything [about it] was not what I was taught was supposed to happen,” he said. “We were just, at first, really intrigued by the biology.”

What started as a side project on heteroresistance soon became the focus of the entire lab. “Once we started to understand more, we thought, ‘Could this be relevant? Could this really be messing up treatment?’” The answer was yes. They found that heteroresistant cells can account for treatment failure in a patient, like antibiotics failing even when tests label the bacteria susceptible.

Combining drugs for a stronger treatment
Here’s how current technology assesses heteroresistant bacteria: If there are more susceptible bacteria than resistant bacteria in a population, the test labels the entire population “susceptible.” A clinician might prescribe an appropriate antibiotic, but treatment would fail due to the presence of a subpopulation of cells resistant to the antibiotic. Normally, when bacteria are not susceptible to any single antibiotic, clinicians will prescribe "combination therapy" by picking two antibiotics and mixing them. But there are no actual guidelines for how to pick the two antibiotics; it is left to the clinician's discretion, leading to varying results.

Weiss’ lab discovered a new way to approach combination therapy: If a bacterial strain were heteroresistant to two antibiotics, scientists could combine those two antibiotics to create an effective treatment. This is because the resistant subpopulations were independent and did not work together to defeat the drugs.

David Mudd, Licensing Associate with the Emory Office of Technology Transfer, praised the new invention, commenting, “This technology could be a crucial part of preventing the rise of the next bacterial superbug.”

Creating a new test for heteroresistance
Current antibiotic susceptibility tests completely overlook heteroresistance, so with his discovery in mind, Weiss looked to create a new test. “We think that the current clinical tests are like going to the IMAX movie without the glasses. With our test, it's like putting on 3D glasses,” Weiss explained. “When everything becomes clearer, and the doctors can make much better treatment decisions.”

To create the new clinical test, Weiss teamed up with Peter Yunker, PhD, a physics professor at Georgia Tech studying interferometry. This is a way to use the interference of two light beams to make precise measurements. Light A is shone on the sample, and Light B is shone on a mirror. Light A reflects off the sample back to its source, while Light B does the same thing off the mirror. Combining the two beams and analyzing how they interfere with each other creates a high-resolution image of the surface of the sample.

Interferometers are usually used in physics and material science, and rarely in microbiology and medicine. But Yunker’s lab looked at bacterial biofilms and studied the surface with biophysics, connecting the surface shape to the activity happening beneath it.

“I met David, and from when we first chatted, it became very clear that they had the problem and we have the tool that can be the solution,” Yunker said.

Together, Yunker and Weiss developed an algorithm to determine whether bacteria are resistant, susceptible, or heteroresistant to an antibiotic based on the shape of the surface of the population. First, they grow the bacteria on an antibiotic for a few hours – much less time than current tests, which grow it overnight or for an entire day. Next, they image the population using the interferometer, which gives them a map of the surface of the population of bacteria. The algorithm takes those results and determines bacteria behavior in response to the antibiotic.

Preventing an oncoming threat
There are roughly 1.27 million deaths worldwide due to antibiotic heteroresistance, more than HIV and malaria combined. Those deaths only increase as time goes on. Unlike COVID-19, antibiotic heteroresistance is a slow-creeping pandemic, though its effects could pose new and perhaps more alarming challenges than COVID. “It's lurking and growing, but it's very slow,” Weiss explained. “So I think it's easier for people to ignore it or lose track of it. But in the end, it could be, in some ways, even a bigger problem, which is hard to imagine.”

Without antibiotics, a patient’s chance of survival can dramatically decrease. In cases with serious antibiotic heteroresistance, some surgeries – even small ones like a knee replacement – may not even be possible. Many would rather hobble around on a bad knee than risk death from an incurable infection.

“Antibiotic resistance could cripple medicine as we know it,” Weiss said. “A lot of the things we take for granted in medicine would not be possible without antibiotics.”

But with interferometry to detect antibiotic heteroresistance, it’s possible to test for and treat heteroresistance. If we can catch it, these bacterial subpopulations won’t remain incurable – and doctors will be able to tame infections, bringing their patients back to health.

— Chaya Tong

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