By Deborah Borfitz
March 16, 2021 | Experts in basic medical sciences, biomedical engineering, and physics at Purdue University have collaborated to develop Doppler spectroscopy—the technique used to reveal the existence of extrasolar planetary systems—to study bacterial invasions of healthy tissue. They recently demonstrated that their testing device can both tell if an unknown microbe is pathogenic and identify the best antibiotic to fight it in just a few hours, according to David Nolte, professor of physics and astronomy.
The feat was made possible by two major breakthroughs, the first by Nolte and John Turek (the basic medical scientist) while researching cancer over 10 years ago. They found that it was possible to measure the Doppler shifts of intercellular motion. More recently, in partnership with Professor Mike Ladisch and research scientist Eduardo Ximenes (the microbiology experts), they applied the biodynamic assay to bacteria using small tissue cultures, or “tissue sentinels,” grown from commercially available immortalized cancer cell lines, says Nolte.
"First we did biodynamic imaging applied to cancer, and now we're applying it to other kinds of cells," Nolte says. "This research is unique. No one else is doing anything like it."
The central clinical problem is that it generally takes about 24 hours to get enough bacteria cultured to identify the culprit pathogen and the standard-of-care antibiotic, Nolte says. By then, a patient’s health could have significantly degraded and sepsis—the body’s often deadly response to infection—could have set in.
Consequently, physicians often treat sepsis with broad-spectrum antibiotics and in doing so make the situation worse for the next patient, Nolte says. Letting bacteria come into close contact with antibiotics that do not kill them only makes them more resistant to that antibiotic and more difficult to fight next time.
After only a few hours of culturing, too few bacteria have grown to be visible even when magnified under a microscope, he explains. But the altered behavior of the sentinels as they respond to the bacterial invasion would be detectable in that short time span. The Doppler signals change quite significantly when cells are pathogenic, and not at all when infection is absent.
Sensing Bacteria
The immortalized cell lines used for the Doppler radar-like diagnostic device are “little pieces of tissue acting as sensors” that detect bacteria in a patient sample, which might be saliva, blood, or possibly some other bodily fluid, Nolte says. In a paper recently published in Communications Biology (DOI: 10.1038/s42003-020-01550-8), the team exposed the cells to Salmonella and E. coli and then used the Doppler effect to spy out how they reacted.
Their device uses a standard 96-well plate, meaning up to 96 tissue sentinels could be exposed to a patient’s fluid with the low bacterial load at one time, says Nolte. Different antibiotics would be applied to each of those wells to see which recovered from the infection and which did not. The identity of the bacteria is irrelevant to the approach.
“The antibiotic that works is just the one that works” and, if unexpected, might even prompt a clinical study, Nolte says. If all 96 antibiotics fail to work, the next step would probably be to run a second panel “of more exotic drugs.”
In the future, the technique might be used on patients thought to be in the early stages of bacterial sepsis to ensure the diagnosis is correct and, if so, the antibiotic most likely to work gets selected. “Antibiotic resistance keeps increasing and it is going to become a really serious health threat by around 2050,” he notes.
“People tell stories about what life was like before antibiotics when you could die from a paper cut, and then antibiotics came along and everyone took them for granted,” says Nolte. “Now we give antibiotics to everything, including feedstock.” Experts predict that most standard antibiotics will no longer work within the next three decades “and we may go back to that time [of death by paper cut].”
The new Doppler-based tool for rapidly testing antibiotics against a pathogen is a potential answer to the growing antimicrobial resistance threat, he says. “There are lots and lots of antibiotics [about 200 are currently approved for use by the U.S. Food and Drug Administration] and the common ones doctors typically use will have the greatest resistance.” As antibiotic resistance increases, they may be forced to switch to the more expensive, novel antibiotics on the market.
Universal Approach
Development of Purdue’s latest biomedical application will likely begin with a few specific types of infections where bodily fluids can be easily obtained and then expand into more general use, Nolte says. “Over time it could become a universal approach.”
The research team wants to expose the tissue sentinels to other types of microbiota, including viruses, as well as nonliving pathogenic cells or dried spores to see how they respond and if there might be treatments to get rid of them, he adds. Anything toxic trying to invade a tissue, including environmental pathogens, could be studied with the technique.
Although their labs at Purdue do not have the requisite biosafety level to work with SARS-CoV-2, the researchers plan to work with other types of viruses in hopes of having the technology ready by the time the next pandemic strikes. They have already identified viral sepsis, which has been discussed in connection with COVID-19, as a future target.
Nolte estimates it will be another two or three years before the device is clinic-ready. The research team still needs to figure out how small the tissue sentinels can be and still generate good Doppler signals, as well as how few bacteria are necessary to change the dynamics of the sentinels. Based on their back-of-the-envelope calculations, he adds, they believe that the sentinels can be small enough to respond with Doppler signals in about three hours.
The speediest results turnaround today—eight hours—is possible with high-tech, state-of-the-art assays but few hospitals can afford them, says Nolte. “Our approach could be very simple and inexpensive. We just shine LED light on the tissues and measure the scattered light without any complicated light sources or detectors.”
Purdue filed a patent application for the technology a few years ago, which will be licensed to Indianapolis-based startup Animated Dynamics, where Nolte serves as chief scientific officer. The company launched as an LLC in 2011 and converted to a C corporation in 2015. It is “on the cusp” of commercializing a product for selecting chemotherapy for cancer patients—closely paralleling its ambitions to create a device to select antibiotics for sepsis patients.