Ryan Hunter, PhD, associate professor of microbiology and immunology at the Jacobs School, and his colleagues created a device to more easily study anaerboic bacteria, described in a recent mBio paper.
Device Creates Oxygenated, Oxygen-Free Conditions
Researchers’ New Platform Enables Simultaneous Study of Anaerobic Bacteria and Aerobic Cells
By Keith Gillogly
Published April 9, 2025
Within our airways, oral cavity, and gut, anaerobic bacteria proliferate. But because these bacteria require oxygen-free conditions for survival, studying them and their interactions with other microorganisms and cells in the body has been difficult.
“The presence and abundance of anaerobic bacteria have long been recognized,” says Ryan Hunter, PhD, associate professor of microbiology and immunology at the Jacobs School of Medicine and Biomedical Sciences. “But whether and how they contribute to both health and disease is poorly understood because of the lack of compatible model laboratory approaches to study their potential contributions to the disease process.”
It’s so challenging to study interactions between anaerobic bacteria and aerobic human cells, which require oxygen, in part because keeping the latter alive in oxygen-limited conditions isn’t easy.
To address this problem, Hunter and his colleagues created a device enabling a dual oxic-anoxic culture (DOAC) approach, which allows for studying bacteria and cells in an environment simultaneously containing both oxygenated and oxygen-free conditions. These conditions simulate the microenvironment inside human airways, particularly during infection, among other sites within the body.
The work is described in a recent paper titled “Dual Oxic-Anoxic Co-Culture Enables Direct Study of Anaerobic-Host Interactions at the Airway Epithelial Interface” published online on April 9 in mBio, an American Society for Microbiology journal.
A Simple, Cost-Effective Approach
The DOAC device uses mostly commercially available components, Hunter says, including gas-permeable culture plates. After placing the device in an anaerobic chamber, the top of the culture plate is cut off from oxygen while the contained bottom portion remains oxygenated. Custom 3D printed components form a tight seal to prevent oxygen inserted below from flowing into the top.
Hunter says that while it’s possible to use organ-on-a-chip nanotechnology to study dual aerobic-anaerobic conditions, that approach requires more specialized engineering and technology, costs more, and doesn’t permit high-throughput results.
The DOAC platform is much simpler. “We wanted something more accessible that we could share with other labs,” Hunter says. “Something that’s easy to use, highly reproducible, and cheap.” The device also permits direct contact between the anaerobic bacteria and aerobic cells being studied.
Hunter notes that development of the device, and subsequent testing using bacterial models, required a multidisciplinary team, drawing on expertise in cell biology, microbiology, engineering, and computational analysis, among other fields. “It’s a great example of team science and an assimilation of various expertise,” he says.
Bacteria and Cells Provide Model for Evaluation
After creating the DOAC device, Hunter and his team evaluated it using airway epithelial cells and the bacterium Fusobacterium nucleatum as a model. This oxygen-sensitive anaerobic bacterium is part of the normal human oral microbiome and also colonizes various mucosal surfaces — such as airways — throughout the body.
Recent evidence, however, has linked F. nucleatum to diseases such as esophageal cancer, colorectal cancer, and chronic airway infections.
Hunter, who’s long studied cystic fibrosis (CF), also notes F. nucleatum’s increased presence in the gut and airways of CF patients. People with CF, Hunter says, are at a five to 10 times increased risk of developing colorectal cancers.
Using an electrode device, the research team measured oxygen within the contained spaces to make sure levels matched the desired aerobic-anaerobic conditions. From there, the investigators evaluated the F. nucleatum’s behavior as it grew on the epithelial cells.
The researchers observed increased production of ethanolamine, a metabolite derived from the epithelial cells, which the bacteria may use for fuel, Hunter says. They also found bacterial gene expression changes linked to increased virulence, among other observations.
DOAC Being Shared, Used for Further Studies
In addition to insight on F. nucleatum biology, this research reveals the potential of the DOAC platform. Hunter’s lab has already begun sharing the device with other researchers focused on airway diseases, including a team at Dartmouth College studying polymicrobial communities and their interactions.
Hunter and his team plan to use the DOAC device to study more traditional pathogens and their interactions with other bacteria and the epithelial interface in oxygen-limited environments. Besides airway epithelial cells, Hunter notes the ability to study different epithelial cell types such as nasal and colonic cells using the DOAC.
“We’re very interested in looking at other sites and tissue types and how certain bacteria might affect the host but also how the host is affecting the behavior of bacteria,” Hunter says. “A whole overarching goal of my lab is to come up with better laboratory models that mimic what’s actually happening in the human body.”
Additional study authors include doctoral student Sara Ahmed of the Jacobs School and researchers from the University of Minnesota, North Carolina State University, and Syracuse University.
This research was supported by the Cystic Fibrosis Foundation, the Esselen CF Trust Fund, and the National Institutes of Health.