Enterotoxigenic bacteria E. coli and Shigella together they cause hundreds of millions of infections each year and are among the leading causes of death from diarrhea, especially in children. Decades of vaccine development efforts have stalled, in part because common vaccine targets vary so much from strain to strain.
New research from Washington University School of Medicine in St. Louis points to a common biological feature of these gut pathogens that could lead to a vaccine that protects against both.
Researchers at WashU Medicine, along with collaborators at the University of Missouri and the International Center for Diarrheal Disease Research in Bangladesh, found that enterotoxigenic E. coli (the leading cause of traveler’s diarrhea), Shigella and other pathogens that cause diarrhea rely on three closely related enzymes to break through the protective mucus layer of the gut and cause infection. Based on samples from infected patients and volunteers exposed to the bugs, the team showed that antibodies targeting a common region of these enzymes can neutralize all three biomolecules and prevent the bacteria from penetrating the gut’s mucus barrier.
The results, appearing June 15 in PNAS, indicate the potential for a single combination vaccine against these major causes of severe diarrhea.
“For something so common and so deadly to young children, it’s striking that we still don’t have a vaccine for any of these pathogens,” said James M. Fleckenstein, MD, professor of medicine in the Department of Infectious Diseases at WashU Medicine and co-senior author of the study. “What’s exciting here is that we found a kind of Achilles heel or shared weakness that we could target to protect against both.”
A common vulnerability
To cause disease, gut pathogens must first penetrate a thick layer of mucus that lines the gut and keeps out even the body’s healthy bacteria. Getting past that barrier is a critical early step in infection—and a point where, Fleckenstein said, harmful bacteria can be stopped without disrupting beneficial microorganisms. Enterotoxigenic E. coli (ETEC) – so named because it causes gastrointestinal disease, unlike other strains E. coli which are harmless – and Shigella manage the job using closely related enzymes that cut the main protein in the gut mucus. Once they breach the barrier, the bacteria can deliver the toxins that cause diarrhea.
Fleckenstein’s lab identified such a diarrhea-causing enzyme for the first time E. colicalled EatA, which appropriately consumes the main structural component of mucus. The team has now shown that two related enzymes – SepA and Pic, produced by Shigella and some other diarrhea-causing bacteria—perform the same mucus-eliminating function.
Working with co-author Ali Ellebedy, PhD, the Leo Loeb Professor in the WashU Medicine Department of Pathology & Immunology, Fleckenstein and colleagues isolated antibodies from patients in Bangladesh naturally infected with ETEC and from volunteers intentionally infected with the bacteria in controlled studies. They found that antibodies blocking EatA also neutralized SepA and Pic. Antibodies are proteins that the immune system produces to recognize a specific target and lock onto it so it can be destroyed.
Structural biologists at the University of Missouri, including first author David P. Buckley, PhD, a postdoctoral research associate, then used cryo-electron microscopy—a technique that freezes molecules to image them in detail—to pinpoint exactly where the most effective antibodies stuck to the enzymes. The spot turned out to be a common area in all three, which explains how a single antibody can disable the mucus-degrading machinery of many pathogens. It also gives vaccine designers a precise target for creating a vaccine that would prompt the immune system to produce such antibodies and have them ready in the event of infection.
This study establishes EatA as a viable vaccine candidate capable of providing protection against multiple pathogens. By identifying the key regions of EatA that are targeted by neutralizing antibodies capable of inhibiting its enzyme function, we have created a basis for rational vaccine design—an important advance toward the development of effective therapeutics that have the potential to save many lives.”
Zachary Berndsen, PhD, assistant professor of biochemistry, University of Missouri and co-senior author on the study
The project builds on previous studies of children in Dhaka, Bangladesh, showing that those who develop natural antibodies against EatA tend to be protected from disease, while children without them are more likely to get sick.
The need for vaccines to protect against these infections is not limited to the developing world. Enterotoxigenic E. coli has caused major foodborne outbreaks in the United States and because it is difficult to distinguish from harmless E. coli in most clinical laboratories, cases often go unrecognized. Reliance on antibiotics to treat these infections also fuels antibiotic resistance, which knows no borders, Fleckenstein noted.
The team is now working to move towards developing a vaccine.
“These bacteria have evolved alongside us and become very good at breaching our defenses,” Fleckenstein said. “If we can block this first step, we have a chance to stop these infections before they even take hold.”
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