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Mounting evidence in animal models, as well as correlative studies in humans, indicate that the microbes present in the gut can shape immune responses. In a study published today (September 5) in Cell, researchers have confirmed that link in humans. They showed that, for people who hadn’t had a flu shot or hadn’t caught the bug in the previous three years, a course of antibiotics just before a flu shot led to fewer antibodies produced in response to the immunization than among study participants who didn’t take antibiotics.

“It’s really important to do these kinds of studies in human because there’s a lot of work that’s been done in animal models,” says Dan Littman, who studies microbiota-immune system interactions at New York University School of Medicine and did not participate in the work. “While those are very valuable—particularly for understanding mechanistic aspects of how the immune system responds to microbiota, to vaccination, to infections of pathogenic microbes—there are clearly going to be differences in human, and we know very little about how humans do respond.”

In 2011, Bali Pulendran, an immunologist who is now at Stanford University, and colleagues monitored gene expression in people who received the seasonal influenza vaccine in order to explore how the flu shot works and why it varies in efficacy in different people. While most of the changes made sense, one stuck out: an increased activity of the gene encoding a receptor in the innate immune system that recognizes the bacterial protein flagellin, the core part of flagella that many microbes use to move. This indicated that perhaps the immune system’s recognition of bacteria—potentially the gut microbes—was somehow playing a role in the its response to the vaccine.

Then in 2014, the researchers gave mouse knockouts of the receptor for bacterial flagellin the flu shot. These mice made fewer antibodies to the flu than their littermate controls. The research team suspected that this reduction was mediated by the receptor sensing flagellin present in the animals’ gut microbes, so in separate experiments they depleted mice’s microbiota with antibiotics before vaccination and gave germ-free mice the vaccine. Both groups of mice did not respond as well to the flu vaccine as controls that had their microbiomes intact. They concluded that the gut microbiota plays a role in generating an optimal antibody response against the flu shot, and the next step was to bring the work back to humans.

“There’s a lot of beautiful mechanistic work in mice, and there’s a lot of elegant work in humans showing correlations between particular species of bacteria and susceptibility to allergic inflammation, cancer, or other inflammatory disorders, but precious little causal evidence in humans in which there has been a deliberate and intentional perturbation of the microbiome and then a consequent examination of what this does to human physiology—not just the physiology of the immune system—but any aspect of human physiology,” Pulendran tells The Scientist.

In the current study, the researchers designed a Phase 1 clinical trial to test the influence of gut microbes on flu vaccine–induced immunity. First, they treated 11 healthy adults with broad-spectrum antibiotics—metronidazole, neomycin, and vancomycin—for five days. On day four of the trial, their subjects and 11 untreated controls received the seasonal influenza vaccine. The people who got antibiotics had a corresponding drop in gut microbe diversity and bacterial load. But when the researchers monitored antibody generation in response to the vaccine, they didn’t see much of a difference in the treated and untreated groups.

There has been “precious little causal evidence in humans in which there has been a deliberate and intentional perturbation of the microbiome and then a consequent examination of what this does to human physiology.” —Bali Pulendran, Stanford University

Because the flu antibody levels in both groups were pretty high to start with, next the authors recruited 11 people who hadn’t had the seasonal flu shot or been sick from the virus in the previous three years. Five of these subjects received the five-day course of broad-spectrum antibiotics, and six served as untreated controls. Everyone got the flu vaccine on day four. This time, the research team observed a marked difference in vaccine-induced immunity between the two groups. Treated subjects made far fewer flu-specific antibodies than their untreated peers, an effect that persisted through the 90 days of monitoring. “If the immune system has very poor immune memory or imprinting, then it is far more susceptible to the effects of the perturbation of the gut microbiota by antibiotics,” Pulendran explains.

“This is for me a first step and [presents] some really compelling data showing that in certain individuals, especially those that have low preexisting immunity, change to microbiota can have profound consequences on the protective response developed in the context of vaccines,” says Yasmine Belkaid, an immunologist at the National Institute of Allergy and Infectious Diseases who did not participate in the work. “What would be really fascinating is to try to understand more [about] the mechanism by which the microbiota does control” immune responses in humans, she adds.

It is possible that people’s immune systems are sensing bacterial flagellin, just like the researchers found in mice. Directly translating the mechanism is difficult, though, because influenza was new to the animals’ immune systems, which is almost never the case for people.

Because they saw such a striking difference in the role for the microbiome in people lacking immune memory of the flu, Pulendran and his team are investigating what the relationship between gut bacteria, vaccines, and a completely new immune challenge—such as a virus the immune system’s never seen before. “The results of this relatively small Phase 1 study demonstrate that the gut microbiota has an important influence on immune physiology in humans and, in particular, the immune response to vaccination,” he says, “but this needs to be explored more broadly in the context of other vaccines [and] in the context of different populations.”

T. Hagan et al., “Antibiotics-driven gut microbiome perturbation alters immunity to vaccines in humans,” Cell, doi:10.1016/j.cell.2019.08.010, 2019.

Abby Olena is a freelance journalist based in North Carolina. Find her on Twitter @abbyolena.