ANTIBIOTICS GIVE RISE TO NEW COMMUNITIES OF HARMFUL BACTERIA

Most people have taken an antibiotic to treat a bacterial infection. Now researchers from the University of North Carolina at Chapel Hill and the University of San Diego, La Jolla, reveal that the way we often think about antibiotics -- as straightforward killing machines -- needs to be revised.

 The work, led by Elizabeth Shank, an assistant professor of biology in the UNC-Chapel Hill College of Arts and Sciences as well as microbiology and immunology in the UNC-Chapel Hill School of Medicine, and Rachel Bleich, a graduate student in the UNC-Chapel Hill Eshelman School of Pharmacy, not only adds a new dimension to how we treat infections, but also might change our understanding of why bacteria produce antibiotics in the first place.

"For a long time we've thought that bacteria make antibiotics for the same reasons that we love them -- because they kill other bacteria," said Shank, whose work appears in the February 23 Early Edition of the Proceedings of the National Academy of Sciences. "However, we've also known that antibiotics can sometimes have pesky side-effects, like stimulating biofilm formation."

Shank and her team now show that this side-effect -- the production of biofilms -- is not a side-effect after all, suggesting that bacteria may have evolved to produce antibiotics in order to produce biofilms and not only for their killing abilities.

Biofilms are communities of bacteria that form on surfaces, a phenomenon dentists usually refer to as plaque. Biofilms are everywhere. In many cases, biofilms can be beneficial, such as when they protect plant roots from pathogens. But they can also harm, for instance when they form on medical catheters or feeding tubes in patients, causing disease.

"It was never that surprising that many bacteria form biofilms in response to antibiotics: it helps them survive an attack. But it's always been thought that this was a general stress response, a kind of non-specific side-effect of antibiotics. Our findings indicate that this isn't true. We've discovered an antibiotic that very specifically activates biofilm formation, and does so in a way that has nothing to do with its ability to kill."

Shank and her team previously reported that the soil bacterium Bacillus cereus could stimulate the bacterium Bacillus subtilis to form a biofilm in response to an unknown secreted signal. B. subtilis is found in soil and the gastrointestinal tract of humans.

Using imaging mass spectrometry, they subsequently identified the signaling compound that induced biofilm production as thiocillin, a member of a class of antibiotics called thiazolyl peptide antibiotics, which are produced by a range of bacteria.

At that point, Shank and her colleagues knew thiocillin had two very specific and different functions, but they didn't know why -- and wanted to know how it worked. That's when they modified thiocillin's structure in a way that eliminated thiocillin's antibiotic activity, but did not halt biofilm production.

"That suggests that antibiotics can independently and simultaneously induce potentially dangerous biofilm formation in other bacteria and that these activities may be acting through specific signaling pathways," said Shank. "It has generated further discussion about the evolution of antibiotic activity, and the fact that some antibiotics being used therapeutically may induce biofilm formation in a strong and specific way, which has broad implications for human health."

GUT BACTERIA THAT PROTECT AGAINST FOOD ALLERGIES

The presence of Clostridia, a common class of gut bacteria, protects against food allergies, a new study in mice finds. By inducing immune responses that prevent food allergens from entering the bloodstream, Clostridia minimize allergen exposure and prevent sensitization -- a key step in the development of food allergies. The discovery points toward probiotic therapies for this so-far untreatable condition, report scientists from the University of Chicago, Aug 25 in the Proceedings of the National Academy of Sciences

Although the causes of food allergy -- a sometimes deadly immune response to certain foods -- are unknown, studies have hinted that modern hygienic or dietary practices may play a role by disturbing the body's natural bacterial composition. In recent years, food allergy rates among children have risen sharply -- increasing approximately 50 percent between 1997 and 2011 -- and studies have shown a correlation to antibiotic and antimicrobial use.

"Environmental stimuli such as antibiotic overuse, high fat diets, caesarean birth, removal of common pathogens and even formula feeding have affected the microbiota with which we've co-evolved," said study senior author Cathryn Nagler, PhD, Bunning Food Allergy Professor at the University of Chicago. "Our results suggest this could contribute to the increasing susceptibility to food allergies."

To test how gut bacteria affect food allergies, Nagler and her team investigated the response to food allergens in mice. They exposed germ-free mice (born and raised in sterile conditions to have no resident microorganisms) and mice treated with antibiotics as newborns (which significantly reduces gut bacteria) to peanut allergens. Both groups of mice displayed a strong immunological response, producing significantly higher levels of antibodies against peanut allergens than mice with normal gut bacteria.

This sensitization to food allergens could be reversed, however, by reintroducing a mix of Clostridia bacteria back into the mice. Reintroduction of another major group of intestinal bacteria, Bacteroides, failed to alleviate sensitization, indicating that Clostridia have a unique, protective role against food allergens.

Closing the door

To identify this protective mechanism, Nagler and her team studied cellular and molecular immune responses to bacteria in the gut. Genetic analysis revealed that Clostridia caused innate immune cells to produce high levels of interleukin-22 (IL-22), a signaling molecule known to decrease the permeability of the intestinal lining.

Antibiotic-treated mice were either given IL-22 or were colonized with Clostridia. When exposed to peanut allergens, mice in both conditions showed reduced allergen levels in their blood, compared to controls. Allergen levels significantly increased, however, after the mice were given antibodies that neutralized IL-22, indicating that Clostridia-induced IL-22 prevents allergens from entering the bloodstream.

"We've identified a bacterial population that protects against food allergen sensitization," Nagler said. "The first step in getting sensitized to a food allergen is for it to get into your blood and be presented to your immune system. The presence of these bacteria regulates that process." She cautions, however, that these findings likely apply at a population level, and that the cause-and-effect relationship in individuals requires further study.

While complex and largely undetermined factors such as genetics greatly affect whether individuals develop food allergies and how they manifest, the identification of a bacteria-induced barrier-protective response represents a new paradigm for preventing sensitization to food. Clostridia bacteria are common in humans and represent a clear target for potential therapeutics that prevent or treat food allergies. Nagler and her team are working to develop and test compositions that could be used for probiotic therapy and have filed a provisional patent.

"It's exciting because we know what the bacteria are; we have a way to intervene," Nagler said. "There are of course no guarantees, but this is absolutely testable as a therapeutic against a disease for which there's nothing. As a mom, I can imagine how frightening it must be to worry every time your child takes a bite of food."

"Food allergies affect 15 million Americans, including one in 13 children, who live with this potentially life-threatening disease that currently has no cure," said Mary Jane Marchisotto, senior vice president of research at Food Allergy Research & Education. "We have been pleased to support the research that has been conducted by Dr. Nagler and her colleagues at the University of Chicago."

GUT BACTERIA ARE PROTECTED BY HOST DURING ILLNESS

To protect their gut microbes during illness, sick mice produce specialized sugars in the gut that feed their microbiota and maintain a healthy microbial balance. This protective mechanism also appears to help resist or tolerate additional harmful pathogens, and its disruption may play a role in human diseases such as Crohn's disease, report scientists from the University of Chicago in Nature on Oct 1.

Both hosts and their gut microbiota can suffer in the case of sickness, but this mutually beneficial relationship is guarded by the host," said study senior author Alexander Chervonsky, MD, PhD, chairman of the Committee on Immunology at the University of Chicago.

When faced with systemic illness, animals eat less to conserve energy instead of foraging for food and to deprive pathogens of nutrients. However, this can harm beneficial gut bacteria, which have an important role in health and disease.

To investigate how microbiota might be supported during illness, Chervonsky and his team focused on a potential internal resource produced by the host -- L-fucose, a sugar which has been shown to affect gut microbes. A host cannot use L-fucose for energy, but when bound to proteins, it can be used by microbes as a food source. Under normal conditions, however, the small intestine of mice produces almost no L-fucose.

The team exposed different types of mice to a molecule that mimicked a systemic infection. The mice became sick -- eating less food, drinking less water and losing weight. Only a few hours after this induced sickness, the researchers observed that L-fucose was produced and present on almost every surface of the small intestine. This effect was seen only in response to illness.

The researchers then tested genetically engineered mice lacking Fut2, the gene responsible for L-fucose production. Healthy under normal conditions, mice without Fut2 regained weight after induced sickness -- a measure of recovery -- much slower than their normal counterparts. However, only mice with both intact gut microbiota and the ability to produce L-fucose recovered efficiently.

"Mice that can produce L-fucose recover better than those that can't," Chervonsky said. "If you remove bacteria the effect goes away."

The team used genetic analyses to confirm that gut microbes were affected metabolically by the production of L-fucose. As part of this analysis, they noted that sick mice without Fut2 had significantly greater expression of harmful microbial genes than normal mice. Hypothesizing that L-fucose production was somehow preventing opportunistic bacteria from expressing virulent genes, they exposed mice to a mild bacterial pathogen and then four days later induced sickness. Under this condition, mice without Fut2 lost significantly more weight than normal, suggesting that the production of L-fucose helps the host tolerate or resist additional harmful pathogens.

Interestingly, around 20 percent of humans lack a functional gene to produce L-fucose, a problem that has been associated with the inflammatory bowel ailment known as Crohn's disease.

"We speculate that without L-fucose, the activation of virulence genes cannot be blocked, and that's why bacteria play a role in Crohn's disease," Chervonsky said. "Whether we can use this toward therapeutics in the future requires further study."