Artificial bacteria could help protect the “good”

CAMBRIDGE, MA — Antibiotics are life-saving medicines, but they can also harm beneficial microbes that live in the human gut. Following antibiotic treatment, some patients are at risk of developing inflammation or opportunistic infections such as Clostridiodes difficult. The indiscriminate use of antibiotics on gut microbes can also contribute to the spread of drug resistance.

In an effort to reduce these risks, MIT engineers have developed a new way to help protect the natural flora of the human digestive tract. They took a strain of bacteria safe for human consumption and engineered it to safely produce an enzyme that breaks down a class of antibiotics called beta-lactams. These include ampicillin, amoxicillin, and other commonly used drugs.

When this “living biotherapeutic” is given with antibiotics, it protects the microbiota in the gut but allows circulating blood levels of antibiotics to remain high, researchers found in a mouse study.

“This work shows that synthetic biology can be harnessed to create a new class of therapies designed to reduce the adverse effects of antibiotics,” says James Collins, Termeer Professor of Medical Engineering and Science at the Institute of Engineering and Science. Medical Sciences (IMES) from MIT. and Department of Biological Engineering, and the lead author of the new study.

Andres Cubillos-Ruiz PhD ’15, a researcher at IMES and the Wyss Institute for Biologically Inspired Engineering at Harvard University, is the lead author of the paper, which appears today in Nature Biomedical Engineering. Other authors include MIT graduate students Miguel Alcantar and Pablo Cardenas, Wyss Institute staff scientist Nina Donghia, and Broad Institute research scientist Julian Avila-Pacheco.

Protect the gut

Over the past two decades, research has revealed that microbes in the human gut play an important role not only in metabolism, but also in immune function and nervous system function.

“Throughout your lifetime, these gut microbes come together into a very diverse community that performs important functions in your body,” says Cubillos-Ruiz. “The problem arises when interventions such as medications or particular diets affect the composition of the microbiota and create an altered state, called dysbiosis. Some microbial groups disappear and the metabolic activity of others increases. This imbalance can lead to various health problems.

A major complication that can occur is infection of It’s hard, a microbe that usually lives in the gut but does not usually cause harm. When antibiotics kill strains that compete with It’s hardhowever, these bacteria can take over and cause diarrhea and colitis. It’s hard infects about 500,000 people each year in the United States and causes about 15,000 deaths.

Doctors sometimes prescribe probiotics (mixtures of beneficial bacteria) for people taking antibiotics, but these probiotics are usually also sensitive to antibiotics and do not fully replicate the native microbiota present in the gut.

“Standard probiotics cannot be compared to the diversity of native microbes,” says Cubillos-Ruiz. “They can’t perform the same functions as the native microbes you’ve nurtured throughout your life.”

To protect the microbiota from antibiotics, the researchers decided to use modified bacteria. They engineered a strain of bacteria called Lactococcus lactis, which is normally used in cheese production, to deliver an enzyme that breaks down beta-lactam antibiotics. These drugs account for approximately 60% of the antibiotics prescribed in the United States.

When these bacteria are given orally, they transiently populate the intestines, where they secrete the enzyme called beta-lactamase. This enzyme then breaks down antibiotics that reach the intestinal tract. When antibiotics are taken orally, the drugs enter the bloodstream primarily from the stomach, so the drugs can still circulate through the body at high levels. This approach could also be used with injected antibiotics, which also eventually reach the gut. Once their work is done, the modified bacteria are excreted through the digestive tract.

The use of engineered bacteria that degrade antibiotics poses unique safety requirements: beta-lactamase enzymes confer antibiotic resistance to host cells and their genes can easily spread between different bacteria. To solve this problem, the researchers used a synthetic biology approach to recode how the bacterium synthesizes the enzyme. They split the beta-lactamase gene into two pieces, each coding for a fragment of the enzyme. These gene segments are located on different pieces of DNA, making it highly unlikely that both gene segments will be transferred to another bacterial cell.

These beta-lactamase fragments are exported out of the cell where they reassemble, restoring enzyme function. Since beta-lactamase is now free to diffuse into the surrounding environment, its activity becomes a “public good” for the intestinal bacterial communities. This prevents the modified cells from gaining an advantage over the native gut microbes.

“Our biocontainment strategy allows the delivery of antibiotic-degrading enzymes into the gut without the risk of horizontal gene transfer to other bacteria or gaining an additional competitive advantage through live biotherapy,” says Cubillos- Ruiz.

Maintain microbial diversity

To test their approach, the researchers gave the mice two oral doses of the modified bacteria for each injection of ampicillin. The modified bacteria traveled to the gut and started releasing beta-lactamase. In these mice, the researchers found that the amount of ampicillin circulating in the blood was as high as that of mice that had not received the modified bacteria.

In the gut, mice that received modified bacteria maintained a much higher level of microbial diversity than mice that received only antibiotics. In these mice, levels of microbial diversity dropped dramatically after receiving ampicillin. Moreover, none of the mice that received the modified bacteria developed opportunism. It’s hard infections, while all mice that received only antibiotics showed high levels of It’s hard in the intestine.

“It’s a strong demonstration that this approach can protect the gut microbiota, while preserving the effectiveness of the antibiotic, because you’re not changing the levels in the bloodstream,” says Cubillos-Ruiz.

The researchers also found that removing the evolutionary pressure of antibiotic treatment made it much less likely that gut microbes would develop antibiotic resistance after treatment. In contrast, they found many antibiotic resistance genes in the microbes that survived in the mice that received antibiotics, but not the modified bacteria. These genes can be passed on to harmful bacteria, compounding the problem of antibiotic resistance.

The researchers now plan to start developing a version of the treatment that could be tested in people at high risk of developing acute illnesses that stem from antibiotic-induced gut dysbiosis, and they hope that eventually it could be used. to protect anyone who needs to take antibiotics for infections outside of the gut.

“If antibiotic action is not needed in the gut, then you need to protect the microbiota. It’s like when you have an X-ray, you wear a lead apron to protect the rest of your body from ionizing radiation,” says Cubillos-Ruiz. “No previous intervention could offer this level of protection. With our new technology, we can make antibiotics safer by preserving beneficial gut microbes and reducing the chances of new antibiotic-resistant variants emerging.

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The research was funded by the Defense Threat Reduction Agency, the Paul G. Allen Frontiers Group, the Wyss Institute, and a National Science Foundation postgraduate research grant.