Keeping the upper hand in the battle with bacteria

Attending critical care physician Deborah Hung knows first-hand the threat of drug-resistant infections. In the Medical Intensive Care Unit at Brigham and Women’s Hospital in Boston, Hung often cares for patients fighting dangerous bacterial infections. Sadly, some of her patients learn that among the array of antibiotic drugs on the pharmacist’s shelf, none can overcome the particular bacteria attacking their bodies.

“In the intensive care unit, I have had to tell a patient or explain to a family that the patient has an infection that is untreatable with antibiotics – that the organism has become resistant to everything we have and there is nothing more we can do,” recalled Hung, who is also co-director of the Infectious Disease and Microbiome Program at the Broad Institute of MIT and Harvard. “The notion that we don’t have antibiotics left to treat [some] infections…is really quite shocking to patients and families.”

Credit: Centers for Disease Control and Prevention

Soon after antibiotics were first given to patients more than 70 years ago, these weapons against infection began losing firepower as disease-causing bacteria quickly evolved resistance to them. Today, many types of bacteria that were once easily treated with cheap medicines have adapted to become impervious to our best pharmacological weapons. Some bacteria (also known as microbes) can now only be conquered with second- or third-line drugs that can be expensive and carry toxic side effects, and still others have become resistant to all drugs available, giving infected patients no treatment options. According to a 2013 report from the Centers for Disease Control and Prevention, each year in the United States at least 2 million people become infected with drug-resistant bacteria and at least 23,000 people die from those infections, with most deaths happening in healthcare settings such as hospitals and nursing homes.

Preventing a post-antibiotic era

The rise of drug resistance stems from misuse and overuse of antibiotics in the clinic and in agriculture. With few new antibiotics in the pipeline and drug-resistant illnesses spreading easily in an increasingly globalized world, the problem calls for new perspectives and new solutions.

At the Broad Institute, Hung and her colleagues take a variety of genomic and biochemical approaches to find new ways of treating, diagnosing, tracking, and preventing drug-resistant infections. The scientists are attacking the problem from all sides, looking into the microbial genome to understand how bacteria become resistant, exploring new methods to quickly diagnose infections, and devising ways of making antibiotics work better.

“It is going to take this type of innovation and perspective to think about new paradigms for antibiotic discovery, new paradigms to treat infection, and really a very different way to think about infection,” said Hung. “The old way of reaching for an antibiotic and thinking the problem is solved, that time is gone.”

Revealing the roots of resistance

Microbes have an array of defensive tactics used to evade treatment — e.g., making enzymes that inactivate the drug, altering cell surface receptors to prevent drug binding, or even actively pumping the drug out of the bacterial cell — but researchers don’t fully understand exactly how bacteria acquire those abilities. A better picture of the evolution of drug resistance can shed light on how it should be diagnosed and how it might even be prevented.

The two main ways that bacteria become genetically resistant to drugs are 1) by acquiring resistance genes from other microbes through so-called “horizontal gene transfer,” or 2) through mutation of their own genes. In a recent study appearing in eLife, Hung and her colleagues coaxed bacteria to develop antibiotic resistance in the lab, so they could track which genes mutate as the cells become drug-resistant. Focusing on Mycobacterium smegmatis, a cousin of the tuberculosis bacterium that becomes resistant through mutations to its own genes, the researchers exposed the microbes to low concentrations of different antibiotics that slowly kill the bacteria.

Some of these cultures developed antibiotic-resistant colonies and in those, the scientists found single mutations in genes encoding different parts of the ribosome, part of the cell’s molecular machinery that helps build proteins. Surprisingly, they found these mutations in bacteria that were exposed to antibiotics that don’t even target the ribosome. The mutations come at a cost, making the bacteria grow slower, but they give the bacteria a survival advantage when exposed to several different antibiotics and make it more likely the bacteria will develop high-level, multi-drug resistance. This and other efforts to learn how bacteria evolve resistance will help preserve the utility of current and future drugs.

On a much larger scale, Broad scientists are surveying pathogenic bacteria from around the globe, to uncover patterns of resistance and, specifically, to track the order in which mutations occur in bacteria that become resistant to more than one drug. One problematic infection found around the globe is multidrug-resistant tuberculosis (MDR-TB), which is impervious to the two widely used first-line tuberculosis drugs, isoniazid and rifampicin. In 2015, 10.4 million fell ill with TB (an estimated 480,000 of those with MDR-TB) and nearly two million died from the disease. MDR-TB can sometimes be cured with second-line drugs, but they are expensive and toxic and must be taken for up to two years.

For a closer look at how tuberculosis becomes impervious to multiple drugs, a group led by Broad scientist Ashlee Earl teamed up with other international TB researchers to collect bacterial isolates from more than 5,000 people around the world. After analyzing the whole-genome sequences of these samples, they discovered that DNA mutations conferring resistance to isoniazid usually emerge first, followed by resistance to rifampicin. Because the current frontline molecular test used to detect drug-resistant TB only looks for rifampicin resistance, patients who have isoniazid-resistant TB may be assumed to have drug-sensitive TB and may be mistakenly given a regimen containing isoniazid, which won’t help them get better. Inappropriate treatment may also allow these bacteria to develop further resistance and become MDR-TB, which is much harder to treat than TB resistant to a single drug. Appearing in Nature Genetics, the study suggests that incorporating tests for these “harbinger mutations,” some of which confer isoniazid resistance, could greatly help efforts to control TB worldwide.

“Just as it's important to understand weather patterns so you can prepare and protect yourself from threatening weather, understanding the factors that drive the rise and spread of antibiotic resistance — and monitoring for those conditions — can help us be prepared and intervene as threats arise,” explained Earl.

Tracking the spread of superbugs in hospitals

Drug-resistant infections are especially dangerous in settings like hospitals and nursing homes, where patients often have weak immune systems or other illnesses. One threat that is particularly urgent is the carbapenem-resistant Enterobacteriaceae (CRE), which are Klebsiella species and E. coli that are resistant to all or nearly all the antibiotics we have today, including last-resort drugs known as carbapenems. In February 2017, the World Health Organization placed CRE bacteria atop its list of drug-resistant pathogens that pose the greatest risk to human health. The ways CRE evolve, diversify, and spread are not fully understood, hindering efforts to design appropriate strategies to slow or contain the pathogens’ spread.

The bacteria on the left plate are drug-sensitive and unable to grow adjacent to the discs of antibiotics. On the right plate, carbapenem-resistant Enterobacteriaceae (CRE) bacteria are resistant to the antibiotics tested and are able to grow near the discs.
Credit: Centers for Disease Control and Prevention, James Gathany

A multi-institutional collaboration including Broad associate member William Hanage and Broad scientists Cheryl Murphy, Gustavo Cerqueira and Ashlee Earl took a deep look at the DNA of CRE isolated from blood, wounds, urine, and respiratory tracts of patients in three Boston-area hospitals and one in California. CREs are so dangerous, in part, because they can readily transfer resistance genes among bacterial species, such as between Klebsiella and E. coli. Writing in PNAS, the researchers report little evidence of spread of resistant bacteria between patients in each hospital and more variation in resistance mechanisms and families of resistant bacteria than expected. Interestingly, they also found carbapenem-resistant bacteria that didn’t carry the usual suspects of resistance genes, suggesting unknown resistance genes or mechanisms still to be discovered. In light of the results, efforts to control CRE infections must include surveillance and monitoring of strains, even among asymptomatic carriers.

For better treatment, wake bacterial cells up

Some bacteria that would otherwise be susceptible to antibiotics can survive exposure to the drugs by restricting their cellular metabolism, i.e., becoming “metabolically dormant.” Scientists like Broad researcher Jim Collins suspect that stimulating the metabolism of dormant bacteria could help overcome this “phenotypic tolerance” and allow antibiotics to do their job.

In Cell Chemical Biology, Collins and his fellow researchers recently described their effort to tune the antibiotic susceptibility of Pseudomonas aeruginosa, a pathogen that plagues cystic fibrosis (CF) patients with difficult-to-treat lung infections. Once a chronic P. aeruginosa infection is established, it is nearly impossible to clear with antibiotics, due to the way bacteria respond to the unique environment in the airways of CF patients.

To uncover the biochemical basis for P. aeruginosa’s tolerance to aminoglycoside antibiotics, the researchers stimulated cultures of the bacteria with different carbon metabolites. They found that cellular respiration modulates the effects of tobramycin, a cornerstone CF drug, and that the carbon metabolite glyoxylate promotes tolerance to tobramycin while the metabolite fumarate promotes drug susceptibility. By shedding light on the non-genetic ways that bacteria withstand drugs, the work suggests that metabolites could be used clinically to make antibiotics better at killing these tolerant cells.

The more quickly we know the enemy, the better

Improper use of antibiotics in the clinic can lead to resistance, but physicians aren’t always able to prescribe the right medicine for each infection. A definitive diagnosis relies upon accurately identifying the pathogenic bug by culturing it in the lab, which takes valuable time. If a physician wants to start treating an infection quickly, they must either prescribe broad-spectrum antibiotics that kill many types of bacteria – good and bad – or make an educated guess and prescribe a narrow-spectrum drug that may target the wrong microbe. And if the patient has a viral infection, antibiotics may be used needlessly.

New tests to rapidly diagnose bacterial infections in the clinic are crucial to controlling the growth of antibiotic resistance. It is important for physicians to know the infectious microbe’s identity, to discern whether it is drug-sensitive or drug-resistant, and to distinguish between pathogens and healthy, beneficial microbes. Roby Bhattacharyya and other Broad scientists are developing methods that analyze the sequence and abundance of bacterial RNA to identify the infectious organisms and help guide treatment decisions, efforts that could make a huge impact clinically.

Broad core institute member Paul Blainey is working on other technological solutions aimed at speeding up the sequencing of microbes, which can help with surveillance of drug-resistant infections. Because the lengthy and costly steps of sample preparation are still a bottleneck to rapid microbial sequencing, Blainey and his colleagues have devised a general-purpose, automated microfluidic system that enables higher-throughput sample preparation from whole cells for sequencing with less starting material, without sacrificing data quality. Featured in Nature Communications, the system could help overcome technical barriers to using genomics in the battle against antibiotic resistance, among other applications.

With innovative approaches and technological solutions, scientists at the Broad Institute and elsewhere hope to preserve the utility of our wonder drugs and prevent a return to the pre-antibiotic era. Antibiotic drugs have made an incredible impact on human health. Through biological understanding, improved surveillance, and new treatment strategies, we may be able to keep the upper hand in our battle with these nimble foes.