New antimicrobial shuts down bacterial growth without harming human cells

Graphic shows how the KKL-55 molecule (in red) inhibits trans-translation, a process that bacteria cells use for quality control during protein synthesis.

By Carol Clark

Scientists have shown how a molecule with broad-spectrum antibiotic activity works by disabling a process vital to bacterial growth without affecting the normal functioning of human cells. mBio, a journal of the American Society for Microbiology, published the work, led by researchers at Emory University and Pennsylvania State University. 

The molecule, known as KKL-55, is one of a suite of recently identified molecules that interfere with a key bacterial mechanism known as trans-translation, essentially shutting down the ability of bacteria to grow. 

“We’re opening a promising pathway for the development of new antibiotics to treat drug-resistant infections,” says Christine Dunham, co-corresponding author of the paper and a professor in Emory’s Department of Chemistry and the Emory Antibiotic Resistance Center. 

Kenneth Keiler, a professor in the Department of Biochemistry and Molecular Biology at Pennsylvania State, is co-corresponding author of the paper. 

First authors are Ha An Nguyen, who did the work as an Emory chemistry PhD candidate and has since graduated and works at Memorial Sloan Kettering, and Neeraja Marathe, a graduate student at Pennsylvania State. 

A growing global threat 

Antimicrobial-resistant infections have long been a public health threat. The situation grew even worse during the COVID-19 pandemic with increased antibiotic use and less prevention actions, according to the U.S. Centers for Disease Control and Prevention (CDC). 

The CDC estimates that at least 2.8 million antimicrobial-resistant infections continue to occur in the United States each year, killing more than 35,000 people. Globally, the World Health Organization projects that these infections will cause up to 10 million deaths annually by 2050 if new antibiotics are not developed. 

While antibiotics can save lives, any time they are used they can also contribute to the problem of resistance. Bacteria keep evolving new weapons as a defense against drugs, even as scientists work on developing new strategies to disarm bacteria. 

Cross-toxicity, or harmful effects on humans, is another key drawback of some of the drugs used in a last-ditch effort to kill antibiotic-resistant bacteria. 

Avoiding cross-toxicity 

Dunham and Keiler are avoiding the problem of cross-toxicity by focusing on the inhibition of a mechanism unique to bacteria — trans-translation. This mechanism is vital to the proper functioning of the bacterial ribosome. 

Keiler, a molecular geneticist and biochemist, first identified trans-translation in bacteria and is an expert in how it functions. Dunham, a structural biologist, is an expert in the human ribosome. She uses advanced biochemistry and structural biology techniques to understand the mechanics of molecular interactions. 

“Our individual areas of expertise mesh well for this project,” Dunham says. “By collaborating, we are able to take the science further, faster.” 

A cellular protein factory 

The ribosome is an elaborate macromolecular machine within a cell that operates like a factory to manufacture proteins. Proteins are the machines that make cells run while nucleic acids such as DNA and RNA store the blueprints for life. The ribosome is made mostly of RNA, which does not just store information but can also act as an enzyme, catalyzing chemical reactions. 

In a human cell, messenger RNA (mRNA), containing the instructions for manufacturing a protein, originates in the nucleus. While still in the nucleus, mRNA undergoes an elaborate quality-control process. It must pass inspection before getting exported to translate the information it contains into a protein. 

“A lot of mRNAs have defects,” Dunham says. “Human cells have efficient ways to test mRNAs and ultimately remove the defective ones.” 

Bacterial cells, however, have no nucleus or organized center for quality control. 

“Bacteria wants to grow, grow and grow, which requires the ribosome to make a lot of proteins,” Dunham says. “But when mRNA has defects, there is little to no quality control. When the ribosome encounters a defective mRNA protein, synthesis gets stalled.” 

The trans-translation process “rescues” ribosomes stalled due to such defects, in order to maintain proper protein synthesis and cell viability in bacteria. 

How KKL-55 works 

Using a high-throughput screening process, the Keiler lab has identified dozens of molecules that inhibit trans-translation in bacteria. 

For the current paper, the researchers focused on understanding how one of these molecules, KKL-55, performs this trick. They used the high-powered structural biology technique of X-ray crystallography to capture KKL-55 in action as it interacted with a protein required for translation. 

The results showed how KKL-55 blocks trans-translation by binding to elongation factor thermos-unstable (EF-Tu). EF-Tu is a protein that interacts with transfer RNA molecules, which play a key role in protein synthesis, and also transfer-messenger RNA, an RNA molecule required for the trans-translation pathway. 

“We got lucky,” Dunham says. “There are dozens of steps involved in the process that KKL-55 could have inhibited and we might have had to test for each one. But the results are clear-cut. It shuts down trans-translation right at the beginning by preventing EF-Tu from binding to tmRNA.” 

Determining the mechanism by which a molecule works to inhibit bacteria is a critical step toward developing a new antibiotic for clinical use. The next step is to test the efficacy of KKL-55 to treat a bacterial infection in a mouse model. 

In 2021, the research team published their finding that a group of trans-translation inhibitors known as acylaminooxadiazoles clear multiple-drug-resistant Neisseria gonorrhoeae infection in mice after a single oral dose. That work is now advancing to clinical trials. 

Dozens more trans-translation inhibitors await the team’s investigation. Each represents a potential new weapon to help humans stay on top in the arms race with drug-resistant bacteria. 

Co-authors of the current paper include Alexandra Nagy (a former National Institutes of Health FIRST Institutional Research and Academic Career Development postdoctoral fellow at Emory who is now at Earlham College); as well as John Alumasa and Michael Vazquez (both from Pennsylvania State University). The work was funded by the National Institutes of Health, the National Institute of General Medical Sciences. 

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Emory breaking new ground for climate-smart agriculture in the Southeast

"Our project is unique in that it focuses on the Southern Piedmont and an often under served piece of our food system, but one that is vital to providing us the nutrients we need — the vegetable sector," says Emily Burchfield, assistant professor of environmental sciences.

Three Emory University researchers received $5,100,000 as part of a United States Department of Agriculture (USDA) project to help measure and promote climate-smart practices that support small-scale, diversified vegetable farmers in the Southern Piedmont. A plateau below the Appalachian Mountains and above the coastal plain, the Southern Piedmont is a banana-shaped region spanning a bit of eastern Alabama, up across part of northern Georgia and into North and South Carolina and Virginia. 

Emory is one of 12 organizations involved in the $25 million project, headed by the Rodale Institute and titled “Quantifying the Potential to Reduce Greenhouse Gas Emissions and Increase Carbon Sequestration by Growing and Marketing Climate-Smart Commodities in the Southern Piedmont.” 

The five-year project is part of the USDA’s Partnerships for Climate-Smart Commodities initiative. “This effort will increase the competitive advantage of U.S. agriculture both domestically and internationally, build wealth that stays in rural communities and support a diverse range of producers and operation types,” USDA Secretary Tom Vilsack says of the initiative. 

The Emory team encompasses three faculty from the Department of Environmental Sciences: Emily Burchfield, Eri Saikawa and Debjani Sihi. 

• Burchfield combines spatial-temporal social and environmental data to understand the future of food security in the United States. 

• Saikawa is an atmospheric chemist who models global soil nitrous oxide emissions and quantifies soil greenhouse gas fluxes. 

• Sihi is an environmental biogeochemist who researches soil organic matter dynamics and greenhouse gas emissions from natural and managed systems. 

Read more here.

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Math trio makes new points about size of the smallest triangle

"It's a very rich area, to study tiny, small-scale shapes and uncover what the math hidden there can tell us," says Emory mathematician Cosmin Pohoata.

By Carol Clark

Think of a square dotted with points. Now imagine the smallest triangle that could be made by connecting three of those points. That’s the Heilbronn triangle problem in a nutshell. 

“The problem is very easy to state and can sound frivolous,” says Cosmin Pohoata, a theoretical mathematician and Emory assistant professor of mathematics “When I have conversations with non-math friends, they often ask me why we should study problems like this. The beauty of them is that they are often more complex than they seem. They can have unexpected connections that open new doors for understanding all sorts of phenomena.” 

 

Pohoata and two MIT graduate students, Alex Cohen and Dimitrii Zakharov, recently opened some of those new doors. They completed a new proof for the Heilbronn triangle problem that shows that the smallest triangle in a confined space is much smaller than was previously realized, breaking a record that stood for 40 years. 

 

Their proof, available online, is submitted to the Journal of the American Mathematical Society and is already making waves in the math world. 

“I think it’s a stunning result,” Anthony Carbery, a mathematician at the University of Edinburgh, told Qanta Magazine. And Thomas Bloom of the University of Oxford told Qanta that he expects the new proof to “prompt a renaissance” of progress on the triangle problem. 

 

“What makes the proof special to me,” Pohoata says, “is the way that we connected the triangle problem to different areas of math. In particular, harmonic analysis, the study of how waves interact with one another and projection theory, or the behavior of fractals under projections.” 

A graphic representation of the Heilbronn triangle problem.

The making of a mathematician 

Pohoata loved math from the time he was a small child growing up in Romania. He cites an elementary school teacher, who encouraged his love for numbers and patterns, as one key influence. 

In middle school he began competing in International Mathematical Olympiads (IMO). Romania is the original home of the IMO, which dates back to 1959, making it the oldest of the International Science Olympiads. Today more than 100 countries compete in the annual event. 

“I thought I knew a lot about math because the problems in class had been so easy for me,” Pohoata recalls. “When I started competing in the Olympiads, I began to realize how little I knew and how much math was out there to learn.” 

He began to think about math as a career. “It’s quite fun to get to think about problems that interest you,” he says. 

Pohoata attended Princeton as an undergraduate, got his PhD at the California Institute of Technology and taught at Yale before joining Emory this fall. 

Patterns in points and lines 

As a theoretical mathematician, Pohoata focuses his research on three specialized fields: Discrete geometry, additive number theory and extremal combinatorics. 

Extremal combinatorics examines how large or small finite objects such as graphs can be, if placed under certain restrictions. For centuries, it seemed like an esoteric endeavor. 

“The breadcrumbs to extremal combinatorics trace back to ancient Greece,” Pohoata says, “but the field didn’t really come alive until the late 20th century with the rise of computers and the internet. Graphs are at the heart of many things to do with computer science and the internet.” 

Facebook “friend” networks, for example, are large collections of data that are essentially graphs. “You can think of people in the world as points on paper and then draw arrows connecting the ones who are friends,” Pohoata explains. “Then you can look at basic questions underlying these structures. If you have at least seven points do you always have triangles? When do you see a big cluster of connected vertices? Are there areas of the world that are less connected than others?” 

Real-life problems, like how to make algorithms run faster, fuel interest in studying problems about graphs and related areas. 

“As a theorist, I’m driven simply by the math behind shapes and the beauty of them,” Pohoata says, “but I do get excited when I hear that some math breakthrough has been used in a cool way to help with a practical problem.” 

The human side of math 

“I wasn’t interested in the history of math when I first started out,” Pohoata says. “Why learn the progression of results if you have the latest result?” 

But when he taught an introductory course to number theory, it forced him to look more closely at the history of the greats and trace the chronology of events. “I started realizing that you can get many new ideas by following the progress of the past rather than just focusing on the latest thing,” he says. “And I personally learn better when I follow the story of the people in the history of math. You feel the math differently, too, when you put yourself in someone else’s shoes.” 

Pohoata’s interest in the Heilbronn triangle problem inspired him to delve deeper into the work of Klaus Roth, a German-British mathematician who won math’s highest honor, the Fields Medal. “Roth does elegant math that has inspired a lot of activity,” Pohoata says. 

In 1951, Roth developed a strategy for finding the smallest possible triangle within the parameters of the Heilbronn triangle problem, or its so-called “upper bound.” Austrian mathematician Wolfgang Schmidt pushed the upper bound further in a paper published in 1972. That inspired Roth to jump back into the game. Roth further improved the result by Schmidt, just a few months after Schmidt’s breakthrough. 

A tiny problem 

“Roth and Schmidt had a kind of rivalry to see who could come up with the best recipe to find even smaller triangles,” Pohoata says. “They were writing beautiful papers, improving on each other’s work. I learned a lot by studying them.” 

In 1980, a trio of mathematicians — Komlos, Pintz and Szemeredi — pushed the envelope even further, finding a new upper bound to the Heilbronn triangle problem. 

While the problem in its simplest form can be thought of as dots and lines drawn on paper, the mathematicians are working with triangles far too tiny to be “seen” without special tools. 

“You can think of these triangles as microscopic,” Pohoata says. “We’re talking about billions of points crammed within a confined space.” 

Just as scientists keep making improvements in microscopy to get an ever more detailed view of the tiniest parts of a living system or of distant galaxies imperceptible to human eyes on Earth, theoretical mathematicians create tools to get closer and sharper views of the math underlying the universe and everything in it. 

Making connections 

Pohoata had pondered the Heilbronn triangle problem for several months with Zakharov. He met Cohen last year in a chance encounter at MIT, where he had traveled to give a presentation. 

“When math people get together, they like to talk about recent problems on their minds,” he says. “We were excited to learn that we were taking similar approaches to the Heilbronn triangle problem. And that we were all stuck in the same place.” 

The trio decided to join forces. Unlike many of their math heroes of the past, who communicated across distances by letter, they exchanged ideas in real time through Zoom and the Discord instant-messaging platform. 

“Math research is becoming more of a social experience as the world has become more connected,” Pohoata says. “Technology facilitates collaboration.” 

Rather than a single, euphoric eureka moment, he describes the process of creating their 40-page proof as a series of smaller insights. “There are many moving pieces to this proof and each one had to come together,” Pohoata explains. “There was a lot of going back and forth to get all the pieces to fit. You can think of it like putting together a really complex Lego structure.” 

Ultimately, their breakthrough revealed new connections between the Heilbronn triangle problem and other areas of mathematics, including harmonic analysis and fractals — figures that are similar and keep repeating one another at smaller and smaller scales. 

Pohoata and his two MIT colleagues are continuing to work on explaining this web of connections in more detail. “It’s a very rich area, to study tiny, small-scale shapes and uncover what the math hidden there can tell us,” Pohoata says. “What makes math fascinating is that it’s the language for how things work in the world.” 

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Analyzing ways to help golden eagle populations weather wind-energy growth

"We are taking basic information about golden eagle ecology in the Anthropocene and developing it into predictive frameworks for how to protect them," says Eric Lonsdorf, Emory assistant professor of environmental sciences.

By Carol Clark

Wind energy is a major component of the U.S. clean-energy goals. Already one of the fastest growing and lowest-cost sources of electricity in the country, it is poised for even more rapid growth, according to the U.S. Department of Energy. 

Wind power, however, does not come without tradeoffs, including some negative impacts on wildlife. Throughout the United States, for example, it’s been estimated that as many as three golden eagles per wind farm are killed each year by wind turbines. 

“Renewable energy sources, including wind energy, are critical for us to achieve a net-zero emissions future,” says Eric Lonsdorf, assistant professor of environmental sciences at Emory University. “We need to address conflicts between renewable energy and wildlife conservation so that we can combat climate change while also limiting damage to biodiversity.” 

Lonsdorf and colleagues are developing data-driven methods to determine how much effort is needed to save golden eagles in order to offset the impact of wind turbines on their populations. 

The Journal of Wildlife Management recently published their latest model for calculating the benefit of one mitigation strategy — removal of large, road-killed animals that can lead to golden eagles getting hit by cars. 

Quantifying the benefits of natural capital

Lonsdorf is an expert in natural capital, or the quantifiable benefits that nature provides humans. He translates ecological principles and data into computer models that enable industry leaders and policymakers to better manage natural resources. 

Co-authors of the current study include James Gerber and Deepak Ray, from the University of Minnesota; Steven Slater, from HawkWatch International; and Taber Allison, from the Renewable Energy Wildlife Institute. 

The U.S. Fish and Wildlife Service (FWS) monitors golden eagle populations, which are protected through the Bald and Golden Eagle Protection Act and the Migratory Bird Treaty Act. Threats to golden eagles include loss of habitat and prey. 

Additional threats that are directly linked to human activities include illegal shootings, electrocution at power poles, lead poisoning from consuming parts of bullets in the entrails of deer carcasses discarded at the site of hunters’ kills, collisions with cars at sites where the birds are scavenging roadkill and collisions with the blades of a wind turbine. 

Across the western United States, hundreds of wind turbines have gone up in sage-brush flats that are part of golden eagles’ core habitat, and many more turbines are planned. In order to meet the permit requirements of the FWS, wind-energy companies must agree to mitigate their impact on the animals by offsetting the predicted number of golden eagles that will fly into their turbines each year. 

Currently, the only offset strategy approved by the FWS for wind-energy companies is to retrofit power poles to prevent golden eagles from becoming electrocuted. 

Adding empirical data

For the past five years, Lonsdorf and his colleagues have combined their expertise to develop a range of potential offset strategies for golden eagle fatalities. 

Their current paper — an updated model for golden eagle mortality due to vehicle collisions based on data from Wyoming — considered myriad factors such as the population density for golden eagles in the region, the number and size of deer roadkill carcasses expected and the traffic volume on the roads. The model also incorporated observational evidence of eagle-carcass roadside interactions obtained by motion-triggered cameras, data that was lacking in a previous model the researchers created. 

The addition of this empirical data allowed the researchers to make estimates for how long a golden eagle typically spends at a carcass, how the decay rate of the carcass affects the number of visits from eagles and the effects of seasonality on the scavenging behavior of the eagles. 

The model results suggest that carcass relocation is a viable golden eagle mitigation strategy that could save up to seven golden eagles annually in some Wyoming counties. On average, the model indicates that the prompt removal of four roadside carcasses would save at least one golden eagle. 

The researchers can make a user-friendly version of the prediction framework available to the FWS and wind-energy companies if the FWS decides to approve carcass removal as an eagle mortality offset strategy. 

“We’re taking basic information about golden eagle ecology in the Anthropocene and developing it into predictive frameworks for how to protect them,” Lonsdorf says. “As wind energy continues to grow, more mitigation strategies will likely be needed. Our goal is to provide scientific evidence for a portfolio of methods to help accomplish a zero-net loss of golden eagles from wind-energy facilities.” 

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Natural compound found in plants inhibits deadly fungi

A 3D illustration of the newly emerged species of fungus Candida auris, which is often drug-resistant and has a high mortality rate. (Dr_Microbe, Getty Images)

By Carol Clark

A new study finds that a natural compound found in many plants inhibits the growth of drug-resistant Candida fungi — including its most virulent species, Candida auris, an emerging global health threat. The journal ACS Infectious Diseases published the discovery led by scientists at Emory University. 

Laboratory-dish experiments showed that the natural compound, a water-soluble tannin known as PGG, blocks 90% of the growth in four different species of Candida fungi. The researchers also discovered how PGG inhibits the growth: It grabs up iron molecules, essentially starving the fungi of an essential nutrient. By starving the fungi rather than attacking it, the PGG mechanism does not promote the development of further drug resistance, unlike existing antifungal medications. 

Laboratory-dish experiments also showed minimal toxicity of PGG to human cells. 

“Drug-resistant fungal infections are a growing healthcare problem but there are few new antifungals in the drug-development pipeline,” says Cassandra Quave, senior author of the study and associate professor in Emory School of Medicine’s Department of Dermatology and the Center for the Study of Human Health. “Our findings open a new potential approach to deal with these infections, including those caused by deadly Candida auris.” 

C. auris is often multidrug-resistant and has a high mortality rate, leading the Centers for Disease Control and Prevention (CDC) to label it a serious global health threat. 

“It’s a really bad bug,” says Lewis Marquez, first author of the study and a graduate student in Emory’s molecular systems and pharmacology program. “Between 30 to 60% of the people who get infected with C. auris end up dying.” 

An emerging threat 

Candida is a yeast often found on the skin and in the digestive tract of healthy people. Some species, such as Candida albicans, occasionally grow out of control and cause mild infections in people. In more serious cases, Candida can invade deep into the body and cause infections in the bloodstream or organs such as the kidney, heart or brain. 

Immunocompromised people, including many hospital patients, are most at risk for invasive Candida infections, which are rapidly evolving drug resistance. 

In 2007, the new Candida species, C. auris, emerged in a hospital patient in Japan. Since then, C. auris has caused health care-associated outbreaks in more than a dozen countries around the world with more than 3,000 clinical cases reported in the United States alone. 

A ‘natural’ approach to drug discovery 

Quave is an ethnobotanist, studying how traditional people have used plants for medicine to search for promising new candidates for modern-day drugs. Her lab curates the Quave Natural Product Library, which contains 2,500 botanical and fungal natural products extracted from 750 species collected at sites around the world. 

“We’re not taking a random approach to identify potential new antimicrobials,” Quave says. “Focusing on plants used in traditional medicines allows us to hone in quickly on bioactive molecules.” 

Previously, the Quave lab had found that the berries of the Brazilian peppertree, a plant used by traditional healers in the Amazon for centuries to treat skin infections and some other ailments, contains a flavone-rich compound that disarms drug-resistant staph bacteria. Screens by the Quave lab had also found that the leaves of the Brazilian peppertree contain PGG, a compound that has shown antibacterial, anticancer and antiviral activities in previous research. 

A 2020 study by the Quave lab, for instance, found that PGG inhibited growth of Carbapenem-resistant Acinetobacter baumannii, a bacterium that infects humans and is categorized as one of five urgent threats by the CDC. 

The Brazilian peppertree, an invasive weed in Florida, is a member of the poison ivy family. “PGG has popped up repeatedly in our laboratory screens of plant compounds from members of this plant family,” Quave says. “It makes sense that these plants, which thrive in really wet environments, would contain molecules to fight a range of pathogens.” 

Experimental results 

The Quave lab decided to test whether PGG would show antifungal activity against Candida. Laboratory-dish experiments demonstrated that PGG blocked around 90% of the growth in 12 strains from four species of Candida: C. albicans, multidrug-resistant C. auris and two other multidrug-resistant non-albicans Candida species. 

PGG is a large molecule known for its iron-binding properties. The researchers tested the role of this characteristic in the antifungal activity. 

“Each PGG molecule can bind up to five iron molecules,” Marquez explains. “When we added more iron to a dish, beyond the sequestering capacity of the PGG molecules, the fungi once again grew normally.” 

Dish experiments also showed that PGG was well-tolerated by human kidney, liver and epithelial cells. “Iron in human cells is generally not free iron,” Marquez says. “It is usually bound to a protein or is sequestered inside enzymes.” 

A potential topical treatment 

Previous animal studies on PGG have found that the molecule is metabolized quickly and removed from the body. Instead of an internal therapy, the researchers are investigating its potential efficacy as a topical antifungal. 

“If a Candida infection breaks out on the skin of a patient where a catheter or other medical instrument is implanted, a topical antifungal might prevent the infection from spreading and entering into the body,” Marquez says. 

As a next step, the researchers will test PGG as a topical treatment for fungal skin infections in mice. 

Meanwhile, Quave and Marquez have applied for a provisional patent for the use of PGG for the mitigation of fungal infections. 

“These are still early days in the research, but another idea that we’re interested in pursuing is the potential use of PGG as a broad-spectrum microbial,” Quave says. “Many infections from acute injuries, such as battlefield wounds, tend to be polymicrobial so PGG could perhaps make a useful topical treatment in these cases.” 

Scientists from the University of Toronto are co-authors of the paper, including Yunjin Lee, Dustin Duncan, Luke Whitesell and Leah Cowen. Whitesell and Cowen are co-founders and shareholders in Bright Angel Therapeutics, a platform company for development of antifungal therapeutics, and Cowen is a science advisor for Kapoose Creek, a company that harnesses the therapeutic potential of fungi. 

The work was supported by grants from the National Institutes of Health, National Center for Complementary and Integrative Health; the Jones Center at Ichauway, the CIHR Frederick Banting and Charles Best Canada Graduate Scholarship and the Canadian Institutes of Health Research Foundation. 

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