by Amanda Siegfried
UPON ACCEPTING HIS Nobel Prize in 1945, the scientist who discovered penicillin warned of the dangers of infectious bacteria becoming resistant to antibiotics. By the late 1940s, the penicillin-resistant bacterium Staphylococcus aureus, which causes pneumonia, sepsis and skin infections, had become a global pandemic.
Dr. Alexander Fleming’s dire predictions have been amplified in modern times: Frontline antibiotics have become nearly obsolete as disease-causing bacteria quickly evolve and adapt for survival. It’s an arms race the human race might lose, unless researchers can develop strategies to stay more than one step ahead of marauding microbes.
Microbiologists at The University of Texas at Dallas are in the fray, discovering the tricks bacteria play to evade antibiotics and developing new approaches to fight infectious diseases.
BACTERIA ARE SINGLE-CELLED organisms that live nearly everywhere on the planet—in the air, water and earth. They break down soil and provide nutrients for plants to grow. They live on the human body, causing body odor, and inside it, digesting food. There are as many bacterial cells as human cells in and on a human body, resulting in a healthy symbiosis.
In the human gastrointestinal (GI) tract, a conglomeration of helpful bacteria, called the gut microbiome, is thought to be unique among individuals.
Wherever they live, bacterial species battle one another for food, and they have evolved defenses to keep their rivals at bay. Scientists have discovered many of the chemical weapons bacteria produce to kill their competitors, and the pharmaceutical industry has subsequently exploited those discoveries to produce lifesaving drugs — antibiotics — that kill bacteria that cause human diseases.
But nearly every antibiotic in use today is based on discoveries made more than 30 years ago, according to the Pew Charitable Trusts’ Antibiotic Resistance Project. Bacteria constantly mutate, and some have developed ways to survive not just one antibiotic, but many, earning the moniker “superbug” for their multi-drug resistance. Such antibiotic resistance is accelerated in part by the overuse and inappropriate use of the drugs, for example taking an antibiotic for an infection caused by a virus. (Antibiotics do not kill viruses.)
Research Goes Viral
HOW EXACTLY DO bacteria develop antibiotic resistance? Dr. Kelli Palmer, an associate professor of biological sciences who joined the School of Natural Sciences and Mathematics faculty in 2012, is investigating the underlying mechanisms by which bacteria acquire antibiotic resistance genes from one another. Her work is funded primarily by the National Institutes of Health.
For example, in a study published in the Journal of Bacteriology, Palmer, her students and her colleagues shed light on a gene-swapping process called conjugation, which, she tells her students, is like bacterial sex.
“Bacteria of different species can exchange antibiotic resistance genes through conjugation, which requires them to meet up and come into physical contact in close quarters, such as in human wounds or on hospital surfaces,” said Palmer, Fellow, Cecil H. and Ida Green Chair in Systems Biology Science.
Dr. Kelli Palmer and members of her UT Dallas laboratory, including research scientist Dr. Yahan Wei, are investigating how bacteria develop resistance to antibiotics.
Research in Palmer’s lab — which she is quick to note is driven and carried out by talented students — also examines how viruses might be used to combat antibiotic resistance.
A virus that only infects and kills bacteria is called a bacteriophage, or phage for short. Phages don’t infect human, animal or plant cells.
Just as bacteria can acquire resistance to antibiotics, they also can quickly evolve resistance to phage infection. In a recent study, led by former biology student Khang Ho BS’18, who is now a graduate student at UT Austin, Palmer’s team observed this happening in a population of Enterococcus faecalis bacterial cells. Like some other species of bacteria normally found in the gut, E. faecalis can cause dangerous infections if it spreads to other areas of the body, like the bloodstream. Some E. faecalisstrains have become highly antibiotic resistant.
“The only thing we did was introduce the phage and these bacteria to each other,” Palmer said. “Then, at low frequency, bacterial mutants arose that were resistant to that phage. It took one day.”
The mutated bacterial cells altered their surfaces to block the docking site where phage would normally attack. But that adaptation came at a price.
“When this strain became resistant to the phage, meaning it could no longer be killed by the phage, it simultaneously became more susceptible to antibiotics,” Palmer said. “That’s fantastic. What this means is, you might be able to develop a combination therapy using phages to kill off disease-causing bacteria and, if any happen to evolve resistance, come in with an antibiotic.”
Waste Not, Want Not
IN ADDITION TO phage therapy, another wave of the future might be fecal microbiota transplantation, Palmer said.
Admittedly, a fecal transplant sounds disgusting, but it’s actually becoming a more mainstream approach to treating certain hard-to-cure bacterial infections, especially infection with Clostridium difficile. When a patient takes a broad-spectrum antibiotic, this opportunistic bacterium often survives in spore form in the gut, emerging after most of the other helpful bacteria have been killed off.
Without any competition, C. diff thrives and can result in chronic diarrhea. The Centers for Disease Control and Prevention considers it “a major health threat” that infects 500,000 people every year in the U.S. and kills 15,000. For patients who experience recurring C. diff infections, fecal microbiota transplants have been shown in published research to be 80% to 90% effective, although the medical procedure is still considered investigational. It essentially restores a patient’s healthy gut microbiome by infusing the GI tract with feces from a healthy donor.
“Fecal transplantation is cheap. It’s just poo,” Palmer said.
“I think that understanding how organisms naturally compete with each other — which they do — in their native environment, and using that to our advantage to out-compete drug-resistant strains is a viable option,” she said.
Case Study: Urinary Tract Infections
URINARY TRACT INFECTIONS (UTIs) are irritating and painful, sometimes debilitatingly so. The majority of UTIs are caused by Escherichia coli, which normally lives in human intestines but sometimes gets into the urinary tract, where it is not welcome.
Roughly 10% of women in the U.S. experience a UTI each year. For most — and these infections predominantly occur in women — they are a temporary condition that can be effectively treated with available antibiotics.
But for some postmenopausal women, UTIs recur so frequently that they become a chronic condition, requiring daily doses of increasingly powerful antibiotics as the infection-causing bacteria gradually become resistant to each new drug.
This microscope image shows bacteria, in green, within the bladder wall tissue of a patient with recurring urinary tract infections.
“For older women, these infections can go on for tens of years,” said Dr. Nicole De Nisco, assistant professor of biological sciences at UT Dallas. “Eventually, a patient’s last resort might be removing the bladder.”
Most of the epidemiological research on UTIs has been done with women in their 20s and 30s, a much earlier age range than the typical onset of menopause. Likewise, the disease has been primarily investigated in juvenile animal models, so knowledge of it in postmenopausal women is severely limited, De Nisco said.
“One of the reasons urinary tract infections have been overlooked is because they affect women, an understudied group in general when it comes to disease, and older women in particular, who are even more understudied,” De Nisco said.
“UTIs are not going to kill you, but the effect on a patient’s life can be profound,” she said. “Many women will withdraw from society and activities, because if you have to use an external bag as your bladder, your quality of life is greatly affected.
“I think it’s a crime that this is the best we can do.”
Dr. Nicole De Nisco conducts research aimed at understanding the basis for recurring urinary tract infections in postmenopausal women. In her lab, students monitor the growth of various bacteria on cell-culture dishes.
PRIOR TO JOINING the UT Dallas faculty in 2018, De Nisco was a Howard Hughes Medical Institute postdoctoral fellow at UT Southwestern Medical Center. A graduate of the Massachusetts Institute of Technology, De Nisco has an ongoing clinical collaboration to study recurring UTIs with Dr. Philippe Zimmern, professor of urology and the Felecia and John Cain Chair in Women’s Health at UT Southwestern.
De Nisco believes that long-term antibiotic therapy to manage recurring UTIs is not only ineffective at permanently clearing the infection, but might exacerbate the problem in postmenopausal women.
“It’s a little controversial. You don’t want to tell a patient, ‘I’m not going to give you antibiotics,’ but it’s becoming more widely accepted that long-term prophylactic treatment with antibiotics, whether you have an infection or not, is not the best way to handle recurring infections,’ she said. “While low-dose antibiotics are generally tolerated by patients, microbiologists know that if you grow bacteria in such subinhibitory concentrations of a drug, they will quickly develop resistance to that drug. This disconnect between scientists and clinicians is a gap my colleagues and I hope to bridge.”
De Nisco’s goal is to identify targets for adjunct, or combination, therapies that more effectively halt UTI recurrence. Her approach is to examine both the behavior of the pathogenic bacteria as well as the host tissue they infect. Host inflammation seems to be a focal point.
She explained that infectious bacteria usually significantly change the host environment. In the case of UTI, invading bacteria like E. coli trigger production of a cyclooxygenase-II enzyme (COX-2) in host cells. This enzyme produces a chemical called prostaglandin E2 (PGE2), which attracts immune cells to the area, resulting in inflammation. This product of COX-2 can be easily detected in urine.
If not resolved, the inflammation can cause permanent damage to the host tissue.
“We’re investigating whether COX-2 levels are elevated in the tissue samples from our patient population. We also can see if the COX-2 product is elevated in their urine,” De Nisco said. “Preliminary results from these patients show that if you have a history of recurring UTIs, levels of PGE2 in your urine are higher than if you don’t have a history.”
These findings suggest that COX-2 inhibitors — a type of nonsteroidal anti-inflammatory drug (such as Celebrex) that blocks the activity of COX-2 — might be an effective therapy to reduce inflammation, giving the tissue a chance to heal and making it less hospitable to E. coli bacteria.
“This is one inflammatory pathway that we would like to target with a clinical trial,” De Nisco said. “If we could attack the cycle of infection at two points — both with antibiotics and selective antiinflammatories — maybe we’ll have a better chance of actually breaking the cycle of recurrence and curing the disease instead of just managing it.”
Another important target for recurring UTIs might be the microbiome.
“We’re also beginning to understand that there are bacteria that normally reside in the urinary tract — a urinary microbiome. An imbalance in this microbiome, often brought on by antibiotic use, likely has a role in recurrence,” De Nisco said.
De Nisco and Palmer are working together to characterize precisely the urinary microbiome in postmenopausal women. They use an advanced technique called whole genome metagenomic sequencing to identify bacterial species in urine by their DNA. The goal is to determine whether there are particular microbiome profiles that are either protective or predisposing to recurrent UTIs.
“By sequencing the entire genomes present, we can begin to get a better understanding of how different microbial species might be working together or against each other to create a healthy versus unhealthy microbiome,” De Nisco said.
“For the science that I want to do and the collaborations I’m involved with, UT Dallas is the perfect place to be,” she said.
From Curious Kids to Serious Scientists
A SELF-DESCRIBED “Girl Scout at heart,” microbiologist Dr. Nicole De Nisco has always loved solving problems.
“I was one of those kids who never had to be told to do their homework,” she said. “I have always enjoyed learning, and, as a scientist, you never stop learning and finding new problems
“It sounds corny, but I want to leave the world better than I found it. This is a core value instilled in me by my parents. Because of this, I strive to always keep a strong connection between my research and the end goal of improving human health.”
WHEN MICROBIOLOGIST Dr. Kelli Palmer entered college, she defaulted to pre-med because “that’s what smart kids who are interested in science were told to do,” she said. The only scientist in her family, Palmer started her research career washing dishes in a microbiology lab.
“I loved the lab so much that I transitioned to research and never left,” she said. “As an undergraduate, I volunteered to do research in a chemistry lab, but I was just analyzing data, and I hated it. The professor could tell and suggested that I needed to be in a biology lab. She was right; I enjoy hands-on research, growing things. I just like the messiness of biology.”
Nicole De Nisco
Dylan McNutt, a senior in neuroscience, and Nhi Nguyen, a senior in biochemistry, learned the techniques scientists use to discover new antibiotics.
Antibiotic Discovery in the Classroom
A REVAMPED MICROBIOLOGY lab course at UT Dallas is teaching undergraduates the investigative skills that might just lead to the next new antibiotic.
The learning process starts with a handful of dirt.
“In the first week, I ask each student to bring a soil sample to class,” said Dr. Iti Mehta, a senior lecturer in biological sciences who leads the course. “From those samples and others I provide, they isolate different bacterial species and see whether they are producing any chemicals that act as antibiotic agents.”
The class, taught for the first time in spring 2019, differs from standard microbiology courses because it takes a project-based approach to teaching microbiology research techniques. The concept was inspired by curricula developed by Dr. Jo Handelsman, director of the Wisconsin Institute for Discovery at the University of Wisconsin-Madison. Her program, called Tiny Earth, encourages college students worldwide to conduct research and essentially to crowdsource antibiotic discovery.
“We took the opportunity to reimagine this class and transform it from a traditional cookbook-style lab class into a true research course,” said Dr. Stephen Spiro, UT Dallas associate provost and the C.L. and Amelia A. Lundell Distinguished Professor of Life Sciences. “In its new format, the class is an open-ended investigative project in which students participate in original research and hopefully make new discoveries. They are also learning important skills in teamwork, critical thinking and communication.”
The students, who work in pairs, begin by isolating bacteria from soil and identifying species using DNA sequencing technology, an advanced technique not typically taught in undergraduate labs. Next, they test whether their bacteria produce any chemicals that can kill clinically relevant bacteria. They also test those chemicals to see whether they have any adverse effects on plant cells.
To increase the chances of discovering novel compounds, a long-term goal is to extend the approach to other environments in addition to soil.
“Students compare what they find to a database of known antimicrobial agents,” said Mehta, who earned her PhD in molecular and cell biology from UT Dallas in 2018. “Even if they don’t find anything novel, they are learning the techniques and hopefully getting excited about the discovery process and possible careers in research.”