Nov. 6, 2019: New research from the EPI and UF’s College of Public Health & Health Professions found that exposing the sexually-transmitted bacterial pathogen Chlamydia to fosmidomycin — an antibiotic which is usually lethal to bacteria — causes Chlamydia to enter a protective bunker-like “persistent” state. The findings could bolster future efforts to intentionally disrupt the molecular changes that induce chlamydial persistence, leading to the prevention of chronic chlamydial infections.
New research from the Emerging Pathogens Institute and UF’s College of Public Health & Health Professions reveals that using the antibiotic fosmidomycin against Chlamydia trachomatis causes the bacteria to enter a protective, bunker-like “persistent” state which helps the microorganism survive harsh conditions. When conditions improve, Chlamydia leave the persistent state and continue to replicate.
Understanding the gene-level changes that take place for Chlamydia to enter the unique state known as persistence could help researchers to eventually develop strategies to block these changes from occurring, making the organism more vulnerable to antibiotics and circumventing chronic chlamydial infections. Researchers believe that Chlamydia’s ability to persist for long periods of time in a human host might contribute to the worst outcomes of infection: infertility and ectopic pregnancy.
Chlamydia trachomatis is the most common bacterial sexually-transmitted disease in the U.S., and it causes about 130 million new genital tract infections globally each year. Between 50 and 90 percent of these cases do not produce symptoms however, and it is thought that these asymptomatic infections may be attributed to people carrying C. trachomatis in a persistent state.
Research led by Jessica Slade, Ph.D., a postdoctoral associate in the lab of EPI/PHHP researcher Anthony Maurelli, Ph.D., originally sought to test whether fosmidomycin might have antimicrobial activity against C. trachomatis. Slade was inspired to test this particular drug because fosmidomycin is known to kill specific types of bacteria and parasites that invade host cells; and Chlamydia bacteria use cellular invasion as a key reproductive strategy: they enter epithelial cells where they can then safely divide.
“I thought that fosmidomycin would prove lethal against Chlamydia,” Slade said. “But it turned out to be a very different story.” The study recently published in PLOS-Pathogens.
In other types of intracellular microbes, fosmidomycin inhibits a metabolic pathway known as isoprenoid synthesis, which then leads to cellular death. But Slade found that when HeLa cells infected with C. trachomatis were exposed to fosmidomycin, isoprenoid synthesis inhibition did not prove lethal. Instead, the drug disrupted the ability of Chlamydia to perform cell wall synthesis and cellular division, likely due to a shortage of a key isoprenoid called bactoprenol, and this disruption led to the bacteria seeking the refuge of persistence.
“Our study also showed that isoprenoids, and most likely bactoprenol in particular, are necessary for Chlamydia to divide,” Slade says, noting that this relationship was previously not identified.
Structured illumination microscopy shows C. trachomatis-infected HeLa cells that were exposed to nothing (top row), exposed to fosmidomycin (FSM, second row) and exposed to ampicilin, a β-lactam antibiotic, (AMP, bottom row). Red coloration depicts the major outer membrane protein, while green depicts peptidoglycan, which is necessary for cellular division.
Understanding the specific steps that induce persistence means that researchers can eventually develop disruptive mechanisms, which might be a key step to preventing many of the adverse outcomes resulting from chronic Chlamydia infection.
“Chlamydia have this unique growth cycle and intracellular lifestyle where they are protected from the osmotic stress that occurs from exposure to antibiotics that can inhibit cell wall synthesis, like penicillins and fosmidomycin,” Slade says. “In other bacteria, that are not osmotically protected, exposure to these drugs ultimately causes lysis of the bacteria. But Chlamydia can survive these insults by entering the persistent state inside a host cell and have refuge from the osmotic stressors that are the downfall of other kinds of bacteria that are exposed to these drugs.”
Current drug treatments, including azithromycin and doxycycline, work well when they reach C. trachomatis cells that are undergoing division. However, most chlamydial infections are asymptomatic, meaning that a patient may be unaware that they have the infection and can transmit it to others, and may not even seek treatment.
Drugs like penicillin are among the most commonly prescribed antibiotics for other infections, such as urinary tract infections. Patients who are unaware that they have a chlamydial infection are then exposed to antibiotics that can potentially cause Chlamydia to enter its persistent state even though the antibiotic effectively treats UTIs. In this scenario, chlamydial infection could last longer and lead to more harmful outcomes.
The response of Chlamydia to fosmidomycin fits a larger pattern of its cunning adaptation and survival, as these bacteria will also seek the refuge of their persistent state when exposed to penicillins, iron scarcity, co-infection with certain viruses, and interferon gamma (a cytokine produced by certain human immune system cells).
“Isoprenoids make up a large group of diverse molecules that participate in a variety of critical functions in cells and we do not yet know all of the ways these molecules contribute to persistence,” Slade says. “Future studies will seek to understand the genetic and molecular mechanisms of entry into fosmidomycin-induced persistence. If we can understand the genetic changes Chlamydia makes in order to enter and leave persistence, then we will be on the right track toward preventing Chlamydia from utilizing persistence as a survival strategy to evade drug treatment.”
When Chlamydia encounter conditions that are stressful for them, they stop dividing and instead switch to a safe persistent state. Yellow and black arrows point to Chlamydia inclusion bodies within infected HeLa cells that were not exposed to antibiotics; and the AB labels pinpoint "aberrant bodies" which formed after exposure to fosmidomycin. The aberrant bodies indicate a stressful environment, and the bacteria enlarge visibly when they enter this persistent state.
Written by DeLene Beeland; Top photo: microscopy image courtesy of Jessica Slade, Ph.D., showing HeLa cells infected with Chlamydia trachomatis that then developed inclusion bodies after exposure to fosmidomycin.
Read Slade and Maurelli's paper, “Fosmidomycin, an inhibitor of isoprenoid synthesis, induces persistence in Chlamydia by inhibiting peptidoglycan assembly” which was published in PLoS Pathogens on October 17, 2019.
Learn more about Chlamydia cell biology and pathogenesis in this Nature Reviews Microbiology paper.