The intestines provide a smorgasbord of nutrients for gut bacteria. Other than ingested food, glucocorticoids in bile are a significant substrate for microbes. Bacteria can convert these steroids into various metabolites that affect host systems.1 In a study published recently in Cell, researchers showed that some gut bacteria can produce sex hormones, including progestins.2 Researchers have previously shown that progestins regulate the menstrual cycle and pregnancy, and alter neuronal activity. This study marks the first report of bacterial production of progestins.
To isolate bacteria capable of converting glucocorticoids into sex hormones, researchers cultured human fecal samples onto agar plates.
Megan McCurry, Harvard Medical School Electron Microscopy Facility
“I’m constantly surprised by the scope of influence of gut microbes on the host, but maybe I shouldn’t be anymore,” said Gerard Clarke, a neurobiologist with an interest in the gut-brain axis at University College Cork.
For the better part of her education, Sloan Devlin trained to be a chemist. However, as a postdoctoral researcher in a lab that studies host-microbiome interactions, she saw an opportunity for someone with a chemistry background to elucidate the effects of bacterial molecules on the host. “There are chemists who are more drawn to physical sciences and material sciences and then there are chemists who are more drawn toward biology,” said Devlin. “I was always more drawn to biology.” Now, in her lab at Harvard Medical School, Devlin is trying to understand the mechanisms by which gut bacteria affect neurological function and behavior.
While scanning the literature looking for information on how gut bacteria metabolize glucocorticoids, the authors realized there was hardly any new data on the topic since the early 1980s—a 40-year gap in knowledge. Some researchers had reported that gut bacteria can convert these molecules into sex hormones. However, since these studies were conducted before the advent of modern genetics, there was no information about the bacterial strains, or the genes involved in the process. Back then, researchers needed between five to 50 milliliters of bile to extract a few microliters of steroids.3 Due to strict patient protection regulations, acquiring such large volumes from patients is no longer possible. So, Devlin had to develop a technique to extract and measure biomolecule levels from limited and precious quantities of bile.
For this, Devlin teamed up with Megan McCurry, a microbiologist at the biotechnology company Holobiome and coauthor of the study. McCurry spent five years developing a highly sensitive chromatography technique that requires 500-times less volume of bile than previous assays. Her technique also proved to be effective at separating and quantifying steroids, which are very potent and are thus present in miniscule quantities.
Using the improved technique, McCurry found that certain glucocorticoids are more abundant in human bile than others. The authors were most intrigued by a group of glucocorticoids that that could be converted into progestins, a class of sex hormones.
To determine if gut bacteria can perform this chemical reaction, the researchers collected feces from mice with and without gut bacteria and cultured the samples with the glucocorticoid. Feces serve as a proxy for the gut microbiome and its products. Devlin and her team observed progestins only in the feces of mice with a gut microbiome, suggesting that bacteria play a key role in producing the hormone.
Exposing Eggerthella lenta bacteria to hydrogen gas (in the yellow balloons) caused them to produce progestin from glucocorticoids.
Megan McCurry, Harvard Medical School Electron Microscopy Facility
Devlin wanted to isolate the bacterial species responsible for converting glucocorticoids into progestins. Based on previous research that showed that the species Eggerthella lenta could perform this reaction, Devlin and her team tested various strains of the bacteria. However, they never observed progestin production. Therefore, Devlin and her team decided to isolate the species from human feces. They cultured human feces with an amino acid that stimulates the growth of E. lenta and its microbial relatives and looked for bacteria that produce progestin. Although the human gut microbiome contains 300 to 500 different species of bacteria, only one, Gordonibacter pamelaeae, did the job, albeit poorly.4 However, when they grew G. pamelaeae along with the gut commensal E. coli Nissle 1917 (EcN) this significantly improved progestin production. Twelve other bacterial strains from the same family—Eggerthellaceae—gained the ability to produce progestins when co-cultured with EcN.
It was clear that EcN was crucial for the chemical reaction, so Devlin hypothesized that it could be creating conducive conditions for progestin production by one of three ways: reducing the redox potential of the medium needed for the reaction, physically interacting with the other bacteria for cooperative metabolism, or releasing extracellular agents that boost bacterial activities.
They tested each of these scenarios using E. lenta, a relative of G. pamelaeae that scientists can genetically modify. After disproving the first two theories, the team had a eureka moment. While testing the impact of extracellular agents, they extracted the supernatant of EcN cultures and filtered them to remove dissolved gases. However, for some cultures, they skipped this step. When they added the supernatant to the E. lenta cultures, the authors observed progestin production only when the bacteria had access to dissolved gases. Since hydrogen gas is predominant in the gut and is produced by EcN, Devlin suspected it could be the key to bacterial conversion of glucocorticoids to progestins. To test this, they added hydrogen gas to E. lenta and observed a similar amount of progestin production.
“All of us pass gas, right?” said Devlin. “But our discovery that this hydrogen production is actually inducing bacterial metabolism of steroids was to me the most surprising thing that came out of the work.”
The researchers captured an image of an Eggerthella lenta and E. coli Nissle co-culture using electron microscopy.
Megan McCurry, Harvard Medical School Electron Microscopy Facility
Next, using comparative genomics analysis on the genome of bacterial strains that produce progestins and those that do not, Devlin and her team identified a four gene cluster involved in the reaction. Expressing this cluster in a bacterial species that cannot convert glucocorticoids to progestins enabled it to do so.
“This is the kind of work that is needed right now in the field,” said Clarke. “We have a lot of observations for microbes doing specific things to impact the host, but we’re missing the resolution that’s needed to intervene and tune things to our favor.”
Other groups have shown that levels of certain progestin-producing glucocorticoid are five to 10 times higher in pregnant people.3 Using human feces from pregnant and non-pregnant individuals, Devlin and her team found that the increase in glucocorticoid concentration translated to exaggerated progestin levels that were two times higher in pregnant people.3 An enrichment of G. pamelaeae, E. lenta, and the gene cluster in these samples strongly suggested that the microbiome contributes to the high levels of progestins.
To further explore how the microbiome churns out higher levels of progestins during pregnancy, the team transplanted feces from pregnant mice to non-pregnant female mice without a microbiome and detected high levels of progestins in the feces of the receivers. Transplanting these mice with only E. lenta and EcN produced the same results.
“One of the things about our work is that it raises more questions than it answers,” Devlin said. She is interested in looking at longitudinal samples throughout pregnancy to uncover the relationship between the gut microbiome and the rates of postpartum depression. Unfortunately, Devlin noted, a biobank of fecal samples is hard to come by. “My message to clinicians would be: Consider collecting feces from your patients,” Devlin said.
