Got gut issues? Research at Oak Ridge National Laboratory may lead to help in the future

The human genome has around 23,000 genes, only about 3,000 more genes than much simpler living systems have. The 1 million or so additional genes that bacteria and other resident microorganisms bring to the human system (our mouth, esophagus, stomach, small intestine and large intestine, including the colon) greatly expand our functional activities. For example, they likely give us the ability to eat salad and other plant polysaccharides without suffering terrible indigestion.

Those were some of the facts presented in a recent talk on the human gut microbiome by Bob Hettich, corporate fellow and leader of the Bioanalytical Mass Spectrometry Group in the Biosciences Division at Oak Ridge National Laboratory. His research is supported by the National Institutes of Health.

He spoke to Friends of ORNL about his pioneering work in using biological mass spectrometry to identify and determine the relative abundances of the gut’s human and microbial proteins (including enzymes), which are products of the hundreds of thousands of genes in the microbiome.

In speaking about some of the samples he subjects to liquid chromatography and electrospray mass spectrometry for analyzing large molecules, such as proteins and peptides, he joked, “I feel like a new father. I’ve got freezers full of baby poop.”

Hettich predicted that within five years, thanks to the work he and others are doing, “you’ll likely give a fecal sample to your doctor so that your microbiome can be sequenced and available for the doctor’s medical reference.”

In other words, your doctor will tell you the types and relative populations of the bacterial proteins you have in your gut. This information on your microbiome, whether it is rich or shallow in a large variety of beneficial bacteria, may be a signature of your health or of a disease, such as cancer, diabetes or autoimmune diseases such as Crohn’s or Parkinson’s.

“Healthy people have an incredibly rich diversity and abundance of bacteria in their gut, predominantly in the colon,” he said. “But in people who have diseases such as diabetes or Crohn’s, their bacteria diversity collapses and the rich complexity they once had becomes very shallow.” Also, he added, in the case of inflammable bowel diseases such as Crohn’s and ulcerative colitis, the mucous layer becomes thin, enhancing inflammation of the colon.

Increased knowledge about the bacterial composition of a patient’s microbiome can also guide a doctor in deciding which drugs will be effective and which will not be in treating the patient’s disorder, he said

Hettich is one of the pioneers of metaproteomics, which is the use of biological mass spectrometry to identify and quantify all the proteins in a complex biological sample, such as a community of microorganisms that make up the human gut’s microbiome. The information is used to understand the functions of the microorganisms in the community.

The reason he's passionate about his research

He is passionate about determining the relationship of the microbiome to inflammatory bowel diseases because one of his two daughters began suffering from Crohn’s when she was only eight years old.

“This is a bit unusual in that most people get this disease in their late teens to early twenties,” he said. “She has had to undergo multiple surgeries and other treatments.”

Hettich said his daughter somehow contracted a bacterial insult during birth, which resulted in a dangerous blood infection.

“She was really in a very critical condition and almost died,” he said. The doctors gave her one antibiotic that didn’t work, so they administered a more powerful one that stabilized her and saved her life. But she was diagnosed with Crohn’s disease eight years later. “I think we messed up her microbiome at a very critical stage of development."

Working with Michael Morowitz, a pediatric surgeon at the University of Pittsburgh, Hettich has studied the evolution of the microbiome in babies. He cited evidence that overuse of antibiotics in babies and young children (for example, with frequent ear infections) can have undesirable effects on their microbiome and health later in life.

In answer to a question, Hettich noted that residents of the Southeast rank among the worst in the nation in receiving too many antibiotics and eating too much sugar and fat, all of which reduce the rich complexity of the microbiome that help make people healthy.

“Such harmful effects on the microbiome can manifest in all kinds of unhealthy conditions, including obesity,” he said.

Hettich spoke about his collaboration with Morowitz, who specializes in pre-term human infants and medical complications they endure. Somewhat surprisingly, most babies are born with sterile, microbe-free guts even though the mothers have tremendous amounts of bacteria in their wombs.

“From birth to about one week, the baby is hit with an onslaught of bacteria it has never seen before,” Hettich said. “The bacteria come from various environmental sources, such as the nurses and mother touching them, the blankets, the pacifiers, the feeding sources.

“The baby’s gut is initially colonized by bacterial anaerobes that thrive without oxygen. They get outcompeted by other bacteria and the baby’s gut microbiome rapidly evolves into richly populated communities that can become stable for the rest of the person’s life.”

Hettich noted that the microbiome of a baby born by a Caesarean section is different from that of an infant who enters the world via a vaginal birth.

“If a baby is born by a vaginal birth, the microbiome looks like the mother’s fecal material,” he said. “If it’s born by Caesarean section, its microbiome looks like the mother’s skin. Whether the baby is breastfed or formula fed can affect the microbiome composition.

“What’s frightening is that some preterm babies can contract a horrible disease called necrotizing enterocolitis, or NEC. The gut becomes perforated, usually causing the infant to die.”

Necrotizing enterocolitis is a serious gastrointestinal condition that primarily affects premature or very low birth weight infants. It can lead to the death (necrosis) of tissue in the intestines.

Research into preterm infants, antibiotics

In a recent study, Hettich and postdoctoral researcher Samantha L. Peters used their mass spectrometry-based metaproteomics approach to examine the developing microbiomes of pre-term infants and the effects of antibiotics. For this study, fecal samples were collected from 16 preterm infants, half of them male and one-half female, over the first three months after birth. Most of the babies were delivered by C-section and were small enough for an adult to easily hold in one hand.

“We studied the microbiomes of 10 healthy babies and six babies that had NEC,” Hettich said. “Every one of them were given antibiotics shortly after delivery, undoubtedly as a ‘cautionary preventative measure.’ Why would you give antibiotics to a healthy baby that is developing a microbiome? Because that’s the accepted medical protocol for preterm infants.”

One baby who was studied contracted NEC twice. The infant received antibiotics and recovered, then got NEC again, received antibiotics and recovered. Hettich’s team tracked the microbiome’s changes during these episodes through analyses of the baby’s fecal samples and observed a complicated transition of the bacterial members and their activities.

In the same hospital, identical twin babies from the same mother were found to have completely different bacterial compositions in their microbiomes, revealing that early-stage microbial colonization is really “the Wild West” even in normal circumstances. One of the twins developed an infection, received antibiotics and survived; the other twin infant developed necrotizing enterocolitis and died on Day 31.

One of the ORNL researchers’ questions was, “Do specific proteins in the baby’s gut influence the dynamics of the gut’s microbiome establishment?” Their studies showed that during the early onslaught of bacteria into the baby’s gut, the human inflammation proteins are elevated to confront the bacterial proteins because they are sensed to be a threat. However, Hettich said, “We found that eventually the human and bacterial proteins settle down into a symbiotic relationship for healthy infants.”

Some of their findings were published in an article in a 2022 issue of Frontiers in Microbiology.

Hettich’s team also found that the human immune system’s white blood cells (macrophages) can use one type of copper to selectively kill certain bacteria and thus help customize the developing microbiome. They found out that most of the surviving bacteria possess the appropriate enzymes to detoxify the copper and thus thrive and expand in abundance.

“This might be one method of internal control of pathogenic bacteria, even in the developing pre-term human infant,” Hettich said

In an answer to a question about the microbiome and inflammation, Hettich stated that most people (maybe about 75%) have Clostridium difficile bacteria in their gut, a microorganism that can cause diarrhea, colitis (inflammation of the colon’s lining) and even death. But fortunately, he added, most of us have in our gut several types of beneficial Bacteroides species, which Hettich calls “the 500-pound gorillas that dominate and thus keep C. difficile at bay.”

Some people in Hettich’s audience may have found it unsettling when he stated that our bodies are estimated to have three to 10 times more microbial cells than human cells. But, by the end of his talk, it was clear that many microorganisms in our bodies are heroic residents that digest our food, protect us against infection, and keep us healthy. To see a video of Hettich’s presentation, visit www.fornl.org and click on “Past Talks” at the top left of the home page.