Why the Placental Microbiome Should Be a Cautionary Tale
A new study suggests that evidence for microbes found on placentas was the result of lab contamination.
For decades, scientists believed that babies encounter microbes for the very first time when they are born. Both a healthy womb and the placenta that nourishes the growing fetus, they said, are sterile. If bacteria do sneak in, they’re intruders and bad news for the fetus. But in 2014, Kjersti Aagaard from Baylor College of Medicine challenged that dogma. In placental tissue samples from 320 women, she found DNA from many kinds of bacteria, which made up a “unique placental microbiome,” she argued. Aagaard suggested that this microbiome is part of natural pregnancy and might seed a fetus’s body with microbes in utero.
“The placenta is not teeming with bacteria, but we can find them,” she told The New York Times, “and we can find them without looking too hard.”
More placental-microbiome studies have since been published. Aagaard’s team and others have looked at how this community of microbes varies with body weight, how it affects the placenta itself, whether it’s linked to diabetes or preterm birth, and whether it’s a “missing link” between gum disease and pregnancy problems.
But other microbiologists were dubious from the start. Many species reported in these studies are easy to grow in labs, they noted, so why had microbiologists largely failed to culture microbes from placentas for decades? And how could fetuses control such microbes when their immune systems are undeveloped? While writing a book about microbiomes, I asked many researchers if there were any discoveries to be wary of, and the most common answer was the placental microbiome.
Skeptics argued that the bacterial DNA most likely came from microbes living in the wider world, which had contaminated the equipment or the samples used in those earlier studies. Four studies that specifically tried to account for such contaminants found no evidence of a placental microbiome. Maybe it’s not a missing link at all. Maybe it’s just plain missing.
Now, a team at the University of Cambridge has weighed in with one of the most comprehensive studies to date, involving placental tissue from more than 500 women. The researchers found no evidence of a consistent community of microbes that live in healthy placentas. The few signs of bacterial DNA they detected came either from contaminants or from harmful microbes that only rarely infect placentas. “We spent a very long time thinking about how to remove and identify every source of contamination,” says Julian Parkhill, one of the study’s leaders. “When we did that, we realized there was nothing left.”
“This study, in my view, settles the matter,” says Marie-Claire Arrieta from the University of Calgary, who wasn’t involved. It’s a reminder that DNA data “can be easily misinterpreted,” she adds, and that “misinterpretations can become quite widespread.”
Marloes Dekker from the University of Queensland, who published a small placental-microbiome study in 2017, says the Cambridge study is bigger, more extensive, and did a better job of accounting for contamination. “It could certainly change the interpretation of our results,” she says. “I am not convinced of the existence of a true placental microbiota.”
Aagaard, meanwhile, stands by her work. She argues that the Cambridge researchers used techniques that would be unlikely to detect actual placental microbes, and that they filtered what they did find too strictly. She also points to other recent studies where researchers also found bacterial DNA in placental tissue, or managed to culture living microbes. Parkhill and his colleagues “are not recognizing, or are naive to, other evidence for colonization,” she says. “We can’t just brush this off and say this is sloppy science.”
Parkhill argues that Aagaard and others are underestimating the issue of contamination—a problem that extends well beyond the placenta. When scientists want to find microbes in a tissue sample, they use a set of chemicals to pull out any DNA, amplify it, and prepare it for sequencing. But these “extraction kits” almost always have traces of microbial DNA in them, too! Even if you use them on a tube of pure water, you’ll find bacterial DNA. That doesn’t matter when looking at organs like the gut, whose extensive communities of microbes overwhelm the faint signals from contaminants. But it very much matters when looking at places where microbes are rare or absent—like the placenta. There’s a huge risk that whatever you find comes from kits, not tissues.
This problem, jokingly known as the “kit-ome,” has led to many spurious discoveries. Bacteria that thrive on sunlight, or that can’t grow at body temperature, have been identified in the human brain. Bradyrhizobium, which lives on plant roots, has been found in many body parts. Another plant-associated species that’s incapable of infecting human cells was identified in breast tissue (of Irish women, but not Canadians). These examples have been compared to finding “blue whales in the Himalayas or African elephants in Antarctica.” They’re becoming more common, because the newfound popularity of the microbiome field, combined with the growing ease of sequencing techniques, attracts researchers who are unaware of “how ubiquitous contamination is,” Parkhill says. “A lot of people will just go and grab a technique off the shelf without realizing the subtleties and pitfalls.”
His team went out of the way to address these issues. When analyzing their 537 placental samples, the researchers also used extraction kits on blank samples to see which contaminants were there. They used two different kits on several samples and sequenced the extracted DNA with two different techniques, cross-referencing all the results. They even spiked their samples with a minuscule amount of the bacterium Salmonella bongori to check that they could detect rare microbes that were actually present.
They also looked for odd patterns. For example, they found Bradyrhizobium in some batches of samples that had been processed together but not in others—a sure sign that it came from the kits, not the placentas. (It showed up in the blank samples too.) Two other bacterial species matched strains that had been analyzed in a completely different study using the same sequencing machines; their DNA had almost certainly stowed away on the machines, and got into the placental samples. The team also detected DNA from vaginal bacteria, but rarely, in tiny amounts, and more often from vaginally delivered placentas than those delivered through C-sections. It’s unlikely that such microbes are regularly colonizing the placenta; either they contaminated the tissues during labor, or they represent rare infections.
Ultimately, the team only found convincing evidence for one species in the placental samples, and even then in just 5 percent of the samples, and at very low levels. It’s called Streptococcus agalactiae (pronounced ay-guh-lack-tee-ay)—a potential pathogen that can pass from mother to infant during birth and cause neonatal sepsis. “This doesn’t conflict with the dogma that the womb is microbe-free in healthy pregnancies, because this bacterium is associated with disease,” writes Nicola Segata from the University of Trento in a related commentary. “This finding provides strong evidence that there is no functional microbiota in the placenta.”
But Indira Mysorekar from Washington University in St. Louis notes that the team only took samples from a region of fingerlike structures called the terminal villi, which transfer nutrients from the mother’s bloodstream into the fetus’s. “That area is not expected to have the bacteria, and there really hasn’t been much controversy about whether there is a microbiome there,” Mysorekar says. When other studies found placental microbes, they looked at different parts of the organ.
Aagaard adds that the Cambridge team used techniques that would minimize the odds of finding resident microbes, from washing samples in saline to using sequencing protocols that are unlikely to find bacteria in low abundances. She also says that the team too readily billed microbes as vaginal contaminants, even when they were seen in placentas that had been delivered through C-sections. “They’ve subtracted out stuff that in my mind doesn’t make a lot of sense to subtract out,” Aagaard says.
“In the end, any one of those things we have decided are contaminants could actually be” in the placenta, Parkhill says. “It comes down to judgment, and a balance of probabilities. Do we throw away decades of biological understanding of the sterility of the placenta, or do we take this very weak and sporadic signal, which [could be] due to contamination, and claim that it’s real?”
In some cases, new evidence has changed his mind. “If you’d asked me 10 years ago, I would have told you that the lung is sterile,” he says. “That’s not the case now. There is a community there.” Other tissues are more controversial: Claims about a brain microbiome have been met with skepticism, and Parkhill says that “a lot of the signals I see in brain-microbiota papers are clearly contamination.”
It is hard to prove a negative, of course, but for the placenta at least, scientists have come close: For decades, they’ve successfully bred sterile mice by removing the uterus of a pregnant female, bathing it in sterilizing chemicals, cutting the pup out in sterile conditions, and raising it in a sterile environment. If placenta microbes exist, and pass from mother to fetus, this technique should be impossible. It plainly isn’t. “It is a bit startling that we are having a debate given that the ultimate experiment for sterility in the womb has been performed repeatedly, and routinely, for more than half a century,” says Maria Elisa Perez-Muñoz from the University of Alberta. (Aagaard counters that this is an extreme procedure that would likely remove any resident prenatal microbes; “It’s awfully darn-tooting hard to make a germ-free animal,” she says.)
Mysorekar feels that the technical side of this debate obscures more interesting questions. Like: If the placenta is sterile, how does that happen? If the fetus develops in a microbiological vacuum for nine months, why doesn’t it go into immunological shock when it’s suddenly exposed to millions of bacteria at birth? Why do fetuses seem to have activated immune cells, which usually only activate on exposure to bacterial molecules? “It makes biological sense that there’s some [prenatal] exposure,” she says, and if not through the placenta, then “where and how is that happening?”
It’s often said that academic fights are so vicious because the stakes are so low, but the dispute over the placental microbiome is more than just a niche academic squabble. As I’ve written about before, entire fields of science have been built upon shaky, nonexistent foundations, to the waste of effort, careers, and taxpayer money. If the human-microbiome field is to avoid making these mistakes, it needs to wrestle with exactly these questions over which methods to use and how data should be interpreted. Despite ballooning popularity, hype, and financial support, it is still a science in its infancy.