Just past the intersection of Centre and Locust in Centralia, Pennsylvania, the microbiologist Tammy Tobin turned the wheel of her aging Prius sharply to the right. As the windshield wipers whipped furiously back and forth to fend off the driving sleet — a reminder that winter had yet to bid farewell — Tobin announced, “We’re here.” We were at the base of a grassy slope nestled behind the SS. Peter and Paul Cemetery. It looked like any of the other countless knolls tucked in the anthracite hills of eastern Pennsylvania. But almost 50 meters beneath our feet lurked a hidden menace. Centralia was burning.
Or rather, the coal seam under what used to be the town of Centralia was burning. The coal has burned for more than 50 years and will likely burn for centuries to come. As we climbed a low rise that hugged the back of the Catholic cemetery, no flames were visible, only puffs of steam where the dirt exhaled excess heat and the grass stubbornly refused to don its icy cap. All but a handful of the town folk had fled when the government revoked Centralia’s postal code in 2002. But Tobin, from Susquehanna University 30 miles west of Centralia, wasn’t here to comb through the wreckage of a once-thriving town.
Instead, she and a group of collaborators had set their sights on something much smaller. The heat and pollution from the underground fire wasn’t just stressful for Centralia’s flora and fauna; it also created a crisis for the area’s microbes. The trillions upon trillions of microscopic single-celled organisms at home in Centralia’s soil suddenly found themselves living in a veritable sauna. It was adapt — or die. Or so scientists thought.
“Centralia is a beautiful sandbox for asking about what happens during an environmental disturbance,” said Ashley Shade, Tobin’s former student, now a microbiologist at Michigan State University and a collaborator on the project. “Even when that disturbance is like a sledgehammer.”
The coal-seam fire at Centralia provides researchers with the perfect opportunity to test a new idea known as a microbial seed bank: that commonly overlooked dormant individuals make up a vast reservoir of biodiversity, ready to spring to life when environmental conditions change. Although scientists had found hints from laboratory and environmental experiments that such a seed bank exists, Centralia represents a rare opportunity to see whether and how a microbial seed bank functions in the real world.
900 Degrees Fahrenheit on the Ground
No one knows exactly how the fire under Centralia started; local legend holds that somebody accidentally ignited the seam while burning trash just outside one of the mine shafts. What is known for certain is that, shortly before Memorial Day in 1962, residents of Centralia reported that a fire had started in the town’s coal mine just east of the Odd Fellows Cemetery. It soon became obvious that even the most aggressive methods wouldn’t stop the spread of the flames. Residents would simply have to wait for the fire to burn itself out. But in an area that dubbed itself “coal country,” there was no lack of underground material to burn through, so the fire outlasted the people. Although residents initially hoped that, because the fire was entirely below ground, they would be able to continue living in Centralia, the release of toxic gases and the opening of sinkholes made it too dangerous.
Most families left by choice or were bought out by the government. A few families, brave or foolhardy (take your pick), continue to live in Centralia, fire be damned. Although Centralia may have had the sharpest reversal of fortune, the economy of the entire region has gone from bad to worse over the past several decades.
Ashley Shade is all too familiar with the travails of central Pennsylvania. She grew up a stone’s throw from Centralia, and although she knew about the fire — you couldn’t really live near Centralia and not know about it, she says — she never really gave it much thought. It wasn’t until her first genetics class as an undergraduate at Susquehanna University in 2002 that she began to think of Centralia as something more than a nearby oddity. The year before, a team of geologists and soil scientists at Susquehanna had approached Tobin, who was Shade’s professor at the time, about setting up a formal study of how the fire was changing Centralia. They asked Tobin if she would help study the soil microbes in Centralia. Although she knew nothing about microbiology, she found the topic quirky and interesting, and so she agreed. She asked her students in 2002 whether any wanted to join her new project in Centralia or an existing study on bovine genetics.
Once the Centralia project started, “it was very hard to get people to work on bovine immunogenetics,” Tobin said drily.
Both Shade and Tobin immediately fell in love with Centralia. The team staked out a range of sites spanning three contrasting areas: one above a spot where the fire had never been, one above where the fire was currently burning, and one where the subsurface flames had already come and gone. This would give the researchers an idea of how the soil microbes changed over time. Some never-burned sites were especially important because the fire was moving in that direction. Tobin and her fellow scientists could track what happened to the soil in real time.
Seventeen years ago, when sequencing the genomes of large numbers of environmental microbes was prohibitively expensive, studying the genetics of soil microbes meant scientists would chop the DNA into small pieces. Each different species of microbe yielded a collection of genetic fragments that could be sorted by size. Using a probe to highlight ribosomal DNA sequences unique to each species, scientists could derive a genetic fingerprint for a microbe and identify its species by comparing their results to a large database of known prokaryotes. Although this “ribotyping” was more time-consuming and less precise than current molecular methods, it nonetheless provided Tobin and Shade with the first clues about what, if anything, may have survived Centralia’s below-ground inferno.
“A place could go from being cool to being very hot pretty quickly, and it fluctuates with all kinds of climatic and geological factors,” Tobin said. “Could things adapt quickly enough?”
Depending on how much oxygen could reach the fire, the flames under Centralia could burn as hot as 1,350 degrees Fahrenheit, and ground temperatures sometimes exceeded 900 F. In 2007, a German documentary film crew bought a single egg from a local cafeteria so they could fry it by a steam vent and eat it for breakfast as an on-camera gimmick. However, the egg didn’t fry. Instead, the soil was so hot that, with a quick sizzle and hiss, the egg charred beyond recognition before the crew could frame their shot, leaving nothing for their toast or their viewers. Under such extreme conditions, Tobin told me as we wound our way along the Pennsylvania byways from her lab at Susquehanna to Centralia, it was entirely possible that nothing had survived. To her delight, she was wrong.
In a 2005 study in Soil Science, Tobin and colleagues showed not only that microbes survive in the soil above actively burning areas, but that some species thrive there. The overall level of diversity was the same in hot areas (with temperatures ranging between roughly 90 F and 170 F) as in areas that the fire had yet to reach. When the researchers looked more closely, they found that although the overall bacterial diversity decreased with higher temperatures, even the hottest samples apparently still held thriving microbial communities. Shade and Tobin also identified heat-loving bacteria (thermophiles) that resembled microbes living near geothermal hot springs in Iceland, though their data wasn’t detailed enough to say just how closely the organisms were related.
What their data couldn’t tell them, however, was whether the microbes living over the fire had lurked there all along in very low numbers or whether they had been blown in or had otherwise arrived from afar, perhaps from other geothermal areas around the globe. It was anyone’s guess as to which might be right.
Not Dead but Dormant
As Tobin and Shade continued chipping away at the microbial mystery in Centralia, the Indiana University biologist Jay Lennon had a mystery of his own. As the cost of genetic sequencing plummeted and computer programs grew more sophisticated, it became possible for researchers to sequence DNA directly from environmental samples, for studies known as metagenomics. For the first time, scientists didn’t need to culture organisms to study them in the lab. Just by sequencing the DNA of microbes in an environment, they could find out what lived there and in what quantity.
But “the abundance of an organism doesn’t tell us whether it is active,” said Alexander Loy, a microbiologist at the University of Vienna. To assay metabolic activity, biologists use strategies such as measuring how much RNA an organism is making; because RNA is a much shorter-lived molecule than relatively durable DNA, it’s a truer indicator of current metabolism and not just the cell’s existence. By analogy, a census taker can count all the buildings on a city block, but that alone won’t say whether they are homes or businesses, or if they’re currently occupied. For those answers, the census worker might need to conduct interviews door to door, or measure water and electricity usage.
When Lennon began looking at biological samples from lake water, soil and even feces in 2010, over and over again he found the microbial equivalent of abandoned buildings. Lots of species were there, but a large proportion of the microbes in seemingly every environment didn’t appear to be doing anything.
These dormant microbes with very reduced metabolic activity exist in a liminal space between life and death. They might not be doing many of the activities typically associated with life, such as growing, eating or replicating their genes, but they are also very clearly not dead — because sometimes they will become animated again. “Go to sleep, if you will, and you have the ability to wake back up,” Lennon said.
The concept of dormant microbes was at least a century old, but biologists thought they were rare. Most of what was known about dormancy came from bacteria that formed hardy spores, including Bacillus anthracis, the soil microbe famous for causing anthrax. The ability to form spores can protect a bacterium from everything: high doses of ultraviolet and gamma radiation, prolonged drought, the vacuum of space. “People have resurrected bacteria from amber,” he said.
The downside of relying on spores as a survival strategy is that it is extremely demanding. Ten percent of the B. anthracis genome is devoted to forming spores, and the process can take more than five hours, start to finish. With such high biological startup costs, this ability only evolved once in a single group of bacteria, as far as researchers know. This suggested that such Lazarus microbes are tiny oddities.
Data from Lennon and other microbiologists, however, indicated that dormancy might be the rule, not the exception. “More than 90 percent of the microbial biomass [in soil] is inactive,” he said.
Dormancy explained how so many microbes — up to 1010 cells per gram of soil — could coexist. In some sense, they didn’t, at least not all at the same time. Rather than using up valuable resources by fighting each other for food and space, microbes could instead enter a dormant phase to wait for better environmental conditions. Dormancy also gave microbes a way to survive the feast-or-famine waves of food and other essentials, as well as the limitations of extreme environments. Dormant organisms aren’t as hardy as spores, but their quiescent state means they don’t have to waste valuable resources coping with stressors. Temperatures that might kill a rapidly dividing organism can become bearable if the microbe doesn’t have to find food, make proteins and attend to other housekeeping tasks. As a result, the quiescent organisms can tolerate a wider range of temperatures and other environmental conditions than if they were growing as usual. Borrowing a phrase from botany, Lennon called this vast reserve of dormant organisms the “microbial seed bank,” which was just waiting around for the right environmental conditions to grow and thrive.
Scientists used to believe that the soil microbes found in the deserts of Antarctica were the same as those found in the Amazon rainforest, but studies showed that, like all organisms, soil microbes are highly adapted to local conditions. For this reason, Lennon doesn’t think the Earth has a global microbial seed bank. Instead, each soil community, such as the dirt in Centralia, has its own local seed bank. Local microbes deposit themselves into the seed bank when conditions are less than ideal. Microbes from elsewhere can also hitchhike into the area, arriving on the feet and feathers of birds or blowing in on the wind. Some of them may try to make a go of it and either thrive or die out, but others will hunker down and wait.
The microbial ecologist Genoveva Esteban of Bournemouth University in the U.K. saw the microbial seed bank at work in Priest Pot, a 10,000-year-old pond in northern England’s Lake District. Esteban brought samples of microbial eukaryotes (small, single-celled organisms with a nucleus) from Priest Pot back to the lab to grow. Like their nucleus-deficient prokaryotic brethren, eukaryotes are challenging to grow in culture. Most just don’t want to grow in the lab. When Esteban peeked at drops of lake water under the microscope, she saw hundreds of types of swirling and swimming creatures. In the lab, she could identify only 20 species growing in the culture bottle. Then she divided the culture and grew it in a range of environments. (“We really squeezed our imaginations” to come up with every possible combination of conditions, Esteban said.) Three months later, she had 135 species.
“There were all these hidden organisms, just waiting for the right conditions to appear,” she said.
The same thing happened when Esteban took samples from Andalusia’s salt pans, those hypersaline remnants of ancient seas in what is now southern Spain. Initially, she could detect only seven microbial species in samples from six different salt flats. She gradually diluted those samples and let them grow for five weeks or more, and the number of species shot up to 95.
In a sense, Esteban’s deliberate environmental manipulations mimicked what happens when conditions shift in the natural world — including what happens as the climate continues to warm. High in the Alaskan Arctic, Janet Jansson, a microbiologist at Pacific Northwest National Laboratory outside of Richland, Washington, was tracking how global warming was affecting microbes in Hess Creek. For thousands of years, the subsurface soil in the area had been permanently frozen, but global warming is changing that, causing the underground soil layers to begin to thaw.
In results published in Nature in 2011, Jansson found that after thawing a sample for just 48 hours, she could begin to see a shift in the community’s DNA. This hinted at a rise in the abundance of carbon-eating bacteria, as opposed to the type of microbe that’s usually found in the permafrost, eking out a living by using iron as an energy source.
Later sampling of both thawed and frozen sites, backed up by RNA analysis, confirmed that the DNA wasn’t lying. In the thawed soil, the iron-reducing microbes had been largely replaced by others using organic carbon for food. These differences, Jansson found, were inherent in the system.
“It’s a very extreme difference in function,” she said. “These organisms are already there, just in low numbers. The environment selects for what’s able to thrive.”
From an ecological perspective, Loy says, seed banks provide the system with a kind of insurance policy. “If you take antibiotics, those with resistance genes can grow and take over those empty niches,” Loy said. Seed banks function the same way, with dormant organisms becoming dominant when environmental conditions change. Tobin and Shade hypothesized that a microbial seed bank could explain some of what they were seeing in Centralia. Their long-term experiments were cranking along, giving them the perfect opportunity to test this idea, when suddenly, disaster struck.
A Backup Plan for Ecosystems
Just as Centralia has attracted plenty of oddball microbes, it has also attracted weirdos of the more bipedal variety. During the freezing afternoon Tobin and I spent traipsing around Centralia, several cars pulled off Locust Avenue to ask us for directions to the fire. “This always happens,” she told me as the cars drove off, disappointed to learn that they wouldn’t be able to see any flames. The old Route 61, which partially collapsed for the final time in 1993 and forced the state to build a bypass, still exists as an asphalt canvas for graffiti artists. Their work ranges from the prosaic “L+L 4EVER” to pieces of a more scatological and sexual nature.
Tobin shrugged off much of this vandalism. But in 2006, a group of treasure hunters tore through much of the land in and around Centralia, digging up soil on their hunt for antique glass. One of the locations they destroyed was the long-term study site that she and Shade had developed. Overnight, half a decade of work was lost.
“It looked like it was a war zone,” Tobin said.
By the time Tobin got her research fully up and running again, Shade had completed her doctorate in microbiology at the University of Wisconsin, Madison, and found a faculty position at Michigan State University. She never forgot her time at Centralia, however, so in 2014 she called up her old professor and asked Tobin to collaborate. That October, Shade and her lab manager hopped on a plane and flew to Pennsylvania.
Armed with trowels, quart-size canning jars, and ample bleach to sanitize their shovels and shoes, the squad descended onto what remained of the town and began taking soil samples. The team dug up soil from several locations once again: over an active fire, over regions that had burned and since cooled, and over parts of the mine that had never caught fire. After carefully packing the filled canning jars into a large cooler, Shade returned to East Lansing and began to study the dirt in her lab.
She started by comparing the species living in each of the three groups. She initially believed that the microbes from the actively burning areas would have the least variation between sites: The challenges of growing in such extreme heat, she figured, would severely limit what kinds of organisms could grow. After the fire burned itself out and the ground cooled, Shade expected that the microbes would return to a more diverse state. In fact, she and Tobin found the exact opposite: Microbial populations in the hot areas diverged and then re-converged as the ground cooled over a period of 10-20 years.
“Microbial communities have an immense capacity to respond and recover,” Shade said. “There seems to be this inherent capacity in the system that’s just sleeping.”
Regardless of how the microbial populations changed, Shade and Tobin hypothesized that Centralia’s microbial seed bank was what allowed the system to respond to the temperature surge from the fire and return to its initial state. A further study in PLOS ONE showed that the seed bank may also have allowed the soil to respond to increased levels of arsenic and other heavy metals that the fire released. To Esteban, that’s the entire point of the seed bank.
“A seed bank means that ecosystem function will never stop. Even if conditions change, the ecosystem can keep going,” she said.
The process also benefits individual species. “Most microbes live on a razor’s edge between life and death,” Lennon says. “And going dormant is better than dying.” Exactly what triggers this dormancy, however, remains unclear. Nor do scientists know whether the entire population of a microbe will opt for dormancy or if some might become dormant as a hedge fund for their brethren that try to make a go of it, even in adverse circumstances.
For now, however, the role of the microbial seed bank and even its very presence remain conjectural. Shade and her grad students drive back to the abandoned town every fall to gather more samples. On her most recent trip, she took soil samples from never-burned sites, brought them back to the lab, and began heating them up under controlled conditions to see how they responded. This set of experiments is still ongoing, but Shade hopes that it can begin to answer some fundamental questions about the role of seed banks. Those answers won’t just provide insight into what’s happening at Centralia or at thousands of other coal-mine fires throughout the world. They could also yield valuable clues as to how the world’s microbes will respond to a warming climate.
Centralia’s position at the fulcrum of climate issues is based on more than just its microbes. Even as the town’s coal continues to burn below ground, several wind turbines have been erected at the top of a nearby ridge. Whether the town will be able to demonstrate the kind of resilience shown by its local microbes remains to be seen. In the meantime, the turbines continue to turn slowly in the breeze.