Ancient bacteria from nearly two miles below Earth's surface: that's what first drew Tullis Onstott to begin his search for life in the most unlikely of places. The geomicrobiologist had just attended a 1992 U.S. Department of Energy meeting about rocks estimated to be more than 200 million years old—older than most dinosaurs. These prehistoric rocks had been unearthed from a gas exploration well, and they turned out to be teeming with bacteria.
“That was pretty amazing to me,” says Princeton University's Onstott. “The idea that these bacteria had been living in these Triassic rocks since they were deposited at a time prior to the age of the dinosaurs, that idea caught my fancy,” he says.
These rocks were among the first substantial evidence that life existed miles underground, and they jumpstarted researchers’ efforts to study life in the so-called deep subsurface. Over the past 20 years, Onstott and others have found that there is a greater variety of life in a lot more inhospitable places than anyone had imagined.
Deep life has been found all over the world and under a variety of conditions—in oil fields and gold mines, beneath ice sheets in Greenland and Antarctica and in sediments and rocks below the ocean floor. These places can be extremely hostile environments, with pressures 10 to 100 times that at the surface. Temperatures can range from near freezing to more than 140 degrees Fahrenheit.
A mile or more below the surface there's no sunlight and very little oxygen. In these austere environments, creatures have to scratch out a living on whatever energy they can muster from their surroundings. This means that the pace of life down there can sometimes be incredibly slow. These microbes can be a thousand- or million-fold less abundant than their brethren above ground. And some may have been around for hundreds, thousands or even millions of years—real microscopic Methuselahs.
These creatures of the deep are diverse, consisting of bacteria and other single-celled organisms called archaea. There are even multicellular animals miles below the surface, including tiny worms called nematodes.
“What has been surprising as we continue exploring this deep hidden universe, is that it’s more complex down there than we could have possibly imagined when we started looking at Triassic samples back in the '90s,” says Onstott.
That complexity has opened up a world of possibilities for researchers, from cleaning up toxic waste to the search for extraterrestrial life. Some of these deep organisms feed directly on metals and minerals, and can affect groundwater by increasing or decreasing levels of arsenic, uranium and toxic metals. Scientists hope that these bacteria can soon be adapted to trap or remove such harmful substances from things like the wastewater leaking from a mine.
But perhaps most tantalizing is the idea that the conditions deep underground are so alien they may give researchers clues about where to find extraterrestrial life—and what that life might look like.
“It directly relates to whether life could be existing below the surface of Mars,” says Onstott. “That’s really what drew me into this field from the get-go, and still is a driver for me.”
Between the extreme environments and the relative scarcity of organisms, researchers go to great lengths—and depths—to study these microbes. They venture into mines and caverns or use drills to extract samples from below terrestrial sites or the ocean floor. In some areas it can take several days to get even a single sample. “Going to the ends of the earth and drilling, or going to the Arctic and going underground a mile to get a sample, it’s not easy,” says Onstott.

Probing the Hellish Depths
Almost a mile below Earth’s surface, deep within South Africa’s Beatrix Gold Mine, Maggie Lau looks for life. It’s hot and humid, and only headlamps breach the darkness as Lau, a geomicrobiologist in Onstott’s group at Princeton University, collects water from boreholes. These are holes drilled into the rock by geologists looking for gas and water pockets in advance of mining operations. Lau fills an assortment of vials with gas and water samples ranging in volume from less than a teaspoon’s worth to just over two pints. 

Maggie Lau collects borehole water in a vial more than two miles below Earth’s surface in South Africa’s TauTona gold mine. (Francois Vermeulen (Geosciences Manager, AngloGold Ashanti Limited))

The gas that Lau collects can reveal how ancient the water is. “The samples I am studying are around 40,000 to 80,000 years old,” she says. The water may have originated at the surface and trickled down through cracks over thousands or even millions of years, bringing microorganisms either from the surface or from shallower regions of the subsurface down with it.
Unlike the water, Lau takes a quicker and more dramatic route to the research site. She heads down a mine shaft in a lift-cage—which drops almost a mile in less than a minute—and then walks a mile or more with a loaded backpack. Some tunnels require researchers to crawl, dragging their packs behind them, or wade through knee- or thigh-high water in flooded sections. Occasionally the lift-cage isn’t available after a hard day’s work, and Lau and Onstott have to take the stairs back up. “We were joking that this was like a stairway to heaven,” she says.
In the hellish depths, where the water can reach 130 degrees Fahrenheit and the rocks themselves are often warm to the touch, there’s not a lot of life to be found. To gather as many living cells as possible for her analysis, Lau leaves some of her vials to filter hundreds to thousands of gallons of water over several weeks to a few months.
About a mile below the surface, Lau can usually find 1,000 to 10,000 cells in less than a teaspoonful of water. That might seem like a lot, but a pinch of soil from your backyard can contain 100,000 to a million times as many cells. At sites more than a mile underground, there might only be 500 cells per teaspoon of water. Lau estimates that she’d have to filter water continuously for 200 days to get enough DNA and RNA for her analysis.
It can be difficult to grow bacterial species in the lab without knowing the specific food they eat or the precise conditions under which they thrive. Scientists have only been able to grow about one percent of the bacteria they find at their deep field sites. As a result, most species are only known from their unique molecular signatures—and DNA or RNA sequencing has revealed a plethora of previously unidentified bacteria in the samples scientists have collected down there. 

Most recently, Lau is going a step beyond finding out what lives down there—she wants to know what they do for a living. Without sunlight and plants to trap the sun’s energy through photosynthesis, these deep-living bacteria have to survive on energy from the chemical reactions between rocks and water. These reactions can produce hydrogen, methane and sulfates, and scientists thought that those three chemicals would fuel the majority of bacteria living in these deep environments.
To her surprise, Lau found that this wasn’t the case. Instead, the chemicals sustain only a minority of the bacteria, which then produce sulfur and nitrates. Bacteria that fed on these secondary chemicals dominated in these environments.
This means that when searching for deep life either on Earth or on other worlds, scientists should look for a broader range of metabolic reactions. “Don’t just focus on the few major processes. We should be more open-minded to look at the full and complete metabolic landscape,” says Lau.
“Being able to actually see what they’re all doing down there now is absolutely the most exciting thing, something that we’ve been always wanting to do and trying to figure out how to do for the last 20 years, and now we can finally do it,” says Onstott.
“[Lau's] first snapshot, it’s like getting the first image back from Mars or something, it’s incredible,” he adds.

A Veritable Zoo
Where there's prey, there are usually predators. And bacteria make a tasty meal for a lot of creatures.
When Gaetan Borgonie heard about these deep bacteria, he wondered if he could find worms called nematodes—which feed on bacteria—in the same subterranean places. Borgonie, a zoologist at Extreme Life Isyensya in Gentbrugge, Belgium, had worked on these worms for 20 years. He knew that nematodes could survive a wide range of conditions at the surface, including extremely hot or cold temperatures and very low oxygen levels, so in theory, they were well suited to conditions deep underground.
Borgonie called up Onstott, who invited him to come explore the mines in South Africa. But finding these worms wasn’t easy. Although they are highly abundant on the surface, in the mines Borgonie had to sample more than 2,500 gallons of water to find a single nematode. “You really need to change your mindset and leave what you know from the surface, because underground is a different planet,” he says.
Borgonie discovered a large number of nematodes living in the mines in 3,000- to 12,000-year-old water from boreholes, as well as in stalactites hanging from the mine's tunnels. These included one new species found nearly a mile below the surface, and another unidentified worm living more than two miles down. These animals were the first evidence of multicellular, eukaryotic life this deep, Borgonie says.
Unlike the unique bacteria found at these depths, the vast majority of the worms belonged to species found on the surface. “These animals are already used to stress, and those that are opportunistic at the surface do very well underground,” says Borgonie.
Deep environments might actually offer some benefits, given the stable conditions and the lack of predators for the worms. “For them it’s like a holiday,” Borgonie says.

White arrows point to bacteria found within biofilms in borehole water from South Africa’s Kopanang gold mine. (Gaetan Borgonie)

Convinced that there must be more such creatures living in the mines, Borgonie left his sampling equipment in South Africa's Driefontein gold mine for two years to filter more than three million gallons of water—enough to fill almost five Olympic-size swimming pools.
“That’s when we found the entire zoo,” Borgonie says. He identified several other multicellular organisms, including flatworms and segmented worms, as well as what appeared to be a crustacean. Nearly all of these species survived by eating bacteria.
The discovery of these organisms is encouraging for scientists looking for extraterrestrial life, Borgonie says. “I think it’s very good that we find such a huge ecosystem underground,” he says. “If we can prove that they can survive indefinitely underground, then it might be very good news for people searching for life on Mars.”
“I would really love [to be doing] this work on the planet Mars,” he says. “That’s why I always say, if they ever give me a one-way ticket to Mars, I’m gone.”

The Alien Deep
Borgonie may not have his ticket just yet, but upcoming space exploration missions could give us a better idea of whether other parts of the solar system could support life.
“One of the things that has given people a sense of optimism where astrobiology is concerned is the finding that there are organisms that can persist in what we would consider very extreme conditions,” says Tori Hoehler, an astrobiologist at the NASA Ames Research Center. Hoehler is a member of the NASA Astrobiology Institute’s Rock-Powered Life team, which studies how reactions between different kinds of rocks and water can generate enough energy to support life.
“One of the most prevalent habitats that is available out there is the one defined by rock and water,” says Hoehler. You can imagine aquifers sitting deep under Mars' surface or the oceans sloshing above the rocky crust of Jupiter's moon Europa or Saturn's moon Enceladus, he says.
NASA’s Europa Multiple Flyby Mission, expected to launch in the next five to ten years, will give scientists a better idea of whether Jupiter's icy moon has any environments that could support life. As for Mars, researchers have gone from asking whether they can find habitable environments to actually looking for evidence of life itself, says Hoehler.
Even though conditions on the Martian surface are currently extremely inhospitable to life, the planet appears to have had an atmosphere and surface water at some time in its past. If life had evolved then, it could have spread to the Martian subsurface, where the environment stayed stable even as the surface turned hostile. It’s possible that life still persists deep underground, waiting for us to dig it out.
An artist's rendering of ESA's ExoMars Rover, which will carry a drill designed to probe down to 6.5 feet below the Martian surface. (ESA)

We won’t have to wait too long to get our first peek beneath the Martian surface. The European Space Agency’s 2018 ExoMars Mission will drill about six feet below the Martian surface to look for signs of life. That may not be deep enough to find living organisms, but it should be far enough below the surface that we could find evidence of life.
More than 20 years since ancient bacteria first gave him a glimpse into Earth's deep life, Onstott can’t wait to see what we find on Mars, especially once scientists can dig a little deeper.
“If there’s a sweet spot on Mars, someplace where you just get the right balance of temperature and water, then there might be organisms surviving under those conditions.”



Read more: http://www.smithsonianmag.com/science-nature/inner-earth-teeming-exotic-forms-life-180958243/#jE0FO5IcWrQ0EL6c.99
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