Pools in the Mexican desert are a window into Earth’s early life


Valeria Souza Saldívar never planned to devote her life to a remote and ancient oasis more than 1000 kilometers north of her laboratory in Mexico City. But a call in early 1999 changed that.

“It’s one of the best cold calls I’ve ever made,” says James Elser, a limnologist at the University of Montana. He had picked up the phone to invite Souza Saldívar to join a NASA-funded astrobiology project in Cuatro Ciénegas—a butterfly-shaped basin with colorful pools, or pozas, in the middle of Mexico’s Chihuahuan Desert.

Neither Souza Saldívar, a microbial ecologist at the National Autonomous University of Mexico, University City, nor her ecologist husband and research partner Luis Eguiarte Fruns, also at UNAM, had ever visited Cuatro Ciénegas in the state of Coahuila. That first trip convinced them to completely change their research plans. “Looking at those mountains and the water, I fell in love,” Souza Saldívar says.

The landscape—more than 300 turquoise-blue pozas scattered across 800 square kilometers, among marshes and majestic mountains—wasn’t the only draw. The waters, whose chemistry resembled that of Earth’s ancient seas, teemed with microbes; unusual bacterial mats and formations called stromatolites carpeted the shallows. When Souza Saldívar first cultured the organisms from the pozas, “The amount of microbes was enormous, as was the diversity of colors and colony sizes,” she recalls. For her, this remote microbial hot spot was an irresistible mystery.

Since then, work by Souza Saldívar, Eguiarte Fruns, and a widening circle of collaborators in Mexico and the United States has shown that Cuatro Ciénegas—which means “four marshes” in Spanish—is one of the most biodiverse places on the planet. “There’s nowhere that has so much ancient diversity of microorganisms,” says Michael Travisano, an evolutionary ecologist at University of Minnesota, Twin Cities, who has collaborated with the Mexican researchers since 2001. Among the most recent additions to that menagerie are hundreds of species of archaea, the ancient microbes that may have given rise to eukaryotes—organisms with complex, nucleated cells.

The diversity includes strains with unusual adaptations, such as the ability to build their lipid membranes with sulfur instead of the usual phosphorus, which is scarce in the waters of the pozas. It includes potential sources of new compounds for medicine and agriculture. And it poses a question that has occupied Souza Saldívar and Eguiarte Fruns for the past 20 years: How did this Noah’s Ark of ancient microbes arise? “It’s a dream for every biologist to know the origin of diversification,” Souza Saldívar says.

But her dream might be short-lived. Since the 1970s, farmers have intensively drained water from the pozas and rivers to irrigate nearby fields of alfalfa, grown for cattle fodder, gradually drying the improbable oasis. Souza Saldívar has galvanized a conservation effort that has slowed the drainage; in the coming weeks, a canal that removes 100 million cubic meters of Cuatro Ciénegas’s water annually is scheduled to close. In the meantime, the researchers have been trying to describe as much as they can, as fast as they can, before their beloved pozas dry up and the precious microscopic life that has survived undisturbed for millions of years dies off.

CUATRO CIÉNEGAS SERVED as a stopping point for hunter-gatherers for thousands of years. To date, 50 archaeological sites with cave paintings—some dating to 2275 B.C.E.—have been found in mountain caves around the basin. Much later, the region made a mark on history when Venustiano Carranza, born in a village at the basin’s margin, became a leader of the Mexican Revolution and president of Mexico from 1917 to 1920. Nowadays, the village is called Cuatro Ciénegas de Carranza after him.

But in the 1960s, Cuatro Ciénegas started to become famous for its biodiversity, as biologists began to describe new species of snails, fish, turtles, and plants found in the pools and marshes—and often nowhere else.

Wendell “Minck” Minckley, a renowned ichthyologist at Arizona State University (ASU), Tempe, was first lured to Cuatro Ciénegas after learning that the world’s only aquatic box turtle (Terrapene coahuila) lived there. Over the years, Minckley made frequent trips to the pozas, describing their snails and fish (Herichthys minckleyi, a cichlid, bears his name) while making connections with the local people.

Minckley also noticed peculiar, rocky structures in the pools. They were stromatolites, biological structures normally found as fossils dating back as much as 3.5 billion years. Colonies of photosynthesizing bacteria, which boosted early Earth’s oxygen, created the layered formations by depositing carbonates and trapping sediment in ancient, shallow seas. But these stromatolites were alive. Also found in other extreme environments such as Australia’s warm, salty Shark Bay, living stromatolites “are sort of a window into early Earth,” Elser says. The pozas also nurture bacterial mats, a soft form of stromatolites normally found deep in the ocean.

As early as the 1970s, Minckley realized the pools and their diversity were under threat: Local farmers were carving canals to tap their water. Thanks in part to his lobbying, the Mexican government in 1994 designated an 85,000-hectare protected area. But the drainage continued. “Minckley knew that Cuatro Ciénegas was going to die,” Souza Saldívar says. He thought NASA might be its salvation.

In 1998, NASA established its Astrobiology Institute, a network of researchers studying life in extreme environments that might resemble conditions on other planets. Minckley saw an ideal astrobiology study site in the waters of the pozas, with their seemingly inhospitable chemistry and living stromatolites. But he was no expert on extreme environments, so he enlisted Elser, who specializes in how water chemistry affects ecosystems and also works at ASU. After they submitted a 1998 proposal to fund the project, however, NASA said they should add experts on microbiology and evolution—and those experts had to be Mexican to help secure permits to obtain samples. Based on colleagues’ suggestions, Elser called Souza Saldívar and Eguiarte Fruns, newly minted professors at UNAM. They joined, and NASA approved the 3-year project.

With two children in tow, the couple met Minckley and Elser at Cuatro Ciénegas. Next to the turquoise-blue waters of La Becerra poza, Minckley told them he believed the ecosystem was a glimpse of deep time. “Do you see these miniature snails in my hand?” Souza Saldívar recalls him saying. “I just scooped them from the springhead, but their direct ancestors were eating sulfur bacteria in hydrothermal vents 220 million years ago in the bottom of the ancient Pacific.”

Based on the water chemistry—low in phosphorus, iron, and nitrogen—and the presence of living stromatolites, Minckley believed Cuatro Ciénegas re-created the marine conditions found worldwide millions of years ago. He challenged the two researchers to explore its mysteries—and to protect its pozas. “Only you, as Mexicans, can save them from the extinction caused by humans,” Souza Saldívar recalls him saying.

Minckley died 2 years later, in 2001.

TO INVENTORY THE FULL DIVERSITY of microbes at Cuatro Ciénegas and trace their relationships, Souza Saldívar needed to study their DNA. To do so, scientists normally take microbial samples from a site and grow them in a lab. But many bacteria and archaea are difficult to culture, and only a few groups at the time had successfully analyzed DNA isolated directly from the environment. High magnesium levels in the water and “slime” from the microbes made isolating DNA from the pozas especially difficult.

But Souza Saldívar and her students Ana Escalante and Laura Espinosa Asuar made a start. In 2006, they reported in the Proceedings of the National Academy of Sciences that they had found 38 distinct groups of microbes—four times as many as in a typical salt marsh—corresponding to 10 major lineages of bacteria and one of archaea. Half the bacterial groups were most closely related to marine microbes. Almost 10% of the groups resembled ones that live on hydrothermal vents—fissures deep in the ocean where microbes thrive despite extreme heat and mineral concentrations.

As Minckley had suspected, Cuatro Ciénegas had somehow preserved ancient marine life forms deep in the desert, more than 500 kilometers from the Gulf of Mexico, at a site where the last seas retreated some 20 million years ago.

“The deep time aspect [of Cuatro Ciénegas] is very surprising,” Travisano says. It is a true “lost world,” preserved by the hostile water chemistry, he and the Mexican team argued in a 2018 paper in eLife. Millions of years ago, they proposed, ancient marine ancestors found their way to the place, adapted to the extreme environment, and didn’t change much.

The pozas themselves are not particularly ancient. The springs that nurture them are fed by deep aquifers in Sierra San Marcos y Pinos, filled with water accumulated during the last ice ages, Eguiarte Fruns says. Now, the water seeps to the surface because of an active fault beneath the basin. It rises through ancient marine sediments, picking up its unusual chemistry along the way. Somehow, the ancient microbes persisted and diversified in a succession of springs that must have appeared and vanished throughout geologic time. As in an ancient clock, Souza Saldívar says, all the original mechanisms are still working together to sustain unusual life.

To Frederick Cohan, a microbial ecologist at Wesleyan University who is not part of the Cuatro Ciénegas project, the fact that many of the microbes are related to marine species and not species found inland is compelling. “I think it’s saying those organisms are anciently there.”

WHEN THE RESEARCHERS looked at the stromatolites, they found even more diversity. Samples from one site, Pozas Azules II, yielded more than 58,000 distinct microbial sequences, predominantly from bacteria—not a direct count of species, but an indicator of biodiversity. In the Río Mezquites, a stream that flows through the northern part of the basin and recharges several pools, they identified 30,000 sequences, mostly from cyanobacteria. More than 1000 sequences from Pozas Azules II appeared to be from archaea, the researchers reported in Environmental Microbiology in 2009. The stromatolites also teemed with bacteria-infecting viruses—strains that were unique to each pool and resembled marine viruses.

Studying the microbes hasn’t been easy. “There are thousands and thousands of new bacteria that we can’t grow in culture,” Souza Saldívar says. They could, however, identify some startling adaptations to the extreme conditions. In one bacterium found only in El Churince, a system of lagoons and pozas on the western part of the basin, researchers sequenced the smallest genome ever found in its genus, Bacillus. The work, led by Gabriela Olmedo Álvarez, a genetic engineer at Center for Research and Advanced Studies of the National Polytechnic Institute, Irapuato, also showed that the microbe—B. coahuilensiscould synthesize membrane sulfolipids. This meant that, like some plants and cyanobacteria, it could use sulfur from the environment—instead of phosphorus—to form its cell membranes.

Lost arks

Shallow, mineral-rich pools and lagoons, with conditions like those in ancient oceans, are hot spots of microbial diversity. Floating mats at Cuatro Ciénegas teem with the primordial microbes known as archaea, leading researchers to call them “archaean domes.”

“It likely ‘stole’ these genes from a cyanobacterium,” Olmedo Álvarez says, enabling it to cope with scarce phosphorus, a condition thought to have prevailed in Earth’s earliest oceans. The microbe’s small genome may also have helped it thrive, as it required less phosphorus to build its DNA. Olmedo Álvarez thinks the organism may offer a glimpse of the stratagems used by early microbes to adapt to their new environment.

“We’re just starting to understand the depth of diversity,” says Olmedo Álvarez, who found that B. coahuilensis is itself starting to split into strains with variations in phosphorus metabolism.

The low phosphorus conditions found in Cuatro Ciénegas not only promoted local adaptations, but also accelerated microbial diversification, Souza Saldívar and Elser argued in a perspective published in 2008 in Nature Reviews Microbiology. Bacteria normally share bits of DNA with their neighbors in a process called horizontal gene transfer, which blurs the divisions between strains. But in Cuatro Ciénegas, the microbes—hungry for phosphorus—essentially consume free DNA rather than incorporating it into their genomes. “They will eat the DNA to get the phosphorus,” Elser says.

Besides offering insights into evolution, Cuatro Ciénegas’s microbial diversity may hold practical payoffs. “Cuatro Ciénegas is one of the richest places on the planet for genetic resources,” Souza Saldívar says. For example, most modern antibiotics are derived from actinobacteria, which are abundant in the pozas. Susana De la Torre Zavala, a biotechnologist at the Autonomous University of Nuevo León (UANL), University City, is searching for potential antibiotics in a library of 350 actinobacteria from the basin. Her team has also found that an extract from a microalga living in the pools shows anticancer activity.

Agriculture, too, could benefit, Olmedo Álvarez says. By 2050, the reservoirs of phosphorus that help sustain global harvests could become scarce, and the microbes’ ability to concentrate the element from different sources could hold solutions. “We’re understanding Cuatro Ciénegas, but we’re also understanding basic principles of ecological interactions that have an application in medicine and agriculture,” she says.

AS THE SCIENTIFIC STORY of Cuatro Ciénegas unfolded, its fate has hung in the balance, with Souza Saldívar fighting a long series of battles over its water with local farmers and landowners, dairy companies, and politicians. Her weapons have been her rising scientific profile and a tireless outreach to the public, especially young people.

Souza Saldívar has drawn fire—during a 2013 microbiology congress, police had to protect her from protesting locals—but she has won a series of victories. In 2007, the daughter of the CEO of LALA, a giant dairy consortium with roots in the state of Coahuila, told her father she wouldn’t speak to him because “he was killing Cuatro Ciénegas,” Souza Saldívar says. The executive promptly scheduled a meeting with the scientist. “You need to change your cows’ diet,” Souza Saldívar says she told him, refusing to accept a courtesy yogurt he offered. “I’ll accept your yogurt when you do so.” He promised not only to stop buying the region’s alfalfa, but also to invest in environmental education projects for local children.

Two years later, she won an unusual ally, the powerful Mexican billionaire Carlos Slim. His foundation collaborated with the World Wildlife Fund (WWF) to buy the land surrounding El Churince in the western basin, and to provide researchers with a 5-year, 18 million Mexican peso ($1.4 million) grant to study Souza Saldívar’s favorite poza. This allowed them to set up the infrastructure to perform long-term experiments. But it did not save the water.

In 2010, Mexico’s National Water Commission (CONAGUA) set out to replace the open, leaky canals, which lose 75% of the drained water, with less wasteful enclosed conduits. But the project was abandoned midway—most likely because of corruption—and the old canals were never closed. As Cuatro Ciénegas continued to dry up, the researchers raced to study El Churince, finding 5167 distinct species of bacteria and archaea in the last remaining pool. A close inspection of the genomes of Bacillus bacteria from one single square kilometer increased the known diversity of the group by more than 20%. By comparing DNA sequences, the team traced the Bacillus diversity to two ancient ancestors, one dating back 680 million years, the other 160 million years. Those dates coincide with the breakup of the supercontinents Rodinia and Pangaea, respectively, and the team thinks the oceans that formed during those convulsions carried the ancestral microbes to what is now the Cuatro Ciénegas Basin, where they have persisted ever since.

Cohan says that’s plausible. Bacillus from elsewhere fail to thrive in Cuatro Ciénegas, most likely because they are outcompeted by the local microbes and can’t adapt to the extreme conditions. And the Bacillus species from Cuatro Ciénegas are not found anywhere else in the world. “It’s just bizarre,” Cohan says, but it makes the pozas so much more valuable and worth saving. “It’s kind of a paleontological microbial park.”

In 2016, El Churince dried up just after the funding from the WWF–Carlos Slim Foundation ended. The researchers felt devastated. Souza Saldívar says it was painful to see turtle shells lying on the now-barren soil. “It’s really sad,” Olmedo Álvarez says. “It’s gone.”

ON THE EASTERN SIDE of the basin, things are looking brighter. In 2000, the conservation nongovernmental organization Pronatura Noreste acquired the Pozas Azules ranch: 2721 hectares hosting about 100 pozas. Pronatura eventually gained rights to the water as well, enabling it to close canals draining the pozas in the ranch. Farmers are now encouraged to adopt water-sparing drip irrigation, and some are growing nopal—an edible cactus popular in Mexican cuisine—which requires much less water than alfalfa.

The researchers have focused their recent studies on Pozas Azules. In 2019, after an unusual spring rain, the team noticed alien-looking structures in the shallow waters of a site near Pozas Azules II: white microbial mats buoyed by gas. The gas appeared to be largely methane, and a genetic analysis showed the mats were teeming with archaea—230 distinct species, they report in a preprint. That makes the spot “the most diverse place of archaea that we know of,” De la Torre Zavala says.

Now, the team hopes to analyze samples from the structures, which it calls “archaean domes,” in search of the elusive Asgard archaea, organisms previously found only in the deep ocean and thought to hold clues to the evolution of simple microbes into complex eukaryotes. Although some in her team are skeptical, Souza Saldívar is convinced they will find them. “Valeria’s usually right,” De la Torre Zavala says.

Such prospects have added to Souza Saldívar’s determination to preserve Cuatro Ciénegas, and she is enlisting young people for support. In every field trip since 2004, her team has spent time with students from the local high school, showing them how to use a microscope and take simple environmental measurements, and teaching them about sustainable agriculture. In 2011, with funding from the LALA Foundation and the WWF–Carlos Slim Foundation, the scientists set up a college-level molecular biology lab at the school, which is now ranked among the best rural high schools in Mexico.

Héctor Arocha Garza is one of its graduates. Inspired by the secrets of Cuatro Ciénegas, he pursued a Ph.D. in biotechnology at UANL with De la Torre Zavala, then returned to his hometown. “My heart was in Cuatro Ciénegas,” he says. Now, he’s leading the scientific branch of a privately funded megaproject called Cuatro Ciénegas 2040 that aims to build a science museum and make Cuatro Ciénegas a scientific tourism destination, while supporting education and medical care for the village’s young people.

The effort comes at a critical moment. More than 90% of the marshes are gone, and some pozas and lagoons are dry. But this year, CONAGUA committed to regulating water usage and closing illegal wells, and Pronatura Noreste will close the Saca Salada Canal, which drains the Río Mezquites, as soon as the COVID-19 pandemic permits.

Those developments, and stories like Arocha Garza’s, give Souza Saldívar hope for the future of Cuatro Ciénegas. “It has been a very complicated, long, and difficult process,” she says. But now, she wrote in a recent book, “There is a revolution occurring in this oasis: Science is the tool and kids are the drivers.”

sciencemag.org, 30 June 2020
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