The search for life in polyextreme brines

Photo of a yellow-brown magnesium chloride evaporation pool at South Bay Salt Works SBSW during 2019 OAST field work.

Looking at life in some of the most extreme conditions on Earth, Oceans Across Space and Time graduate student Luke Fisher of Scripps Institution of Oceanography explores how microorganisms function in exotic environments on our own planet. This kind of research is key to helping scientists develop methods to search for life on other worlds.

In the February 2021 paper in Environmental Microbiology, “Current state of athalassohaline deep-sea hypersaline anoxic basin research-recommendations for future work and relevance to astrobiology”, lead author Fisher and a team of OAST investigators explore polyextremophiles – which are microbial communities that are adapted to multiple extremes (pressure, anoxia, salt stress, desiccation) – and how scientists can better understand the limits of life as we know it.

Polyextremophiles are found in places like the Kryos, Discovery, and Hephaestus basins in the Eastern Mediterranean Sea on Earth – these areas are known as deep-sea hypersaline anoxic basins (DHABs). Like the famous Dead Sea and the Great Salt Lake, salt deposits became concentrated in these areas when now-extinct oceans or seas evaporated. Later, exposure of these ancient evaporites, salts and minerals, dissolve; creating an extremely dense brine that collects in the deep-sea basins. Due to the lack of mixing, the brine quickly becomes anoxic.. DHABs are unique because they present a combination of stressors for life that aren’t found elsewhere: osmotic stress, anoxia, elevated hydrostatic pressure and low water activity.

“Too much salt causes cells to dehydrate and die, as osmotic pressure moves water from the cell to the salty environment ,” Fisher explained.  This means that in dense brines like DHABs, very little water is available for life to use. “A lack of available water affects all the machinery of the cell, proteins, DNA replication, etc. All life needs water,” said Fisher.

The concentration of salts increases as depth increases in DHABs, and so do other stressors on microbial life, such as higher pressure and less available oxygen. It’s this combination of stressors that make DHABs one of the most extreme places we can find life on Earth.

At a certain point, the stressors become too much for microbial life, and not even polyextremophiles can survive. This cutoff point has been studied in places like the Kryos, Discovery, and Hephaestus basins, but Fisher points out in the paper that some inconsistencies in measurements need to be corrected in future work, in order to clarify the limits for life in DHABs.

Photo of a yellow-green evaporation pond at South Bay Salt Works in southern California is tinted yellow-green because of the high concentration of MgCl2, with white NaCl crystal structures visible beneath the surface and protruding above the water.
An evaporation pond at South Bay Salt Works in southern California is tinted yellow-green because of the high concentration of MgCl2, with white NaCl crystal structures visible beneath the surface and protruding above the water.

On the other side of the world, South Bay Salt Works in southern California is a company that harvests salt by allowing the sun to evaporate water from a series of ponds, leaving concentrated salt brine behind. By investigating salt-loving microbes from these ponds, especially ponds dominated by MgCl2, Fisher’s work with the OAST team is improving our understanding of how microbes can adapt and survive in extreme environments like DHABs.

“I find it interesting to work in these places, especially the MgCl2 brines, because they are much more rare,” Fisher said. “There might be life living here – and if they are, they likely have some crazy adaptations thereby changing our perception of life and what is considered biologically habitable.”

One way Fisher is using the SBSW pond samples to study extreme life on Earth, is by culturing salt-loving microbes in the absence of oxygen – one of the many stressors present in DHABs.

“Anaerobic microbes, meaning microbes that don’t use oxygen, are generally found in deeper sediment mud (usually black mud, black coloration generally means anoxia as chemical reactions in the absence of oxygen turn sediments black). I collected this type of mud at SBSW to try to culture methanogens, which are microbes that produce methane. We are interested in culturing halophilic methanogens – salt loving methane producing microbes – because they are relevant in DHAB environments. Being able to collect and grow them in-lab means we can learn more about how they survive and adapt,” Fisher said.

Not only does Fisher’s research improve our understanding of polyextremophiles on Earth, it could also further our understanding on where life might exist elsewhere in the universe.

“These sites are interesting because they offer windows to extraterrestrial environments (Martian brines for example) that teach us more about what these extraterrestrial environments might be like,” Fisher said.

Several moons that orbit Saturn and Jupiter are thought to harbor oceans beneath their icy surfaces. The presence of highly concentrated salts like magnesium and sulfur-rich compounds on Europa (one of Jupiter’s moons) means that understanding these exotic brines – like those dominated by MgCl2 at SBSW and in deep ocean basins – may be a crucial step in the search for life on other oceans across space and time.  Moreover, evaporite basins exist on present day Mars, such that work in DHABs and SBSW can help reveal what happened to these environments—and any microbes that may have once lived there–as Mars lost its water.

Lethal MgCl2 brines could be a great place to search for evidence of life on other worlds

Photo of a yellow-green lake of magnesium chloride, with a small dirt landmass on the right with several people standing on it, wearing orange vests and taking samples.

In a new paper, “Microbial diversity and activity in Southern California salterns and bitterns: analogues for remnant Ocean Worlds”, lead author Benjamin Klempay and a team of investigators from the Oceans Across Space and Time project are seeking to improve our understanding of the limits of life in brines and how that could guide our search for life on other habitable worlds.

Published in Environmental Microbiology in February 2021, Klempay and the team explained how lethal MgCl2 brines could, counterintuitively, be great places to look for signs of life on other planets. Klempay, a graduate student at Scripps Institution of Oceanography working towards his Ph.D. in biological oceanography, studies how microbial communities respond to the intense physiological stresses of living in extremely salty environments.

“Chaotropic stress (the same feature that makes these brines uninhabitable in the first place) preserves DNA, making these brines repositories for genetic fragments from surrounding environments,” Klempay said of the MgCl2 brines.  This is one of the key reasons that OAST is so interested in hypersaline envrionments.

Ponds of blue (seawater dominated by NaCl salt), white, pink, yellow, and green (lethal MgCl2-rich brines) are also the subject of Klempay’s research at South Bay Salt Works in southern California. SBSW, like other solar salt harvesting facilities, relies on evaporation to concentrate the salts found naturally in seawater, in order to harvest them.

Fig 1. An aerial map of South Bay Salt Works. Inlay shows the position of our study site within SBSW, located at the South end of the San Diego Bay, Chula Vista,CA. Satellite imagery from Google Earth. [Color figure can be viewed]
Fig 1. Map of SBSW. Inlay shows the position of our study site within SBSW, located at the South end of the San Diego Bay, Chula Vista,CA. Satellite imagery from Google Earth. [Color figure can be viewed]

“Being involved with OAST and working locally in the San Diego area gave me the amazing opportunity to advance my research with several field excursions to South Bay Salt Works,” Klempay said. “The chemical conditions in these brines changes dramatically over the course of this evaporation process, mirroring the changes that would have occurred on ancient Mars as the planet transitioned from wet to dry.”

Because the ponds span the chemical gradient from life-filled seawater to lethal MgCl2-saturated brines, Klempay and the OAST team could sample from various ponds at any given time.

“This makes SBSW a very valuable field site and Mars analogue, because it allows us to study the entire process of evapoconcentration and desiccation,” he said. “As evaporation removes water from the brines, the dissolved salts become increasingly concentrated. Table salt (NaCl) is the most abundant salt in seawater, but there are other salt species as well, which have different chemical properties. Eventually, NaCl becomes supersaturated and begins to form crystals, which fall out of the solution (and later get harvested for us to eat!). After NaCl, the next most abundant salt in seawater is MgCl2. It is more soluble than NaCl, and it stays dissolved in the brine even after all the NaCl has precipitated out. The result is highly chaotropic, MgCl2-rich brines.”

Kemplay and the OAST team used filters with tiny microscopic pores to capture all of the microorganisms living in the brines (or dead and preserved) at SBSW. Then, they took the samples back to their lab and extracted DNA from them, which allowed them to determine what was living in each of the ponds.

“We used this to form a picture of how the microbial community evolved over the course of evaporation, and at what point the limit of life was reached,” Kemplay said.

Up until a certain point, life can still exist even as the amount of dissolved salts in the seawater increases through evaporation. This life is characterized by the proliferation of highly specialized halophilic microorganisms. But at a certain point, the brines become lethal and all that’s left are the remnants of what used to live there.

But these hypersaline environments may be the perfect place to search for evidence of past life.

“Within our solar system and perhaps beyond, salty environments are among the most promising targets in the search for extraterrestrial life. Not only are hypersaline brines quite good at preserving remnants of past life, but they are also among the most likely homes for current life in the solar system,” Kemplay said.

In places like Mars, where the planet experienced a shift from being a wet world to a dry one, hypersaline brines would have been the last places for life to find liquid water.

“Liquid water is a fundamental requirement of life on Earth, so a leading strategy in the search of extraterrestrial life is to follow the water,” he said. “As for current ocean worlds, the most promising candidates in our solar system are moons of Jupiter and Saturn, which are too far from the Sun for pure water to stay liquid. But salt lowers the freezing point of water, allowing these moons to have vast briny oceans beneath their icy shells.”

By understanding the limits of life in hypersaline brines on Earth, and how evidence of past life can be preserved in these environments, we may be one step closer in our search for life (or the remnants of it) on other worlds.

The paper Klempay was lead author on made a splash in the world of hypersaline brines, but the work he and the OAST team are doing is ongoing at SBSW.

“We went back to SBSW in 2020 to conduct new experiments and to repeat some of the same experiments with new and improved methods. We’re now able to determine much more effectively which cells are actually alive and reproducing versus which ones are dead/inert. I am also zooming in on the proliferation of halophilic microbes in order to understand how these incredible organisms are able to evolve and adapt so quickly to the rapidly changing conditions in evaporative brines,” Kemplay said.

Video: South Bay Salt Works

During our annual meeting in 2019, Oceans Across Space and Time (OAST) team members conducted field work at South Bay Salt Works in Chula Vista, California. We collected samples to study the contents and life in different salt ponds at SBSW, and learn about life detection techniques that may help the search for life on other worlds.

South Bay Salt Works: 2019 Field Work

A dirt path separates two salt ponds on either side. On the left, a deep pink-red pond with still water, and on the right, a white-pink lake with still water.

The August 2019 team meeting in San Diego, CA was a great success, with collaboration, planning, and field work the focus of an extremely successful week. Scientists, engineers, and other OAST (Oceans Across Space and Time) team members met for productive meetings and planning sessions, before heading out for field work in two different locations: South Bay Salt Works in Chula Vista, and the ocean off the coast of San Diego near Scripps Institute of Oceanography.

Life detection methods took center stage at SBSW, while another small team took the Icefin robot out into the ocean for test runs. Icefin, a robot oceanographer designed and operated by several team members, is technology used both OAST as well as several other projects, including MELT and RISE UP.

Over at the salt works, a group of 16 OAST members collected samples of the salts, water, and sediment, in several salt ponds, in order to study how microbial ecosystems are organized across gradients of environmental stress, with the goal of aiding life detection methods on other worlds, such as Mars and Europa.

Several days at the salt works yielded exciting samples for the team to analyze and study over the upcoming year. The salt ponds ranged from approximately the salinity of the ocean, all the way up to the point where certain salts reach saturation and start to precipitate out, resulting in colors that vary from blue, to white, to pink, to yellow and green.

Some ponds – whose salinities similar to ocean water – are dominated by sodium chloride. When that sodium chloride precipitates out, it leaves other salts like magnesium chloride, which interacts with the ecosystem, and life, in a very different way than the sodium chloride in Earth’s oceans.

The intensive pink color of the sodium chloride ponds reflects the density of life in those environments. By contrast the green coloration of the magnesium chloride ponds is caused by highly toxic minerals. Trying to demonstrate the presence or absence of life in these ponds is very similar to the challenge of life detection on other planets.

“These ponds will break any instrument that you put in them” said Jeff Bowman, a biological oceanographer at Scripps Institution of Oceanography and OAST deputy PI. “But that’s what makes them such a good model.  If we can make an assay or experiment work here, we can make it work anywhere.”

In addition to identifying the different salts present in each of the ponds they sampled, team members are also working to identify the specialized life that can survive in sodium chloride, and determine if life can even survive in high concentrations of magnesium chloride at all.

Because of the range of salinities and conditions from pond to pond, some where we know life can exist, and others where we’re unsure, South Bay Salt Works is a great place to test life detection strategies for places like Mars and Europa.