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STUDYING LIFE at the EXTREMES Researchers Have Invented Methods to Study Microbes That Thrive in the World’S Most Inhospitable Environments

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STUDYING LIFE at the EXTREMES Researchers Have Invented Methods to Study Microbes That Thrive in the World’S Most Inhospitable Environments Work / Technology & tools STEFAN GREEN/XMP STEFAN Researchers are looking for microbes in soil samples from this flaming gas crater in Turkmenistan, known as the Door to Hell. STUDYING LIFE AT THE EXTREMES Researchers have invented methods to study microbes that thrive in the world’s most inhospitable environments. By Amber Dance icrobes cling to life in some of Earth’s identify, culture, genetically manipulate and and whether they might make compounds that most extreme environments, from observe extremophiles, researchers often could work as antibiotics. toxic hot springs to high-altitude tweak the methods used in more run-of-the- In their search for new microbes to analyse, deserts. These ‘extremophiles’ mill organisms. Whereas some techniques can the scientists have travelled to extreme locales, include organisms that can survive be easily transferred — from one thermophile including the toxic hot springs of Ethiopia’s Mnear-boiling heat or near-freezing cold, high to other heat-lovers, say — others have to be Danakil Depression, which are loaded with pressure or high salt, as well as environments adapted for each new organism. salt, acids and heavy metals, and a flaming steeped in acids, alkalis, metals or radioactivity. “Each extremophile is going to have its own gas crater in Turkmenistan. Coercing these organisms to live in labora- set of challenges,” says Jocelyne DiRuggiero, But their main challenges arose back in the tories creates many challenges. Nonetheless, a microbiologist at Johns Hopkins University lab: “Massive hurdles, actually,” says Tighe. papers published on extremophiles have dou- in Baltimore, Maryland. “Every time you think Microbes tough enough to withstand extreme bled in the past decade. Some scientists are about doing something, you have to think, how conditions resisted scientists’ attempts to break drawn to the novelty of the organisms, search- am I going to adapt this for my organism?” them open and recover their DNA. So Tighe ing for ones that are undescribed or that might and the XMP team developed a six-enzyme harbour useful enzymes for industrial processes Growing at extremes cocktail — now commercially available as or antibiotics to save lives. Others simply find In 2014, Scott Tighe, a microbiologist at the MetaPolyzyme — to break down any cell sur- that the best organism for their scientific ques- University of Vermont in Burlington, gathered face they might come across, adding detergents tions happens to have extreme preferences. international collaborators for an ongoing ini- and organic solvents to collect nucleic acid on It’s a circumstance that has forced research- tiative known as the Extreme Microbiome Pro- magnetic beads1. “It got to be a fairly exotic ers who study extremophiles to invent new ject (XMP). The researchers hope to find genes DNA-extraction technique,” says Tighe. Thus laboratory methods for handling them. To that might indicate how extremophiles survive armed, the researchers are working through a Nature | Vol 587 | 5 November 2020 | 165 ©2020 Spri nger Nature Li mited. All rights reserved. Work / Technology & tools freezer full of samples, says Tighe. borrowed a gene-swapping system from another it would be so squishable,” says Eun, now a Studying living extremophiles in the lab thermophile to introduce the desired sequence principal data scientist at Janssen in Titusville, creates challenges, too. DiRuggiero’s group, into C. thermocellum. At the same time, they New Jersey. A microfluidics device called the for example, works with salt-loving halophiles, changed nearby sites that normally allow a CellASIC ONIX, from Millipore Sigma in Burl- chipped out of rocks from Chile’s high-altitude CRISPR–Cas system to recognize and cut the ington, Massachusetts, pins microbes using Atacama Desert. Some such organisms are DNA, rendering them invisible to CRISPR. Then, soft polydimethylsiloxane, but the material easy to cultivate, she says: just add salt. they applied CRISPR–Cas to slice any unmodi- proved toxic to Eun’s cells. DiRuggiero has also worked with anaerobic fied genes, eliminating unedited microbes. She finally succeeded by fabricating tiny hyperthermophiles, which grow at high heat agarose chambers to confine the cells gently. and in the absence of oxygen. To culture them, “The main thing is, just try. At last, Eun could see that the archaeon main- she used heat-resistant, NASA-made culture tains its size in much the same way as bacteria jars in an incubator set to 95 °C — so toasty, the The ‘extremophily’ is often do, with each newborn cell adding a consistent team jokingly called it the “pizza oven”. Stand- not as much of a barrier as length to its rod before dividing again6. ard agar in plastic Petri dishes would melt, so you first might think.” scientists turn to glass dishes packed with a Get your glow on gellan-gum derivative called Gelrite, which Microscopists must also find ways to label can withstand the heat. The strategy has up to 94% efficiency, and proteins of interest under extreme conditions. should work for other thermophiles, too, says For example, an international team faced Manipulating genes Eckert. The necessary microbial strains are a challenge with a halophile called Haloferax Altering extremophiles’ genes can require available from New England Biolabs and the volcanii, found in the Dead Sea, because it extra effort. The tools and techniques that American Type Culture Collection, and Eckert makes a pigment that naturally fluoresces. work in lab favourites such as Escherichia coli is working to add the plasmids to the Addgene That made it tricky to use fluorescent protein — including the plasmids that are used to trans- repository. tags for tracking single molecules, the group fer genetic material, methods to insert that reported in a preprint in July7. So, first, the team material into new microbes and compounds Live on camera deleted a gene involved in synthesizing that pig- to select microorganisms that have success- Microscopes are also getting a makeover to ment, creating colourless but otherwise normal fully integrated new genes — are frequently investigate extremophile cell biology. microbes. ill-suited to high levels of salt, roasting tem- Buzz Baum, a cell biologist at the Medical Then, the researchers tackled the genetic peratures and other extreme environments. Research Council Laboratory for Molecular milieu of H. volcanii. Like many halophiles, Scientists often create the genes they want Biology in Cambridge, UK, is interested in cell H. volcanii has DNA containing a relatively to use in E. coli, isolate the relevant plasmids, division in the thermophile and acidophile high proportion of guanine and cytosine and then transfer them into the extremophiles. Sulfolobus acidocaldarius. But live cultures bases, so its codons — the triplet sequences Some extremophiles can take up nucleic acids cool as quickly as a cup of tea on the micro- that encode amino acids — also tend to use from the surrounding media, says Carrie scope stage, says Baum, and the microbes go those bases. Furthermore, genes that use Eckert, a synthetic biologist at the University into suspended animation. “We spent many the same codons will be expressed at higher of Colorado Boulder and the National Renew- years failing.” levels. So the team optimized the codons able Energy Laboratory in Golden, Colorado. The team decided to borrow a technique used for fluorescent proteins that work But if that doesn’t work, she recommends used in polymerase chain reaction machines, in non-extreme organisms to match the electroporation, a pulse of electricity to open to warm the cultures from the top as well as the extremophile’s preferences. up holes in the cells’ membranes. bottom. They recruited a lab member’s father “Most of them worked,” says co-author Iain Another tip, she says, is to determine the and brother, both aerospace engineers, to fab- Duggin, a molecular microbiologist at the methylation patterns on the extremophile’s ricate a chamber out of aircraft aluminium. University of Technology Sydney in Australia. genome, through sequencing2. That’s impor- The researchers have published diagrams Versions of the crimson-coloured protein tant because microorganisms will often for this ‘Sulfoscope’5, but any double-heated mCherry and the green-to-red photoswitch- destroy nucleic acids that have the ‘wrong’ chamber should do, says Baum. The system able tag Dendra2 were especially useful for pattern, to protect themselves from invaders. allows the team to investigate proteins that single-molecule tracking. “We’re really excited Scientists are learning to modify those methyl- maintain symmetrical cell division. about how this can be applied to studies of cell ation systems in E. coli to make new DNA match Halophiles, too, are tricky to work with division and shape and the cytoskeleton,” says that of the host3. under the microscope. They lack the rigid Duggin. Once they’ve provided correctly methylated cell walls found in bacteria, and survive by Indeed, with perseverance, scientists DNA, scientists must pick out the extremo- maintaining the same osmotic pressure as interested in extremophiles can make impos- philes that take it up. To investigate the genes their environment, giving them all the rigid- sible-seeming experiments work. “The main they’re interested in, microbiologists often ity of a limp balloon. That means they’re easily thing is, just try,” says Duggin. “The ‘extremo- insert them into plasmids that contain genes deformed when sandwiched between a micro- phily’, if you will, is often not as much of a for antibiotic resistance; colonies that grow scope slide and coverslip, making it difficult to barrier as you first might think.” on media containing that antibiotic have taken study the size and shape of the cells. up the genes. But many antibiotics fail to func- Ye-Jin Eun encountered this problem during Amber Dance is a freelance science journalist tion under extreme conditions. An alternative a postdoc at Harvard University in Cambridge, in Los Angeles, California.
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