Interview with Dr. Jocelyne DiRuggiero

As mentioned on the email list, this year we’re attempting to do a little astrobiology public outreach and education.  Our first series of efforts are interviews with faculty and researchers here at Johns Hopkins who are actively involved in astrobiology-related research.  These interviews will be featured on the once-a-day podcast site, 365 Days Of Astronomy.  We’ve done two interviews so far, and the first one goes live on that site tomorrow.  It’s an abridged, 12-minute version of our discussion with Dr. Jocelyne DiRuggiero of the Biology department about her research with halobacterium and hyperthermophiles, extremophiles that live in high salt and high temperature regions, respectively.  Below I’m attaching the full, 24-minute interview as well as the transcript.

Let us know, either through the comments section below or on the email list, if you’re interested in helping out by suggesting someone to interview, being interviewed yourself, or anything else you’re interested in trying (it’d be nice if we had a theme song….).  Our next interview is with Dr. Naomi Levin of the Earth & Planetary Science department.  It should be going up in the next month and will be featured on 365 Days Of Astronomy in April.

Now, on with the podcast!

Studying extremophiles on Earth to understand life in space

With the Kepler Mission’s discovery of 4 potential Earth-sized planets orbiting in their host star’s habitability zones, the main question about life is no longer “Is there life out there somewhere?”  Instead we must ask, “Exactly what sort of life could exist on these strange planets?”  For today’s 365 Days Of Astronomy podcast, the JHU Astrobiology Forum’s Adam Fuller begin answering this question by speaking with Dr. Jocelyne DiRuggiero, an associate research professor in the Biology department at Johns Hopkins University, about her research with microorganisms here on Earth that live in environments so hellacious, they could easily be thought to be from another world.

Full Transcript

JDR: I am Jocelyne DiRuggiero.  I am a professor in the Department of Biology at Johns Hopkins University.  My research focuses on extremophiles.  Extremophiles are microorganisms that live in places that we would feel very uncomfortable in.  So, for example, a microorganism living in a deep sea hydrothermal vent, where the temperature is very, very high.  Or they’re living in high salt environments—we’re talking about saturating salts such as the Great Salt Lake or the Dead Sea.  Organisms that can live in acids, basically, very, very low pH. And also organisms that live in very dry environments.  We know that water is essential for life, and when there is no water, it makes it’s really, really difficult for microorganisms to live.

AF: Now, your research specifically focuses on what aspect of all that?

JDR: So we’re interested—really, the lab has two main topics.  One of them has to do with the way those organisms respond to environmental stresses….So we’re interested in how these organisms respond to environmental stresses, how they can protect the micro molecules in the cells—in particular the genetic material which is essential for the cells to survive and multiply.  We’re looking at extremophiles because they have very robust adaptations to extreme conditions, so this is a place where we can really dissect those processes.  We’re particularly interested in DNA repair mechanisms but also stress response in general to the environment.

AF: What sort of extremophiles do you work with?

JDR: So right now in the lab we have two types of extremophiles.  Some are called hyperthermophiles.  They grow above 90°C.  They typically grow in the absence of oxygen, and sometimes we need to add elemental sulfur so they don’t respirate oxygen.  They produce found something that is called hydrogen sulfide, which this very smelly gas.  So that’s one group of organisms.  And we’re looking at particular DNA repair proteins in these organisms.  The other group of organisms we’re using are halophiles, and those grow in high salt conditions, so we have to put 4 molar salt, which is about 20% salt in the culture medium for them to grow.  If we add some water it basically lyses the cell; they just pop open.  They really cannot survive without very high salt.

AF: Okay, so even if—oh, that’s interesting—so the water, it has to have a very high salt content.  Normally, how do cells react in, like, say, like a normal bacterium—

JDR: So, if you put a normal bacterium in very high salt, you have very high osmotic pressure—what’s called osmotic pressure is the pressure outside the cells due to the salt.  What this does is it draws out the water from the cell into the medium, so the cells shrivel and dry.  Those organisms resist a very high salt because they also accumulate very high salt inside the cells; they balance the very high osmotic pressure outside the cells by accumulating very high salt.  Now, what is very interesting is that the salt in the environment is sodium chloride, the same salt we use for our table salt, but inside they accumulate potassium chloride but to the same level.  This means that all the micro molecules of the cells have to be adapted to function in very high salt, and usually if you put a protein in very high salt, it precipitates.  So we have an adaptation: the proteins that allow those proteins to stay in the soluble form at very high salt and function perfectly well in this environment.

AF: What sort of environments on Earth, what sort of places would you find extremophiles like these?

JDR: As I mentioned earlier, the Great Salt Lake.  That’s where the organism we’re using most in the lab, called Halobacterium salinarum, and has been isolated.  But you can also find them in the Dead Sea—any places with very high salt.

AF: Could someone just go and, with a cup, just skim water off, and have these extremophiles, or would you have to dive deeper into the water to find these?

JDR: No, you just scoop water out of the Great Salt Lake, and if you can make up a medium that has a high salt, you can grow the organism.  So they have this very nice color that is pink, and sometimes you can find this—for example, in salt urns, which are places where they make salt, sea salt.  You get salt from the ocean, you let the water evaporate in very shallow ponds, and then the salt concentration increases.  Then, depending on the concentration of salt in the pond, you’re going to see different colors.  The higher salt concentrations have a pink color, which is due to the presence of Halobacterium.  So this organism is so good it can even survive inside the salt crystals.  So you have pink salt crystals.  If you put them in a culture media they’ll start to grow again.

AF: Now, what about the other extremophiles you mentioned, the—

JDR: So the high-temperature hyperthermophiles.  Those are found in deep sea vents, deep-sea volcanoes.  They’re also found in places like Yellowstone National Park.  Every time you have a very high temperature in an aquatic environment, you’re going to find those microorganisms.  There are a few places on the earth and also a few places where we can find them in shallow reefs.  For example, there is an island in Italy called Volcano Island, and right off the beach you can find some of those very hot small vents and you can isolate those microorganisms from there.

AF: So, as far as how those organisms came to be, are these, would you say that these are evolutionary adaptations these organisms have gone through to adapt to their environment?  Or are these reflective of some prehistoric condition that hasn’t changed for these organisms as they’ve stated in one spot?

JDR: This is actually a very good question, and it’s really comes back to the debate about the origin of life.  There are several theories about this, but one that is quite predominant says that the first type of organisms were hyperthermophiles, because they evolved very well protected at the bottom of the ocean, so they were high-temperature organisms.  And it makes sense in many ways because if you look at the tree of life, which is this tree that represents all the life on Earth and is based on molecular data, all the organisms that are close to the root of the tree, the deepest branches of the tree are occupied by hyperthermophiles.  This brought the idea that early organisms were hyperthermophiles and from that evolved the rest of the organisms.  The other argument in favor of this theory is that the Earth at the time was heavily bombarded; it was very unstable at the surface.  If you can imagine that if you have a very deep environment, that might be a good place for life to evolve.  But there are quite a few problems with this.  One of them is that if you look at the adaptation that hyperthermophiles have evolved, they’re actually very sophisticated.  For example, the membrane is quite different than all the other membranes of organisms we can find on Earth.  They also have enzymes that are very specific to those organisms and also very sophisticated.  This really contradicts the idea that those features were primitive features and actually support the idea that those evolved afterwards.  We had very basic simple cells, and then as life multiplied and diversified, you had colonization of those niches with organisms that evolved adaptations to specifically survive in those environments.

AF: Can you tell me about some sort of experiment you’re working on right now in your lab?

JDR: We’re interested in this halophile that I was talking about, Halobacterium.  In particular we want to understand how the organisms can go through periods of desiccation when the water evaporates and the organisms are trapped in the salt crystals, so there’s very little water.  And when water comes about, they survive—they survive very well in the salt crystal, and then they start going again.  So, what are the metabolic changes that they undergo for those periods of desiccation, how do they resist desiccation?  To do that, we don’t desiccate the cells.  We actually irradiate them with ionizing radiation.

AF: You irradiate them?

JDR: We irradiate the cells with Cobalt-60, which is a proxy for desiccation.  It’s much more reproducible, and we can very accurately dose the level of radiation.  When you try to desiccate a culture, depending on the temperature outside—if it’s Baltimore in the summer, it’s still going to be very humid, so it’ll be very difficult to control.  So, because there’s a very strong relationship between resistance to desiccation and resistance to radiation, and we know that the cellular damage in both conditions are very similar, we use radiation as a proxy.  It’s much easier to do accurate dosing in a reproducible experiment.  So we’re looking at how the cells are resisting to the irradiation and similarly to desiccation what we found is what desiccation and irradiation does to the cells is basically a very big oxidative stress.  It oxidizes a lot of the macromolecules in the cells.  So we’re trying to understand what is the difference between Halobacterium that can resist very well for irradiation and desiccation and organisms like Pseudomonas that are very sensitive to both.  So what is different in the cells that make one organism survive perfectly well and the other one basically dies.  What we found is that all the enzymes that are typically present in the cells to fight the oxidative stress are not the most essential things in the cell.  There are actually very small molecules that are able to scavenge, to basically take out those free radicals and then protect the cells.  So the cell doesn’t get as much damage because it’s able to basically knock down all those reactive oxygen species before they cause too much damage, in particular to the proteins.  We’re looking more in particular at the molecular pathways that control the production of small molecules.

AF: What sort of environments…to take this back out to a more broader, astrobiological sense, what sort of environments would you—not to say they are there—but if you were to find a certain environment, what environment would you say would be conducive to hosting extremophiles like these?  For example, Titan’s surface, we have a somewhat clear, somewhat foggy idea of what’s on the surface….

JDR: What other places…how are those halophiles and hyperthermophiles relevant…all right.  So if we look at hyperthermophiles, we’re talking about hydrothermal vents, we’re talking about a hydrothermal source deep in a planet.  You could think of places even like Europa that might have a liquid ocean that could be the result of hydrothermal activity.

AF: I think it’s with Europa where we’re fairly certain there’s a huge underground saline ocean. So, halophiles, I would imagine…

JDR: It depends on salinity.  The ocean is only a few percent salt, sodium chloride, and we’re talking about ten times more for this organism.  If you have very high salt, you may find those organisms that have adapted to very high salt.  But I’m thinking also in terms of energy sources.  The hydrothermal vent can produce the heat but also minerals and things like sodium sulfide that could—also, hydrogen—that could be used by microorganisms as an energy source.  So hyperthermophiles could be found in places where you have a hydrothermal activity.  Now, halophiles can be found in many other places, in particular, thinking about Mars and the idea that at some places on Mars you have salt flats.  Maybe not now but at some point in the history of Mars those might have been warm enough that you had enough water and then halophiles could be able to survive in those conditions.  If they’re not there now, they could have been at some point in time because those are very high salt environments.  The other thing that we’re looking at with the halophile but also with other organisms is desiccation, and we’re doing quite a bit of environmental work in deserts.  We have a site in the Atacama Desert in northern Chile, and we’re also getting samples from collaborators from the Dry Valley of Antarctica, in particular, in the upper valleys, which are extremely dry.  This is very cold and very dry.  So we have a desert that is fairly temperate, the Atacama Desert, extremely dry.  And then a desert that is extremely dry but very cold, which is in the Dry Valley of Antarctica.  And what we’re doing in those environments is looking at what is the diversity of the microbial population we find.  What can we find in the soils?  Is there anything alive?  What are the different types of microorganisms?  Are they living, metabolizing very slowly?  Are they just sitting there because they happen to be deposited there?  Or are they sitting there waiting for next rain that might occur?  We’re using a lot of molecular methods to look at those, and what we’ve found so far is that there is not that much difference between the diversity of the organisms we found in the Atacama Desert and Antarctica, although we have a little more diversity in Antarctica.  That leads us to think that maybe most of those organisms are just dormant, sitting there and not metabolizing much.  But this is very important if we think in terms of astrobiology in places like Mars where there is almost no water.  So what type of organisms could we find in the soil?  In what conditions might they be?  Is there any metabolic activity at all?  By looking at a very dry environment on Earth, we can get insight in what we might or might not find in other places that are very dry.  This is more an environmental work, but it’s really merged together with our interest in desiccation and the halophiles.  We can find the halophiles, but, of course, at some point, to get the halophiles to grow you need fairly high salt.

AF: When you’re looking at extremophiles, how they’ve adapted to their local environment doesn’t really tell us much about the global environment.

JDR: What do you mean, the global environment?

AF: Extremophiles living on a thermal vent versus 30 degrees outside and snowing.  I guess the last question I would want to ask is as far as how these extremophiles would evolve over time, would there need to be a change in the environment to spur genetic diversification of these extremophiles, or, if, and if that is the case, if we were to find extremophiles living in, say, Mars’s soil or a few feet underground, would we expect to find that same type of extremophile over the entire planet…I guess, how does a changing environment change….

JDR: So micro organisms….Well, microorganisms are absolutely fantastic.  They can adapt to many different conditions and changing conditions in the environment.  Even within a population of microorganisms that live in, say, in a hydrothermal vent, you do have multiple variants and you have genetic changes happening all the time in these populations, random mutations that may have consequences on specific pathways of the metabolism or the way they use a specific energy source.  So they’re constantly evolving all the time, and you have all those multiple variants in one population.  If the conditions change to some extent, the dominant organisms might not be dominant anymore, but another part of the population that is better adapted will rise and then colonize this whole environment.  What I’m saying is that microorganisms have a tremendous capacity to adapt.  If there’s another niche that opens—for example, suddenly there’s an import of new carbon sources, you will find organisms that will be able to use those carbon sources and then colonize this niche.  And it’s true for higher temperature and lower temperature—although we think there is a limit for the biochemistry that we know as far as temperature.  But organisms in a population of hyperthermophiles are not all the same, and some might be able to grow at a lower temperature.  So let’s say your thermal source decreases in temperature.  Then you’re not going to have the same dominant organisms that you’re going to have otherwise.

AF: Right.  I guess what I was thinking of was Europa—or is it Enceladus…one of those two—shoot, now I’m embarrassed—one of those two has the geothermal vents, but the rate at which they’re spewing out mass leads us to think that this isn’t a steady state for the object because over time, at the rate spewing mass, it would exhaust all its mass in just a few million years—

JDR: Right, so you think it’s intermittent—

AF: So, yeah, the geysers are intermittent—

JDR: Are you talking about the ones at the surface, those big geysers at the surface?

AF: Yeah, the one where they’re at the South Pole.

JDR: That’s Enceladus.

AF: So, the question would have been, do we have examples of extremophiles where we’ve taken them into the lab or changed their environment, and they’ve adapted, or…

JDR: Yeah, all the time.  You can do that very easily with microorganisms.  I’ll give you an example.  The organism we’re working on at the moment is Halobacterium.  They’re fairly resistant to radiation.  We measure the resistance to radiation as the D10, which corresponds to the radiation doses for which 10% of a population survive.  So the D10 of the organism, that is called the wild type.  The regular organism is five kilo Gray—that’s measured radioactivity—which is pretty high.  This is 5000 Gray, and humans are killed by five Gray.  Those survive 5000 Gray humans died with five Grays.  But it’s okay, 5000 Gray is pretty good.  But what we did was we took those organisms and then we exposed them to nine rounds of very, very high levels of radiation.  We exposed them, came back to the lab, grew the survivors, exposed them, came back to the lab, grow the survivors.  We did that nine times.  And at the end of those cycles of radiation, we raised the D10 of the organisms to 16000 Gray.  So we increased the radiation resistance by an enormous amount.  We went from 5000 to 16000, which is a lot, just by subjecting them to a selective pressure.  So you can do that all the time with microorganisms.  The questions with those intermittent geysers are different in the sense that you would have a heat source but also a nutrient source for a while.  Then you won’t have anything very long time.  And then something is happening again.  Well, there are microorganisms that can survive several million years in salt crystals.  That’s work from Russell Freeland who revived microorganisms had been trapped in salt crystals from a salt mine and that were several million years old.  So microorganisms can survive for a long time being dormant.  And when the right conditions happen again, they can grow again.  So they are tremendously good at adapting to different environmental conditions.  They’re also very good at surviving for very long periods in the dormant state.  I mean, spores of Bacillus, for example—we all know about anthrax—those spores are very resistant and can resist heat, acid, lots of different thing.  You can imagine sporulating microorganism, with spores sitting in the environment for millions of years.