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Down the Rabbit Hole: Ancient Environmental DNA – Benjamin Vernot, Max Planck Institute of Evolutionary Anthropology

Benjamin Vernot is Group Leader at the Max Planck Institute of Evolutionary Anthropology, Germany. His group works in the field of ancient environmental genomics, with a particular focus on studying ancient DNA extracted from sediments. In fact, this work was recently cited in the decision-making process for the 2022 Nobel prize in Physiology and Medicine, which was awarded to the department’s director, Svante Pääbo.

Please note the transcript has been edited for brevity and clarity.

FLG: Hello, and welcome to the latest “Down the Rabbit Hole” interview. Today I’m joined by Benjamin Vernot, who is going to tell us all about the fascinating field of ancient environmental genomics. So, Benjamin, could you tell us a little about yourself, and perhaps give us an overview of your work?

Benjamin: Yeah, sure. Thank you very much for having me. I’m a scientist here at the Max Planck Institute for Evolutionary Anthropology, and I lead a group studying ancient environmental genomics. I got my start in this field studying Neanderthal DNA – looking at the interactions between humans and Neanderthals. So, I have a long history in ancient DNA. But these days, we’re looking at DNA in sediments.

FLG: That’s great. So, I guess let’s start with the basics – what is ancient environmental genomics?

Benjamin: The idea behind ancient environmental genomics is that we leave DNA everywhere we go – that includes us and other animals. Typically, for ancient DNA practices, what we do is we find a bone or a tooth, we drill into it, and we get the DNA out in that way. Using this method, we can study a single person or an animal. But we don’t have to do that. If we can get DNA from the environment, it can be beneficial for a lot of reasons. But also, in a lot of circumstances, we just don’t have those bones around. In this way, we’re developing ways to get our DNA from the places where we lived, the places where we used to be.

FLG: You touched on certain situations where this might be useful. Could you elaborate on that? Why can environmental genomics help in certain research situations?

Benjamin: A lot of my research is on Neandertals – on really old human populations. Back then, it’s not clear whether people buried their dead. If they did bury their dead, we certainly haven’t been able to find the places where they did that. This means for a lot of time periods, we just don’t have any bones. If you don’t have any bones or teeth from people, you can still see that their tools are there, you can see that they lived there, even for tens of thousands of years. But without their bones, we just can’t study them genetically. We can’t tell what population they were, who they’re related to, or how they got there, etc. If we can get their DNA from the dirt instead, then that can fill in some of those gaps. So that’s one big way in which we can use ancient environmental genomics.

FLG: Is there other information you can get from environmental DNA that you might not be able to get from bones or teeth?

Benjamin: Yeah. There are some things you can figure out. An environmental sample often has DNA from multiple individuals, so you get maybe a broader look at things. However, the amount of DNA we can get out of a sediment sample is really small, so we would always prefer to have a bone if we could. In some of our work, we’ll do these time series where we know that people (Neanderthals) have occupied a site for 50,000 years, and we can take sediment samples going all the way through those 50,000 years, representing every 1000 years. And there’s no way we can do that with bones. We just wouldn’t find the human remains sort of spaced out in that in that time series. There are a lot of things we can do with this information, but that’s one pretty obvious example.

FLG: How and why did you first get interested in environmental DNA?

Benjamin: I’ve been studying ancient DNA and ancient populations for a long time. And it essentially became clear that we were missing out on a lot of human history – that there was a whole lot of human history where we didn’t have the bones, and we weren’t able to answer the questions we wanted to answer. For example, how did Neanderthals get to this particular place? When did humans meet them? This sort of stuff. The hope was that by getting into the sediment and extracting the DNA, we could fill in these gaps. So, the original motivation was to try to fill in the gaps in the genetic archaeological record.

FLG: And then what inspired you to look at sediments specifically as a source of ancient DNA?

Benjamin: Well, there’s sediments everywhere, essentially. In these cave floors, the dirt builds up over time, and even if we look outside of caves, we have these stratigraphies pretty much everywhere – every archaeological site has sediments. If you can learn about people who lived in a place by looking at the sediments, then then you have a very abundant resource.

FLG: You mentioned the issues with small sample sizes. Is there any technology that is coming out, or that you’d be really interested to see, that could perhaps help you get more sample out of the dirt?

Benjamin: There’s technical stuff we still need to work on and make better. One of the things we do that really enables us to get the DNA out of the sediments is we “capture” it. This means we sort of fish out the DNA molecules that look like human molecules (the ones we want), and we leave the rest behind. This enables us to get human DNA from a sediment sample where it’s maybe one in a million, or one in 10 million of the molecules in the sediment sample.

There are all sorts of ways that we can make that better. A lot of times the sediment samples have these chemicals in them that inhibit the processes that we use to copy the DNA. And we’re trying to figure out ways to get around that inhibition. We would also like to target more DNA. As I said before, we only get a very tiny amount of human DNA out of each sediment sample. And we think we can get better at that. Essentially, the technology is always improving.

FLG: I bet you’ve got quite a few things on your wish list! Are there other challenges involved in this type of research?

Benjamin: Yeah, the contextualization of the data is a really big challenge. Things move around in these caves, they move around in a system – the sediment moves, we even know that bones and teeth can move through the stratigraphy. So, this has always been a challenge for ancient DNA. I think that while this is a big challenge for sediment DNA, it’s actually a place where we can help. We take a lot more samples, so the chance that in five different places in the cave, the exact same accidental movement has happened, is really low. On the other hand, if you find one tooth, maybe a mouse picked it up and carried it through the cave or something like that. And so, this is a place where our research can really help.

FLG: That’s really interesting. I hadn’t thought about that aspect of it. I wanted to ask you more about your paper, the one on the Neanderthal DNA in caves. Could you give us the top line summary of that research paper?

Benjamin: Yeah, this is a prime example of one of these places where we knew that Neanderthals had been there for, in this case, 50,000 years. This is longer than modern humans have been in Europe. These Neanderthals made tools in the cave, they left fragments of the tools around, but we didn’t have any bones that we could get ancient DNA from, so we had no way to study them. Also, during this time period in Spain, there also weren’t any other Neanderthal bones from the area that we could study.

Therefore, our team (along with our collaborators – the archaeologists who have excavated that site for decades now) went and took sediment samples in a very detailed, centimeter by centimeter process going down through the stratigraphy. You can see the layers on the side of the cave, where they’ve been excavated, and we took the sediment samples at different points. And one of the neat things that we found – really, the primary thing that we found – is that we could get Neanderthal DNA for the whole time period. So every couple of 1000 years.

The second significant finding was that that there were two groups of Neanderthals that lived there. We found that at one point, one of those groups died out or moved away, or were pushed out or something, and a new group of Neanderthals moved in. And the new group of Neanderthals that moved in seem to be the ancestors of all the Neanderthals that spread out and populated the rest of Europe and even western parts of Asia. So, we think this was a pretty significant event. It happened around the time of some climate change events as well, and we don’t know if it’s tied to that. That is an area of future research. But these are the primary results that we got there.

FLG: Could you tell us more about how would you investigate that further?

Benjamin: Well, I think one of the things that is suggested by that result is that there was some place in Europe where Neanderthals were hanging out, were able to survive. We would call that a refugium – where they were able to survive through the glaciation that came down and covered a lot of Europe with ice. Unfortunately, we don’t know where that is, but it’s a place we want to find. We don’t know if there’s just one place or two places – it sort of looks like there’s just one place because really, all the rest of the Neanderthals that we find in Europe were descended from this group. Now, it could also be that we’ll find some other Neandertals that were not descended from the group. There’s so much that we don’t know about it. But for me, that’s the thing I would really like to figure out – where did they survive? Where was their “oasis” during the time of the glaciers?

FLG: Oh, wow. And that involves going out in the field quite a lot, I guess.

Benjamin: Yeah. I mean, the idea is to then just look at a lot of lot of caves!

FLG: What kind of things would you be looking for? What would get you excited and make you think – this might be the cave?

Benjamin: Well, I mean, I think it’s unlikely that we would find the cave, but we might find the region. And I’m sure there are things that archaeologists would know to look for. They would know to look for a particular type of tool that is found in a time period that we wouldn’t expect them to be, or something like that. That’s not my expertise. And we lean quite a bit on the archaeologists, because without that interpretation, without the context, it’s just really a pile of dirt that we’re analyzing, right? It doesn’t have all that significance without the archaeologists. But in terms of our research, what we would be looking for is DNA that is the oldest that we can find that still looks like it’s the common ancestor of all of the Neanderthals in Europe.

FLG: You mentioned there that you work with lots of different researchers from different areas. Who’s in that team when you go out to a cave to sample the dirt?

Benjamin: Some of the most important people are the geoarchaeologists. These are people who can look at the sediments, look at the stratigraphy, and tell us what’s going on. I mentioned that one of the things that we worry about is the movement of DNA through the stratigraphy. One of the tools that we have, and that our collaborators have, is we can make what’s called a micromorphology block. This is where we essentially take a block of sediment, impregnate it with resin so it’s hard as a rock (it’s essentially made from plastic) and then you can really look at the sediments. Using this method, we can see evidence of tiny particles moving through the stratigraphy, or everything might look pretty stable. We can even drill into those blocks and take micro samples and see whether the DNA in the sediment changes from one point to the next. The collaboration with geoarcheologists has been crucial in interpreting the results.

FLG: Wow. It’s amazing the high resolution that you can get by doing that. I think this is something you mention in the paper, but what are some of the problems around using mitochondrial DNA versus the benefits of using nuclear DNA instead?

Benjamin: Our field has a long history of starting with mitochondrial DNA. The first Neanderthal sequences were mitochondrial sequences. And one of the reasons for that is that every single cell in your body has tens, or hundreds of copies of the mitochondrial genome, but only two copies of the nuclear genome. And so, you know, it’s just easier to get that DNA. Nuclear DNA is also a much smaller target size, so mitochondrial DNA is often the first thing people study – but it’s not super informative for population histories, right? If we want to say that this group was replaced by this other group, or this group came from here and was related to this other group, you can’t with mitochondrial DNA, because it only carries the DNA from your maternal ancestors – your mother and your grandmother and her grandmother and her grandmother. And that leaves out a lot of the story. Whereas the nuclear genome, you inherit from all your ancestors. If you imagine the significance of this – with every generation, you’d double the number of people that you’re related to with the nuclear genome. It’s really the full picture of your genetic ancestry.

FLG: What were some of the key takeaways from the Neanderthal paper from your perspective?

Benjamin: In addition to what we were able to tell about the specific Neanderthals who lived at that site, I think for me, it was mostly a proof of concept. This is the first time we were able to get nuclear DNA from sediments. It’s the first time that we were able to do proper population genetics – which is what we call our field – where we look at the genetics of large groups of people and are able to do that from sediments. So this really opened up the field. You know, as I said before, every archaeological site has sediments, this really is a technique that we can apply pretty much anywhere.

FLG: What are you working on at the moment?

Benjamin: Well, I can’t talk about specific sites or projects. But the thing that we’ve done so far with the sediment DNA is we’ve used it in situations where we didn’t have bones. We’ve used it as a replacement for bones. I think there are a couple of really key things we can do going forward. For one, there are some ancient bones that we shouldn’t sample, that we shouldn’t drill into, because it wouldn’t be ethically correct to do that. There are cultural artifacts from the communities where those bones were found where they’re the only known example of their type. Even though we actually only take a tiny, tiny amount of bone powder, we still shouldn’t do that in some circumstances. And there are some cases where maybe you could use sediment DNA instead to circumvent that and still be able to study these people in a more ethical way.

But what I’m really excited about is that when we get DNA from a bone, we essentially get it from where someone died – we have a record of their death, a record of their final resting place. But they have this whole life that we don’t know about. I’m leaving DNA in my office; you’re leaving DNA in your office. And by getting DNA from the sediments, we can study where people lived. We can study, for example, if there’s a workplace, who worked in that workplace versus who lives in the village nearby, stuff like that. I think that by using sediment DNA – this is the stuff that’s really exciting to me now – we can study where people actually lived and spent their lives, as opposed to just their deaths.

FLG: Yeah, you don’t really think about how traditional approaches only allow you to study where those people died, rather than where they spent their entire lives. That’s really fascinating. Are there any emerging areas in the field that you’re particularly excited about?

Benjamin: I mean, there are all sorts of things that people are doing with sediment DNA these days. We’ve tended to focus on human DNA in the sediments, but there are a bunch of researchers who are doing this for broader environmental questions. You know, looking at how the ecosystem changes over time and using the sediment DNA as a fine scale record of that – again, taking advantage of the fact that we can do many, many samples down a stratigraphy.

I think there are probably a lot of applications in the field of forensics as well. And some of my colleagues have done some of that work, where they use the methods we have developed for looking at ancient DNA to help with cold case, for example. I think in general, the technology that we develop is useful in a lot of different areas.

FLG: Are there any developments that you’d like to see in the future that might help with your research or overcome some of the many challenges that we’ve already discussed?

Benjamin: I guess I would just get greedy, you know. Typically, from a cave, we might look at between 100 and a few 100 sediment samples. I would really like to go bigger than that. But in order to do that, we need to make it cheaper, we need to develop the technology so we can process more samples. We’re also working on ways to target even rarer species that might be in the samples. With some sediment samples, you can’t find the human DNA, but you know people were there. So, it would be great if we were able to target them more accurately. I mean, the wish list is long, I guess. But in general, I would just like more data.

FLG: Yeah, I’m sure that would be on most people’s wish list. That’s actually all we’ve got time for today. I’ve certainly learned a lot about ancient environmental DNA that I did not know before. So, I would just like to say thank you again for taking the time to answer these questions.

Benjamin: Thank you for having me.

Watch more interviews in our series here.