"Ours, it turns out, is a repeated history of migrations and mixing," says Cosimo Posth, a junior professor of archaeo- and palaeogenetics at the University of Tübingen in Germany. "Much of what we thought we knew about human history – that populations lived for centuries mostly without mixing – was overturned thanks to modern-day genomics."
In March 2023, Posth and his colleagues analysed the largest ever ancient genetic data set (356 – including 116 from new-found remains in 14 countries) of prehistoric hunter-gatherers, from which they derived new insights about the groups' migration and survival.
A mere decade or so earlier and their work wouldn't have been possible. Breakthroughs in collecting and deciphering ancient human DNA during this time have rapidly accelerated discoveries about the origin of our species – Homo sapiens – often, re-writing history.
"These technologies have enriched the field tremendously," says Chris Stringer, a British anthropologist at the Natural History Museum who has worked in the field for over 50 years. "They've given us so much more information and they're developing all the time."
Cosimo Posth was able to show that early modern humans and Neanderthals had mixed more than 200,000 years ago
Image credit: University of Tübingen
It all started at the turn of the century. Revelations started to come thick and fast following a major breakthrough in ancient genome sequencing, explains Stringer.
Back then, it was thought human evolution over the last 500,000 years was fairly well understood. It was believed to begin with Homo heidelbergensis, known to be from Africa and western Eurasia between about 600,000 and 400,000 years ago. These gradually evolved into Neanderthals, in western Eurasia, and Homo sapiens, in Africa.
Our species later dispersed from Africa, about 60,000 years ago. By around 30,000 years ago, we had completely replaced the 'inferior' Neanderthals across Eurasia, with little or no interbreeding.
Or so it was thought. Then, in the mid-1990s, Swedish geneticist Svante Pääbo deciphered a relatively short component of mitochondrial DNA from a Neanderthal male. A feat that seemed like "science fiction" at the time, says Stringer.
"No one could have imagined that in ten years we'd have whole genomes from fossils."
But that is exactly what happened. By 2010, off the back of advances in genome sequencing for healthcare and medicine, Pääbo drafted a whole genome of a Neanderthal. This, and subsequent work, demonstrated – rather shockingly – that Neanderthals and other extinct hominids made a significant contribution to our ancestry.
In other words, Homo sapiens did interbreed with other species living at the time and humans have inherited about 2 per cent of their DNA from Neanderthals.
Other discoveries and revelations followed: that of a previously unrecognised kind of human, the Denisovans, from a whole genome recovered from a finger bone fragment found in a Siberian cave. That the arrival of Homo sapiens in Europe was 150,000 years earlier than previously understood was confirmed by the dating of a fossil found in Greece in 2019.
Being able to sequence ancient human DNA has been game-changing, but working with it is vastly different from that of modern humans, explains Dr Matthias Meyer, a biochemist who was part of Pääbo's group developing those early breakthroughs. He is now head of the Advanced DNA Sequencing Techniques group at the Max-Planck-Institute for Evolutionary Anthropology.
Ancient DNA typically has chemical modifications, is extremely short, and often only small quantities are recoverable, making it very difficult to extract from a biological sample for processing by a sequencer. For example, DNA from blood can produce thousands to millions of DNA base pairs, compared to around 40-60 from ancient DNA.
Image credit: Jürgen Vogel/LVR-LandesMuseum Bonn
"Special methods are needed to get as much DNA as possible from a sample, as well as sophisticated analysis tools to interpret the sequences and minimise errors," Meyer explains.
The study Posth led at the University of Tübingen, which analysed 356 ancient human DNA samples, used such methods previously developed by Meyer.
Posth's team first extracted 30-50mg of bone powder to test for DNA. "The bones have been lying around for thousands of years and are therefore packed with environmental bacteria and non-endogenous DNA, so we needed to essentially fish out the human DNA," he explains.
This is done by a process called targeted enrichment, Meyer's method. In an ancient DNA lab, probes laden with synthetic oligonucleotides (synthetic sequences of DNA) fish out the DNA of interest. Preparing the probes is meticulous and time-consuming, but Posth says some emerging companies can provide them as ready to use.
"Before capture we might have 1 per cent of human DNA and after around 40 per cent, but it will never be 100 per cent," he says.
Once obtained, ancient human DNA can be identified using high-quality genome sequences from modern humans, says Meyer. These sequences can be used as a reference on which to map the short fragments of ancient DNA using a powerful computer, a process known as mapping assembly.
"Once enough coverage across the reference genome has accumulated, a researcher can obtain firm information about the ancient individual," he explains.
In a day, using powerful sequencers, computers and sophisticated software, billions of DNA sequences can be generated.
"Ten years ago, I wouldn't have imagined one can go as fast as this now," says Meyer.
But the process is much more costly than sequencing modern human DNA. Samples with almost no microbial contamination can generate high-quality genomes for around €10,000, according to Meyer.
From their ancient human DNA analysis, Posth and his team could compare samples and group some individuals together from the same archaeological site to form a population. These could then be compared between sites and through time, Posth explains.
The research, which lasted six years, determined that, during the last glacial maximum, which occurred between 19,000 and 20,000 years ago in Iberia, humans survived after migrating to fairer climes. But in the Italian peninsula, the population was replaced – how or why is not known – most likely from another known climatic refugium from the Balkans.
The reach of ancient DNA is destined to be limited, however. Its preservation is entirely dependent on the environment. It has recently been shown to be preserved in permafrost for about two million years, but in more temperate climes, such as Central or Southern Europe, the current record is 400,000 years from a deep cave system in Northern Spain.
Posth believes techniques for working with ancient DNA are improving, though – and in the study they used a new method that allowed them to extract shorter molecules, called the single-stranded library protocol – but future major breakthroughs will be in the computational analysis, he believes.
"This is where new methods will emerge, pushing the limits of the resolution in our inferences to identify more distantly related relatives, such as individuals that share a common ancestor one hundred years apart. It's a matter of developing an algorithm," Meyer says.
For DNA recovery, he feels there's limited potential in optimising methods by "yet another order of magnitude" and certainly not to reach "millions of years outside permafrost". Meaning it's unlikely there will ever be DNA from really early humans.
These human fossils, found on a Dutch coast, originally came from Doggerland, a now submerged land under the North Sea, where European hunter-gatherers lived, a DNA analysis found
Image credit: National Museum of Antiquities (RMO)
Researchers are therefore exploring new avenues of understanding. Stringer and his colleagues at the Natural History Museum are working to retrieve genetic data from fossil proteins, which can have a much longer survival time and potentially unlock ancient history from two to five million years ago.
Stringer refers to a robust and primitive jawbone fossil found a couple of years ago in the Tibetan plateau of China. No DNA could be extracted from it, but protein material was retrieved which matched the coding in the Denisovan genome rather than Neanderthal or Homo sapiens.
"That jawbone, which has a big molar tooth that matches what we know of the molar teeth from the Denisova cave, is probably from a Denisovan. This shows that protein evidence can be used to identify fossils," says Stringer.
He hopes to test more fossils to see how they relate to people based on proteins, as well as ancient DNA.
"It's early days but hopefully it will develop into a field that will enrich knowledge of human evolution, just as much as the ancient DNA," he says.
Although much has been discovered, there is still an abundance more to learn about human evolution; vast areas of Africa have still not produced a single ancient fossil bone or skull, though stone tools show people were there, and the Indian subcontinent has only one significant ancient human fossil.
These finds will only come from funding exploration in the right areas, says Stringer. But he cautions that more is needed, especially in the UK which, due to Brexit, has seen a reduction in real-terms science funding, as well as lost connections with many European partners, he says.
Posth believes the surface of what's possible "is just being scratched".
"Now we're getting an overview of all this human movement through time, we need to try to understand its causes. There's much to be done to get an even better glimpse of our past."
Bones, sediment and artefacts
Finding ancient human remains is rare, which is why researchers, including Meyer and Posth, are experimenting with extracting human DNA from other areas, such as earth sediment.
Sediment is already used to date an archaeological find by a process called palaeomagnetism. This determines the state of the Earth's magnetic field, which fluctuates through time, within the sediment to know when deposits originated.
During the past few hundred thousand years, humans visited and occupied caves leaving DNA signatures in them.
Meyer and his group found that by taking samples every few centimetres, in columns, effectively creating a grid, it's possible to find samples containing human DNA.
"This is fascinating because it allows us to put ancestry information to the cultural records of a site, so you can know who produced the stone tools there," he says.
Methods to retrieve the human DNA from the sediment are similar to those already used for bones but the process is trickier due to the complex mixtures and contamination, including of other mammal DNA.
"We know many positions in the human genome where humans are sufficiently different from other species. At these positions we can have a high confidence of identifying human DNA and then make inferences about the population history."
There are projects under way that are already starting to challenge assumptions archaeologists had made about who produced which tools, Meyer adds.
In the Siberian cave where Denisovans where first discovered, Meyer and his group of researchers took more than 700 sediment samples and identified more than 150 samples that contain Neanderthal, Denisovan, or modern human DNA.
"We showed that Denisovans were probably the first occupants of the site, and that Neanderthals came later, and then there were periods of a mixture of the two, until finally, modern humans replaced them all," explains Meyer.
His group are also looking at ways to extract DNA from artefacts, including stone-age pendants.
"One pendant we examined contained ancient human DNA, enabling us to produce a very detailed genetic profile of the wearer, which I think is really quite spectacular."
It is, however, very difficult to find DNA on non-porous artefacts like stone tools, Meyer adds.
"This integration between archaeology and genetics is moving on and there will be more and more archaeological questions that genetics can help to answer," he says.
Posth, who is also working on developing techniques to retrieve human DNA from sediment, says the possibility is "incredible" as it "gives us an opportunity to analyse human DNA in the absence of fossils".
Why does any of this matter today?
"Understanding what has happened in the past gives us an understanding about what is happening today," says Posth. He refers to patterns of human migration and even climate change. "It's not new to human history; but the reasons for it change."
On a more practical level, insights into ancient human DNA can provide useful knowledge about modern humans. It has been shown that different bits of Neanderthal DNA confer protection and increased risk of human hospitalisation from Covid-19 and can be linked to some autoimmune diseases in modern humans.
Dr Philipp Gunz, a paleoanthropologist at Max-Planck-Institute for Evolutionary Anthropology, and his team are using genetic human data and brain scans from 4,500 people to explore how Neanderthal gene fragments affect the shape of the human brain. They hope to uncover differences in people who have these gene fragments.
They're currently expanding their work to include data from the UK Biobank, which will allow them to explore the entire genome as well as other body parts.
"Such data sets are game-changing," says Gunz. "We are only scratching the surface – I expect new fossil discoveries and insights into the biological underpinnings of the changes we see in human evolution and to start to see a better understanding of what this means for health and disease in living people."
