Spot that horse!

Roughly 25,000 years ago in what is now southwestern France, human beings walked deep into a cave and left their enduring marks. Using materials like sticks, charcoal and iron oxides, they painted images of animals on the cave walls and ceilings, lions and mammoths and spotted horses, walking and grazing and congregating in herds.

Today, the art at the Pech-Merle cave, and in hundreds of others across Europe, is a striking testimony to human creativity well before modern times. But what were these cave paintings, exactly? Were prehistoric artists simply sketching what they saw each day on the landscape? Or were the images more symbolic, diverging from reality or representing rare or even mystical creatures? Such questions have divided archaeologists for years. Now, a group of researchers has used distinctly modern techniques to help decipher the mystery, at least in the case of Pech-Merle’s famous spotted horses.

By comparing the DNA of modern horses and those that lived during the Stone Age, scientists have determined that these drawings are a realistic depiction of an animal that co-existed with the artists.

The research, published in the journal Proceedings of the National Academy of Sciences, grew out of an effort to discern the coat colours of ancient horses to help figure out when the animals were domesticated, a pivotal moment in the development of human societies.

In general, domesticated species exist in a far greater variety of colours than wild ones, so understanding colour variation in fossil animals can help pinpoint the timing.

Previous research on DNA from the bones and teeth of horses that lived 7,000 to 20,000 years ago showed that those animals were either black or bay (a brown coat with a black mane and tail).

That work was published in the journal Science in 2009. Since then, geneticists have deciphered the underlying code for the spotted pattern, known as leopard, in modern horses. So the scientists went back to their samples, looking for the leopard sequence in horses that lived in Europe 11,000 to 15,000 years ago.

Cumbersome process

“There is a striking correspondence between the coat-colour patterns of horses painted in Paleolithic caves of France with what geneticists found in the genotypes” – the specific genetic sequences – “of colour genes,” said Hopi E Hoekstra, an evolutionary biologist at Harvard who studies pigmentation. Hoekstra was not involved in the study but called it “very convincing.”

An author of the study, Michael Hofreiter, an evolutionary biologist at the University of York in England, said: “Why they took the effort making these beautiful paintings will always remain a miracle to us. It’s an enigma, but it’s also nice to see that if we go back 25,000 years, people didn’t have much technology and life was probably hard, but nevertheless, they already endeavoured in producing art. It tells us a lot about ourselves as a species.” Extracting DNA from such old material is a complex process, and the potential for contamination is huge.

Early studies of Neanderthal DNA were marred by contamination from humans and led to skepticism about the field’s future. Since then, researchers have adopted strict procedures to ensure they are not contaminating ancient samples with modern-day DNA. The procedures include analysing ancient and contemporary material in physically separate facilities and replicating results multiple times. “This is a whole different level of clean,” said Jessica L Metcalf, a postdoctoral researcher at the University of Colorado who also works with the Australian Center for Ancient DNA in Adelaide, where the labs that work on ancient and modern DNA are more than half a mile apart. “We have sealed rooms with HEPA filter air flow, UV lights that sterilise when you come in,” she said.

“We spend over half our time cleaning. We use a lot of bleach. You’re in this ridiculous-looking clean suit with a face shield on.” In fact, Hofreiter said, researchers who work with ancient horse genes should not even go horseback riding. “Traces of DNA,” he said, “they just stick to people.” Hofreiter, 38, began his career working with a pioneer of ancient DNA research, Svante Paabo. Although he intended to study taxonomy, he was so intrigued by the idea of extracting DNA from ancient material that he switched his focus. “You have this 30,000-year-old piece of feces in your hand,” he said, adding: “Well, you should wear gloves. And you can actually get to the genetic code from the animal. And I thought, this is so fascinating.” He and his colleagues did not set out to study cave art.

They were simply continuing their work on coat colour in prehistoric horses. Only after they found the spotted horse gene in their ancient samples did they realise they could say something about archaeology. “What we found is that there were really only these three colour patterns – spotted or dappled; blackish ones; and brown ones,” he said.

More symbolic than realistic

“These are the three phenotypes we find in the wild populations. And then we realised these phenotypes are exactly the ones you see in cave paintings.” Terry O’Connor, an archaeologist at the University of York who collaborated on the study, said that spotted horses in particular had been used to argue that cave art was more symbolic than realistic, and that as a result, the finding could cause a stir. But now it is clear that some horses had a gene for that coat colour. “People drew spotty horses,” he said, “because they saw spotty horses.”

Last summer, exploring a cave in the Dordogne region, O’Connor said, he became transfixed by a series of line drawings of mammoths. “They were absolutely superb, some using contours of the cave itself, capturing the size and shape and movement,” he said.
“You look at that and say, ‘These guys know what the animals looked like, and they can draw.”’

As techniques for working with ancient DNA have matured, scientists are now using it to answer an increasing variety of questions about the past, from what happened to a species’ genetic variation as its environment changed to how humans recolonised Europe after the last ice age to what type of microbes lived in the guts of people and animals thousands of years ago. “One of the things that most pleases me about this paper as a piece of ancient DNA science,” O’Connor said, “is it kind of begins with a question. These spotty horses, were they magical or real? “And then science answers that. It’s not just, ‘Let’s rip the DNA out of ancient bones and see what it tells us.”