Burial I6113 was the only one that yielded ancient DNA from the Indus Valley civilization.Vasant Shinde

The climate of South Asia is not kind to ancient DNA. It is hot and it rains. In monsoon season, water seeps into ancient bones in the ground, degrading the old genetic material. So by the time archeologists and geneticists finally got DNA out of a tiny ear bone from a 4,000-plus-year-old skeleton, they had already tried dozens of samples—all from cemeteries of the mysterious Indus Valley civilization, all without any success.

The Indus Valley civilization, also known as the Harappan civilization, flourished 4,000 years ago in what is now India and Pakistan. It surpassed its contemporaries, Mesopotamia and ancient Egypt, in size. Its trade routes stretched thousands of miles. It had agriculture and planned cities and sewage systems. And then, it disappeared. “The Indus Valley civilization has been an enigma for South Asians. We read about it in our textbooks,” says Priya Moorjani, a computational biologist at the University of California at Berkeley. “The end of the civilization was quite mysterious.” No one alive today is sure who the people of the Indus Valley civilization were or where they went.

A pair of newly published papers use ancient DNA to shed light on the Indus Valley civilization and the entire history of people in South and Central Asia. The first study is a sweeping collection of 523 genomes—300 to 12,000 years old—from a region spanned by Iran, Russia, and India. By comparing the results with modern South Asians’ genomes, the study showed that South Asians today descended from a mix of local hunter-gatherers, Iranian-related groups, and steppe pastoralists who came by way of Central Asia. It’s the largest number of ancient genomes reported in a single paper, all made possible by an ancient DNA “factory” the geneticist David Reich has built at Harvard. (Moorjani completed her doctorate in Reich’s lab and is a co-author on this paper.)

The second study focuses on just a single genome from the Indus Valley civilization: I6113, a woman who died more than 4,000 years ago. Her skeleton was the only one—out of more than 100 samples the researchers tested from 10 different Indus Valley–civilization sites—that yielded ancient DNA, but even then it was contaminated and of poor quality. “We had to squeeze, squeeze, squeeze the sample really hard, more than we’ve done in any other sample we’ve ever tried,” says Reich, who is also a senior author of the second paper. The team ultimately tried to sequence DNA from I6113’s ear bone more than 100 times, each time yielding a tiny dribble of genetic data. That I6113 gets her own paper is a testament to both the technical difficulty of sequencing her DNA and the importance of the Indus Valley civilization. Even before publication, rumors were swirling in India about what the ancient DNA would show, and how it would play into the politics of the Hindu-nationalist ruling party.

What’s intriguing about I6113’s DNA is what she lacks: any of the steppe ancestry that is widespread in contemporary South Asians. Instead, she appeared to have a mix of Southeast Asian hunter-gatherer and Iranian-related ancestry.

The two studies piece together a history of how the people of the Indus Valley civilization are related to South Asians today. After the decline of the civilization 4,000 years ago, people with a genetic makeup similar to I6113 mixed with people of Southeast Asian hunter-gatherer ancestry to form what has been called Ancestral South Indians. From 4,000 to 3,000 years ago, other people descended from the Indus Valley civilization mixed with people of steppe-pastoralist ancestry, who likely brought horses and the Indo-European languages now spoken on the subcontinent, to form a group that has been called Ancestral North Indians. These two ancestral groups then mixed as well, giving rise to the great diversity of ethnic groups in South Asia. Go back far enough, and both sides trace to the Indus Valley civilization, which appears to be the single largest source of ancestry for modern South Asians.

The team studying I6113 noticed something intriguing about the Iranian-related portion of her ancestry, too. It appears to date to before the advent of farming in the Fertile Crescent. This suggests that farming did not, as many have thought, spread to South Asia through the migration of people from the Near East. It may have arisen independently in South Asia or spread through cultural contact.

Of course, this is a lot to rest on a single genome. “That would be like taking a single sample from Tokyo and trying to generalize about the whole ancestry of Japan,” Reich admits. But the team’s confidence in its results was bolstered when the researches found that I6113 was genetically similar to 11 people from the 523-genome paper who were buried not in South Asia, but in what is now Iran and Turkmenistan. These 11 people were also “outliers” in their own burial sites. The team thinks they may have been migrants or the children of migrants from the Indus Valley civilization. Archaeological evidence suggests people traveled between these regions as well.

The cities of the Indus Valley Civilization were cosmopolitan places, which also makes it harder to generalize from one genome. J. Mark Kenoyer, an anthropologist at the University of Wisconsin at Madison who was not an author of either study, cautions that only a small number of people who lived in these cities were buried in cemeteries—probably elites. The rest might have been cremated, or their bones simply left uncovered and thus scattered over time. “The cemeteries of the Indus civilization do not represent the people of the Indus civilization. They represent one community,” he says.

Still, more cemetery samples would be better than just one. The research team behind I6113 is trying to sequence more bones from the Indus Valley civilization. Vasant Shinde, an archeologist at Deccan College whose team excavated I6113, says the attempts to get ancient DNA from Indus Valley–civilization sites have been a years-long learning process. To prevent contamination with modern DNA, team members now wear gowns and masks even while excavating in the field. They do not reuse excavation instruments from burial to burial. Niraj Rai, a geneticist who was a visiting fellow in Reich’s lab, also set up an ancient-DNA lab at the Birbal Sahni Institute of Palaeosciences in Lucknow, India, where I6113’s DNA was extracted. “This is beginning,” Shinde says. “This is not the end.” He expects more ancient DNA to come.

In India, ancient DNA has generated intense interest, says Tony Joseph, the author of Early Indians: The Story of Our Ancestors and Where We Came From. He told me his book, published last December, is already in its seventh printing. After a preliminary version of the large Central and South Asian genomes study was posted on bioRxiv last March, it became the site’s most downloaded preprint of 2018. The preprint generated controversy, too, especially the finding that many Indians have ancestry from steppe pastoralists. Hindu nationalists, as Joseph has written, believe that Aryans—who originated in India and spread through Europe and Asia—are the source of Indian civilization. This is contradicted by ancient DNA that finds the population history in India itself contains far more mixing and migration. (Further complicating things, Nazis co-opted the term Aryans to mean something different, a master race of European origin.) A prominent MP even attacked Reich when the preprint came out, tweeting out an article titled, “There Are Lies, Damned Lies and (Harvard’s ‘Third’ Reich and Co’s) Statistics.” Reich, who has experienced how fraught talking about genetics and identity can be, acknowledged the political interest in his work, but declined to get into it.

Ancient DNA has captured the public imagination precisely because it promises an answer to questions like Where did we come from? and Who are we?—questions that also have deep political undercurrents. To sequence I6113’s DNA is to draw genetic connections between an ancient civilization and the people who live in the region today, to add fuel to arguments about who can lay claim to a cultural inheritance. All this, contained in a half-inch wisp of an ear bone.

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