CSI and detecting the presence of ancient humans

Enter a room, even for a few minutes, and dead skin cells will follow you like an invisible cloud to settle on exposed surfaces. Live there and a greyish white, fluffy dust builds up in every room. Even the most obsessive cleaning will not remove it, especially under a bed or on the bathroom floor. Consider a cave as a home, but one without vacuum cleaners, any kind of sanitation, paper tissues, panty liners, nappies or wet wipes. For pre-modern human dwellings can be added snot, fecal matter, sweat, urine, menstrual blood and semen among all the other detritus of living. A modern crime-scene investigator would be overwhelmed by the sheer abundance of DNA from the host of people who had once dwelt there. CSI works today as much because most homes are pretty clean and most people are fastidious about personal hygene as because of the rapidly shrinking lower limit of DNA detection of the tools at its disposal. Except, that is, when someone from outside the home commits a criminal offence: burglary, GBH, rape, murder. We have all eagerly watched ‘police operas’ and in the absence of other evidence the forensic team generally gets its perpetrator, unless they did the deed wearing a hazmat suit, mask, bootees and latex gloves.

Artistic impression of Neanderthal extended-family life in a cave (credit: Tyler B. Tretsven)

Since 2015 analysis of environmental DNA from soils has begun to revolutionise the analysis of ancient ecosystems, including the living spaces of ancient humans (see: Detecting the presence of hominins in ancient soil samples, April 2017). It is no longer necessary to find tools or skeletal remains of humans to detect their former presence and work out their ancestry. DNA sequencing of soil samples, formerly discarded from archaeological sites, can now detect former human presence in a particular layer, as well as that of other animals. In many cases the ‘signal’ pervades the layer rather than occurring in a particular spot, as expected from shed skin cells and bodily fluids. The first results were promising but only revealed mitochondrial DNA. Now the technique has extended to nuclear DNA: the genome (Vernot, B. and 33 others 2021. Unearthing Neanderthal population history using nuclear and mitochondrial DNA from cave sediments. Science, v. 372, article eabf1667; DOI: 10.1126/science.abf1667). Benjamin Vernot and colleagues from 7 countries collected and analysed cave soils from three promising sites with tangible signs of ancient human occupation. Two of them were in Siberia and had previously yielded Neanderthal and Denisovan genomes from bones. The other is part of the Atapuerca cave complex of NW Spain that had not. The Russian caves yielded DNA from more than 60 samples, 30 being nuclear DNA consistent with that from actual Neanderthal and Denisovan bones found in the caves. Galería de las Estatuas cave in Spain presented a soil profile spanning about 40 thousand years from 112 to 70 ka.

Teasing-out nuclear DNA from soil is complicated, from both technical and theoretical standpoints. So being able to match genomes from soil and bone samples in the Russian caves validated the methodology. The Spanish samples could then be treated with confidence. Galería de las Estatuas revealed the presence of Neanderthals throughout its 40 ka soil profile, but also a surprise. The older DNA was sufficiently distinct from that from later levels to suggest that two different populations had used the cave as a home, the original occupants being replaced by another genetically different group around 100 to 115 ka ago. The earlier affinity was with the ancestors of sequenced Neanderthal remains from Belgium, the later with those from Croatia. That time is at the end of the last (Eemian) interglacial episode, so one possibility is a population change driven by climatic deterioration. This success is sure to encourage other re-examinations of caves all over the place. That is, if there is the analyical capacity to perform such painstaking work in greater volume and at greater pace. Like many other palaeo-genomic studies, this one has relied heavily on the analytical facilities pioneered and developed by Svante Paäbo at the Max Planck Institute for Evolutionary Anthropology in Leipzig, Germany. Covid has forced genetics to the front page for a year and more. And it has led to many advances in anlytical techniques, particularly in their speed. It would nice to think that a dreadful experience may end-up with positive benefits for understanding the full history of humanity.

Detecting the presence of hominins in ancient soil samples

Out on the plains countless herbivores fertilise the ground by continual urination and defecation. A friend’s sheep are doing just that in the small field that came with my current home while they are keeping the grass under control.  Millions of hectares of prime agricultural land in China are kept fertile through disposal of human night soil from ‘honey wagons’ every day; it is even fed to fishes in small ponds. Such a nice economy also donates the DNA of the animal and plant inhabitants to the soil system. In 2015 analysis of environmental DNA from permafrost in Siberia and Alaska produced ‘bar codes’ for the now vanished ecosystems of what was  mammoth steppe during the climate decline to the last glacial maximum and the subsequent warming. The study revealed mammoth and pre-Columbian horse DNA and changes in the steppe vegetation, from which it was concluded that the steppe underwent regional extinction pulses of its megafauna linked to rapid climate ups and downs connected with Dansgaard-Oeschger cycles. It was but a small step to see the potential for studying distribution and timing of various hominins’ occupation of caves from the soils preserved within them, without depending on generally very rare occurrences of human skeletal remains.

Tourists at the entrance to Denisova Cave, Rus...
Tourists at the entrance to Denisova Cave, Russia (credit: Wikipedia)

The Max Planck Institute for Evolutionary Anthropology in Leipzig, now famous for extracting DNA from Neanderthal, Denisovan and possibly H. antecessor fossils, has applied the environmental DNA approach to sediments from 7 caves in France, Belgium, Spain, Croatia and Russia that span the period from 550 to 14 ka (Slon, V. and 30 others 2017.  Neandertal and Denisovan DNA from Pleistocene sediments. Science, v. 356 (online publication); doi:10.1126/science.aam9695). The sites had previously yielded fossils and/or artefacts. All of them contained mitochondrial DNA from diverse large mammals, four including archaic human genetic material supplied by Neanderthal individuals and Denisovans in the case of the Denisova cave. A key finding was Neanderthal mtDNA in one sedimentary layer that contained no skeletal remains – decay of a body was probably not involved. In two cases the DNA was from more than one individual. A variety of tests showed that surprisingly large quantities of DNA survive in soil and that it is spread evenly in sediment rather than being present in spots – an indication of derivation from urine, excreta or decayed soft tissue.

Although the study does not add to knowledge of hominin genetics, it confirms that the methodology is sufficiently advanced and efficient to detect hominin presence in fossil-free sediment. So this approach seems set to become a standard for many sites, such as that from California reported in the previous post, which suggest a human influence, or any cave sediments for that matter. Although skeletal remains are essential for reconstruction of bodily characteristics, hominin phylogeny seems set to cut loose from fossils. Hitherto suspected species’ presence in the time period where DNA analysis is feasible may be detected, such as Asian H. erectus. It may become possible to map or extend the geographic ranges of Denisovans and Neanderthals. Perhaps species new to science will emerge.

More on late Pleistocene hominin genetics here

Wade, E. 2017. DNA from cave soil reveals ancient human occupants. Science, v. 356, p. 363.

Wade, E. 2017. DNA from cave soil reveals ancient human occupants. Science, v. 356, p. 363.