Method can identify the species of thousands of archaeological bone fragments

Tech Science 21. aug 2022 2 min Postdoctoral Fellow Patrick Rüther Written by Kristian Sjøgren

A new method for analysing proteins in bones can help archaeologists in classifying the thousands of bone fragments they find in archaeological excavations. The method can provide greater insight into what our ancestors ate and the livestock they kept, says a researcher.

Archaeological excavations almost always find hundreds of unidentifiable small bone fragments. These may contain important knowledge about the past. What did our ancestors eat? What types of farm animals did they have?

However, identifying which animals the various bone fragments come from is often virtually impossible. An expert could identify them one by one, but they need to be a certain size and identification takes a considerable time. Another option is whole-genome sequencing each fragment, but this is very expensive.

The result is mostly that none of the above happen, and museum basements are therefore often filled with sacks of thousands of bone fragments from excavations from Stone Age settlements, Neanderthal caves and other sites.

However, a new method could help archaeologists to illuminate the unknown.

Using a new protein analysis pipeline, researchers can determine the species of 200 bone fragments per day. This enables researchers to rapidly determine the origin of the bone fragments and to use this information for interpreting history or prehistory.

“This method was originally developed to obtain rapid results for proteins related to disease. We have adjusted the method to use it on archaeological bones and could further tweak it for processing teeth, skin or hair,” explains a researcher behind the study, Patrick Rüther, Postdoctoral Fellow, Novo Nordisk Foundation Center for Protein Research, University of Copenhagen in the Mass Spectrometry for the Quantitative Proteomics group led by Jesper Velgaard Olsen.

The research behind the development of the method has been published in Nature Communications.

Shaves hours off analysis time

Patrick Rüther has streamlined various existing methods for analysing the protein content of a biological sample.

The whole process normally involves three steps:

  • extracting the proteins from the sample using various chemicals;
  • analysing the sample using mass spectrometry, which can be used to determine which proteins are present in a given sample; and
  • processing the data from mass spectrometry using advanced computer programs to link proteins to biology, which here means determining the species.

Previous methods typically took 1–3 hours for a single sample and were thus unsuitable for analysing large sacks of bone fragments.

By streamlining all the steps in the process, Patrick Rüther and colleagues have reduced the time to 7 minutes.

“This means that we can analyse 200 samples a day, and the cost of analysing each sample has been reduced enough to make the process financially viable,” he says.

Identifying 156 species so far

An important step in the new research was developing a method that automatically determines the most likely species based on the identified proteins.

This has required researchers to develop new models that compare the findings from mass spectrometry with databases for proteins in various species.

For example, the researchers use the models and databases to link the discovery of a specific protein in collagen in donkeys to the databases and thereby know that their sample originates from a donkey.

In total, the researchers can use their method to identify 156 species based on protein present in bones.

“Previous methods have been good at classifying broader species families by mass spectrometry. In this study, we had to add an extra layer of protein separation and thereby distinguish even closely related species,” explains Patrick Rüther.

Analysing bones up to 60,000 years old

Patrick Rüther says that the researchers plan to expand the database of animals so that they may be able to identify even more in the future.

The 156 species currently include most mammals, but some are so similar that distinguishing between, for example, a domesticated cow and its ancestor, an aurochs, is still difficult.

Nevertheless, researchers can distinguish between a donkey and a horse or humans and chimpanzees. Other methods have had great difficulty with this, and this technology therefore provides a more detailed picture of the past.

Patrick Rüther says that some of the samples the researchers have examined are between 40,000 and 60,000 years old and originate from a Neanderthal residence in Portugal.

Using the method, the researchers identified which animals our distant cousins hunted and ate.

“The method also opens up the possibility of analysing the bone fragments during the course of the excavation, thereby guiding it. If some of the bone fragments originate from humans, whole-genome sequencing will also be relevant to determine much more about the people who lived in this place many millennia ago,” concludes Patrick Rüther.

SPIN enables high throughput species identification of archaeological bone by proteomics” has been published in Nature Communications. Several authors are affiliated with the Novo Nordisk Foundation Center for Protein Research, University of Copenhagen.

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