IAP-26-033
Human ancestry and social organisation at the dawn of the Iberian Megalithic phenomenon
The transition to farming represents one of the most profound transformations in human history. Beginning in Southwest Asia around 12,000 years ago, farming practices subsequently spread into Europe after the 7th millennium BC. Ancient DNA (aDNA) studies have shown that the arrival of these early farming populations resulted in a substantial genetic turnover across much of Europe, often replacing a large proportion of the indigenous hunter-gatherer (WHG) gene pool with new Anatolian Early Farmer (AEF) ancestry. However, this process was not uniform, and in several regions, including parts of the Iberian Peninsula, interactions between farmers and local hunter-gatherers led to varying degrees of admixture, creating genetically diverse communities (1). A particularly striking pattern emerged during the Middle and Late Neolithic, when populations across Europe began to show an increase in WHG ancestry from their preceding populations. While the genetic phenomenon is now well documented, the social processes underlying this shift remain largely unexplored. In Iberia, this broadly coincides with the emergence of funerary megalithic traditions. Archaeogenetic research suggests that these monumental funerary spaces were not simply communal burial grounds but may have been structured around biological kinship. Studies from different parts of Neolithic Europe have identified close familial relationships among individuals buried within the same monuments, while some evidence suggests preferential burial of individuals linked through paternal descent lines, raising the possibility that certain monuments were used by socially prominent lineages or emerging elites (2). However, it remains unclear whether such patterns were also present in Iberia, how they varied between different funerary traditions, and whether they were connected to the demographic processes that produced the resurgence of WHG ancestry (3). Equally unknown is whether health and exposure to disease varied based on ancestry, sex, or social affiliation within these communities.
This project will address these questions through the study of genomic data from Middle Neolithic individuals buried in caves and megalithic monuments across western and northern Iberia, combining the analysis of ~75 newly sampled individuals with the reanalysis of previously published genomes. One third of the newly sampled individuals have already been screened in our laboratory demonstrating sufficient endogenous DNA preservation (4).
Specifically, the project will address three interconnected questions:
1. How was the resurgence of WHG ancestry distributed among Middle Neolithic populations of Iberia, and what does this reveal about the processes of admixture shaping these communities?
2. Did different funerary traditions, particularly caves and megalithic monuments, display distinct patterns of biological relatedness, suggesting different forms of social organisation or access to burial space?
3. Were ancestry, sex, or patterns of kinship associated with differences in pathogen presence and broader indicators of health when considered alongside archaeological and osteological evidence?
Using genome-wide ancient DNA data, the project will reconstruct ancestry profiles, identify biological relationships within burial sites, determine biological sex, and screen for ancient pathogens where preservation allows. These results will be interpreted alongside existing archaeological and bioarchaeological evidence to explore how demographic change was experienced at the level of communities and families.
Methodology
Skeletal samples will be selected, prioritizing pars petrosa of the temporal bone and teeth, and drilled. DNA will be extracted from 30-50mg of bone/tooth powder and transformed into double-stranded, double-indexed UDG-treated libraries as per (5,6,7). These libraries will be shotgun sequenced and screened for human DNA and pathogen content using nf-core/EAGER (8) and aMeta (9). Libraries with aDNA signatures of authenticity and sufficient endogenous content will be taken forward for in-solution capture enrichment or in-depth shotgun sequencing. Pre-library laboratory work will be conducted at the aDNA facilities in the Department of Archaeology, Durham University and post-library work at the modern DNA facilities in the same department and at Newcastle University. Standard methods for population structure and admixture inference will be followed, including PCA (10), f3/f4 statistics, qpAdm and qpwave (11), and biological kinship will be reconstructed with READv2 (12). New ad-hoc methods will be devised and applied, if relevant. The project will be developed in collaboration with Prof. Antonio Faustino Carvalho, U. Algarve, and Dr. Cristina Tejedor Rodriguez, U. Valladolid. They will provide access to archaeological material as well as detailed contextual information relating to chronology, funerary practices and site formation processes. They will play a key role in the interpretation of results, contributing specialist expertise on the archaeological context of the studied assemblages and supporting the integration of genetic data with archaeological evidence.
Project Timeline
Year 1
-Months 1-2: Sample acquisition and photographic documentation. Laboratory training.
-Months 3-12: DNA extraction, library preparation, shotgun sequencing screening. Assessment of aDNA quality and content.
Year 2
– Months 12-18: Human and pathogen DNA capture enrichment and sequencing. Training in human population genetic analyses. Building of comparative human genomic database. Start human population genetic analyses.
– Months 18-24: Complete human population genetic analyses. Training in pathogen phylogenomics. Building comparative pathogen dataset(s). Start pathogen phylogenomic analyses. Draft paper 1.
Year 3
-Months 24-30: Complete pathogen phylogenomic analyses. Write thesis methods. Finish and submit paper 1.
-Months 30-36: Write thesis introduction and results. Conference presentation. Draft paper 2.
Year 3.5
-Months 36-42: Complete thesis writing and submission. Finish and submit paper 2.
The project timeline may require adjustment depending on the DNA preservation in the samples.
Training
& Skills
The student will be responsible for both the laboratory and bioinformatic components of the project, including the handling, sampling, and documentation of archaeological human remains. DNA laboratory skills and knowledge of the Unix command line are essential for this project and due to the analytical complexity of the project, priority will be given to applicants with previous demonstrable experience in any of these areas. Additional training will be provided on human population and pathogen genomics by the supervisors and through public courses, including those offered by the Advanced Research Computing group at Durham University.
References & further reading
1. Allentoft, M. E. et al. Population genomics of post-glacial western Eurasia. Nature 625, 301–311 (2024).
2. Sánchez-Quinto, F. et al. Megalithic tombs in western and northern Neolithic Europe were linked to a kindred society. Proc Natl Acad Sci U S A 116, 9469–9474 (2019).
3. Carvalho, A. F. et al. Hunter-gatherer genetic persistence at the onset of megalithism in western Iberia: New mitochondrial evidence from Mesolithic and Neolithic necropolises in central-southern Portugal. Quaternary International
4. Laidlaw, G. Mitochondrial and Metagenomic Analysis of Iberian Individuals from the Neolithic and Chalcolithic Periods. (Durham University, Durham, 2025).
5. Dabney, J. et al. Complete mitochondrial genome sequence of a Middle Pleistocene cave bear reconstructed from ultrashort DNA fragments. PNAS 201314445 (2013)
6. Kircher, M., Sawyer, S. & Meyer, M. Double indexing overcomes inaccuracies in multiplex sequencing on the Illumina platform. Nucleic Acids Res 40, e3–e3 (2012).
7. Meyer, M. & Kircher, M. Illumina Sequencing Library Preparation for Highly Multiplexed Target Capture and Sequencing. Cold Spring Harbor Protocols 2010, pdb.prot5448-pdb.prot5448 (2010).
8. Fellows Yates, J. A. et al. Reproducible, portable, and efficient ancient genome reconstruction with nf-core/eager. PeerJ 9, e10947 (2021).
9. Pochon, Z. et al. aMeta: an accurate and memory-efficient ancient metagenomic profiling workflow. Genome Biology 24, 242 (2023).
10. Patterson, N., Price, A. L. & Reich, D. Population Structure and Eigenanalysis. PLOS Genetics 2, e190 (2006).
11. Patterson, N. et al. Ancient admixture in human history. Genetics 192, 1065–1093 (2012).
12. Alaçamlı, E. et al. READv2: advanced and user-friendly detection of biological relatedness in archaeogenomics. Genome Biology 25, 216 (2024).
