Abstract
Scandinavia was one of the last geographic areas in Europe to become habitable for humans after the last glaciation. However, the origin(s) of the first colonizers and their migration routes remain unclear. We sequenced the genomes, up to 57x coverage, of seven hunter-gatherers excavated across Scandinavia and dated to 9,500-6,000 years before present. Surprisingly, among the Scandinavian Mesolithic individuals, the genetic data display an east-west genetic gradient that opposes the pattern seen in other parts of Mesolithic Europe. This result suggests that Scandinavia was initially colonized following two different routes: one from the south, the other from the northeast. The latter followed the ice-free Norwegian north Atlantic coast, along which novel and advanced pressure-blade stone-tool techniques may have spread. These two groups met and mixed in Scandinavia, creating a genetically diverse population, which shows patterns of genetic adaptation to high latitude environments. These adaptations include high frequencies of low pigmentation variants and a gene-region associated with physical performance, which shows strong continuity into modern-day northern Europeans. Finally, we were able to compute a 3D facial reconstruction of a Mesolithic woman from her high-coverage genome, giving a glimpse into an individual’s physical appearance in the Mesolithic.
Main text
As the ice-sheet retracted from northern Europe after the Last Glacial Maximum (LGM), around 23,000 years ago, new habitable areas emerged (1) allowing plants (2, 3) and animals (4) to recolonize the Scandinavian peninsula (hereafter Scandinavia). There is consistent evidence of human presence in the archaeological record from c. 11,700 years before present (BP), both in southern and northern Scandinavia (5–8). At this time, the ice-sheet was still dominating the interior of Scandinavia (8) (Fig. 1A, Supplementary Information 1), but recent climate modeling shows that the Arctic coast of (modern-day) northern Norway was ice-free (9). Similarities in late-glacial lithic technology (direct blade percussion technique) of western Europe and the oldest counterparts of northernmost Scandinavia (10) (Supplementary Information 1) have been used to argue for a postglacial colonization of Scandinavia from southwestern Europe. However, studies of a new lithic technology, ‘pressure blade’ technique, which first occurred in the northern parts of Scandinavia, indicates contacts with groups in the east and possibly an eastern origin of the colonizers (6, 11, 12) (Supplementary Information 1). The first genetic studies of Mesolithic human remains from central and eastern Scandinavia (SHGs) revealed similarities to two different Mesolithic European populations, the ‘western hunter-gatherers’ (WHGs) from western, central and southern Europe and the ‘eastern hunter-gatherers’ (EHGs) from northeastern Europe (13–19). Archaeology, climate modeling, and genetics, suggest several possibilities for the colonization of Scandinavia, including migrations from the south, southeast, northeast and combinations of these, however, the early post-glacial peopling of Scandinavia remains elusive (1–12, 14–17, 20, 21). In this study, we contrast genome sequence data and stable isotopes from Mesolithic human remains from western, northern, and eastern Scandinavia to infer the post-glacial colonization of Scandinavia – from where people came, what routes they followed, how they were related to other Mesolithic Europeans – and to investigate human adaptation to high-latitude environments.
Results and Discussion
We sequenced the genomes of seven hunter-gatherers from Scandinavia (Table 1 and Supplementary Information 1-3) ranging from 57.8× to 0.1× genome coverage, of which four individuals had a genome coverage above 1×. The remains were directly dated to between 9,500 BP and 6,000 BP, and were excavated in southwestern Norway (Hum1, Hum2), northern Norway (Steigen), and the Baltic islands of Stora Karlsö and Gotland (SF9, SF11, SF12 and SBj) and represent 18% (6 of 33) of all known human remains in Scandinavia older than 8,000 (22). All samples displayed fragmentation and cytosine deamination at fragment termini characteristic for ancient DNA (Supplementary Information 3). Mitochondrial (mt) DNA-based contamination estimates were <6% for all individuals and autosomal contamination was <1% for all individuals except for SF11, which showed c. 10% contamination (Table 1, Supplementary Information 4). Four of the seven individuals were inferred to be males, three were females. All the western and northern Scandinavian individuals and one eastern Scandinavian carried U5a1 mitochondrial haplotypes while the remaining eastern Scandinavians carried U4a haplotypes (Table 1, Supplementary Information 5). These individuals represent the oldest U5a1 and U4 lineages detected so far. The Y chromosomal haplotype was determined for three of the four males, all carried I2 haplotypes, which were common in pre-Neolithic Europe (Table 1, Supplementary Information 5).
The high coverage and Uracil-DNA-glycosylase (UDG) treated genome (to reduce the effects of post-mortem DNA damage) of SF12 allowed us to confidently discover new and hitherto unknown variants at sites with 55x or higher sequencing depth (Supplementary Information 3). Based on SF12’s high-coverage and high-quality genome, we estimate the number of single nucleotide polymorphisms (SNPs) hitherto unknown (that are not recorded in dbSNP (v142)) to be c. 10,600. This is almost twice the number of unique variants (c. 6,000) per Finnish individual (Supplementary Information 3) and close to the median per European individual in the 1000 Genomes Project (23) (c. 11,400, Supplementary Information 3). At least 17% of these SNPs that are not found in modern-day individuals, were in fact common among the Mesolithic Scandinavians (seen in the low coverage data conditional on the observation in SF12), suggesting that a substantial fraction of human variation has been lost in the past 9,000 years (Supplementary Information 3). In other words, the SHGs (as well as WHGs and EHGs) have no direct descendants, or a population that show direct continuity with the Mesolithic populations (Supplementary Information 6) (13–17). Thus, many genetic variants found in Mesolithic individuals have not been carried over to modern-day groups. Among the novel variants in SF12, four (all heterozygous) are predicted to affect the function of protein coding genes (24) (Supplementary Information 3). The ‘heat shock protein’ HSPA2 in SF12 carries an unknown mutation that changes the amino acid histidine to tyrosine at a protein-protein interaction site, 107which likely disrupts the function of the protein (Supplementary Information 3). Defects in HSPA2 are known to drastically reduce fertility in males (25). Although SF12 herself would not be affected by this variant, her male offspring could carry the reduced fertility variant, and it will be interesting to see how common this variant was among Mesolithic groups as more genome sequence data become available. The high-quality diploid genotype calls further allowed us to genetically predict physical appearance, including pigmentation, and to use a model-based approach trained on modern-day faces and genotypes (26) to create a 3D model of SF12’s face (Supplementary Information 9). This represents a new way of reconstructing an ancient individual’s facial appearance from genetic information, which is especially informative in cases such as for SF12, where only post-cranial fragments were available, and future archaeogenetic studies will have the potential to many individuals appearance from past times.
Demographic history of Mesolithic Scandinavians
In order to compare the genomic data of the seven SHGs to genetic information from other ancient individuals and modern-day groups, data was merged with six published Mesolithic individuals from Motala in central Scandinavia, 47 published Upper Paleolithic, Mesolithic and Early Neolithic individuals from other parts of Eurasia (Supplementary Information 6), as well as with a world-wide set of 203 modern-day populations (15, 23). All 13 SHGs – regardless of geographic sampling location and age – display genetic affinities to both WHGs and EHGs (Fig. 1A, B, Supplementary Information 6). This is consistent with a scenario in which SHGs represent a mixed group tracing parts of their ancestry to both the WHGs and the EHGs (14–16, 19, 27).
To investigate the postglacial colonization of Scandinavia, we explored four hypothetical migration routes (primarily based on natural geography) linked to WHGs and EHGs, respectively (Supplementary Information 11); a) a migration of WHGs from the south, b) a migration of EHGs from the east across the Baltic Sea, c) a migration of EHGs from the east and along the north-Atlantic coast, d) a migration of EHGs from the east and south of the Baltic Sea, and combinations of these four migration routes. These scenarios allow us to formulate expected genetic affinities for northern, western, eastern, and central SHGs (Supplementary Information 11). The SHGs from northern and western Scandinavia show a distinct and significantly stronger affinity to the EHGs compared to the central and eastern SHGs (Fig. 1). Conversely, the SHGs from eastern and central Scandinavia were genetically more similar to WHGs compared to the northern and western SHGs (Fig. 1). Using a model-based approach (15, 16), the EHG genetic component of northern and western SHGs was estimated to 55% on average (43-67%) and significantly different (Wilcoxon test, p=0.014) from the average 35% (22-44%) in eastern and south-central SHGs. This average is similar to eastern Baltic hunter-gatherers from Latvia (28) (average 33%, Fig. 1A, Supplementary Information 6). These patterns of genetic affinity within SHGs are in direct contrast to the expectation based on geographic proximity with EHGs and WHGs and do not correlate with age of the sample (Supplementary Information 11).
The archaeological record in Scandinavia shows early evidence of human presence in northern coastal Atlantic areas (12). Stable isotope analysis of northern and western SHGs revealed an extreme marine diet, suggesting a maritime subsistence, in contrast to the more mixed terrestrial/aquatic diet of eastern and central SHGs (Supplementary Information 1). Combining these isotopic results with the patterns of genetic variation, we suggest an initial colonization from the south, likely by WHGs. A second migration of people who were related to the EHGs – that brought the new pressure blade technique to Scandinavia and that utilized the rich Atlantic coastal marine resources –entered from the northeast moving southwards along the ice-free Atlantic coast where they encountered WHG groups. The admixture between the two colonizing groups created the observed pattern of a substantial EHG component in the northern and the western SHGs, contrary to the higher levels of WHG genetic component in eastern and central SHGs (Fig. 1, Supplementary Information 11).
By sequencing complete ancient genomes, we can compute unbiased estimates of genetic diversity, which are informative of past population sizes and population history. Here, we restrict the analysis to WHGs and SHGs, since only SNP capture data is available for EHGs (Supplementary Information 7). In current-day Europe, there is greater genetic diversity in the south compared to the north. During the Mesolithic, by contrast, we find higher levels of genetic diversity (Supplementary Information 7) as well as lower levels of runs of homozygosity (Fig. 2A) and linkage disequilibrium (Fig. 2B) in SHGs compared to WHGs (represented by Loschbour and Bichon, (15, 29)) and Caucasus hunter-gatherers (CHG, represented by Kotias and Satsurblia, (29)). Using a sequential-Markovian-coalescent approach (30) for the high-coverage, high quality genome of SF12, we find that right before the SF12 individual lived, the effective population size of SHGs was similar to that of WHGs (Fig. 2C). At the time of the LGM and back to c. 50,000 years ago, both the WHGs and SHGs go through a bottleneck, but the ancestors of SHGs retained a greater population size in contrast to the ancestors of WHGs who went through a more severe bottleneck (Fig. 2c). Around 50,000-70,000 years ago, the effective population sizes of the ancestors of SHGs, WHGs, Neolithic groups (represented by Stuttgart (15)) and Paleolithic Eurasians (represented by Ust-Ishim (31)) align, suggesting that these diverse groups all trace their ancestry back to a common ancestral group which likely represents the early migrants out-of-Africa, who likely share a common ancestry outside of Africa.
Adaptation to high-latitude environments
With the aim of detecting signs of adaptation to high-latitude environments and selection during and after the Mesolithic, we employed three different approaches that utilize the Mesolithic genomic data. In the first approach, we assumed that SHGs adapted to high-latitude environments of low temperatures and seasonally low levels of light, and searched for gene variants that carried over to modern-day people in northern Europe. As we have already noted, modern-day northern Europeans trace limited amount of genetic material back to the SHGs (due to the many additional migrations during later periods), and any genomic region that displays extraordinary genetic continuity would be a strong candidate for adaptation in people living in northern Europe across time. We designed a statistic, Dsel (Supplementary Information 10), that captures this specific signal and scanned the whole genome for gene-variants that show strong continuity (little differentiation) between SHGs and modern-day northern Europeans while exhibiting large differentiation to modern-day southern European populations (32) (Fig. 3A; Supplementary Information 10). Six of the top ten SNPs with greatest Dsel values were located in the TMEM131 gene that has been found to be associated with physical performance (33), which could make it part of the physiological adaptation to cold (34). This genomic region was more than 200kbp long and showed the strongest haplotypic differentiation between modern-day Tuscans and Finns (Supplementary Information 10). The particular haplotype was relatively common in SHGs, it is even more common among today’s Finnish population (Supplementary Information 10), and showed a strong signal of local adaptation (Supplementary Information 10). Other top hits included genes associated with a wide range of metabolic, cardiovascular, developmental and psychological traits (Supplementary Information 10) potentially linked to physiological (34).
In addition to performing this genome-wide scan, we studied the allele frequencies in three pigmentation genes (SLC24A5, SLC45A2, having a strong effect on skin pigmentation, and OCA2/HERC2, having a strong effect on eye pigmentation) where the derived alleles are virtually fixed in northern Europeans today. The differences in allele frequencies of those three loci are among the highest between human populations, suggesting that selection was driving the differences in eye color, skin and hair pigmentation as part of the adaptation to different environments (35–37). The SHGs show a combination of eye and skin pigmentation that was unique in Mesolithic Europe, with light skin pigmentation and varied blue to light-brown eye color. This is strikingly different from the WHGs – who have been found to have the specific combination of blue-eyes and dark-skin (15, 17, 18) (Fig. 3B) – and EHGs – who have been suggested to be brown eyed and light-skinned (16, 17) (Fig. 3B). The unique configuration of the SHGs is not fully explained by the fact that SHGs are a mixture of EHGs and WHGs as the frequencies of the blue-eye and one light-skin variant are significantly higher in SHGs than expected from their genome-wide admixture proportions (Fig. 3B, Supplementary Information 10). This could be explained by a continued increase of the allele frequencies after the admixture event, likely caused by adaptation to high-latitude environments (35, 37).
Conclusion
By combining information from climate modeling, archaeology and Mesolithic human genomes, we were able to reveal the complexity of the early colonization process of Scandinavia and human adaptation to high-latitude environments. We disentangled migration routes and linked them to particular archaeological patterns, demonstrate greater genetic diversity in northern Europe compared to southern Europe – in contrast to modern-day patterns – and show that many genetic variants that were common in the Mesolithic have been lost today. These finds reiterate the importance of human migration for dispersal of novel technology in human prehistory (13–16, 21, 27, 38–45) and the many partial population turnovers in our past.
Materials and Methods
All samples were processed in designated clean labs and sequenced on Illumina HiSeq machines. Sequences were mapped to the human reference genome. More details on the data processing and the population genomic analyses can be found in Supplementary Information.
Acknowledgements
We thank Rachel Howcroft, and Nicci Arosén for help with isotope laboratory work and Heike Siegmund at the Stable Isotope Laboratory, Stockholm University, Lena Ideström at Gotlands museum and Sabine Sten at Uppsala University Campus Gotland in assisting in sampling the Stora Bjers material, Leena Drenzel at The Swedish History Museum for assistance with the Stora Förvar material, and the Norwegian Maritime Museum for excavating Hummervikholmen. MS and PC thank the participants who volunteered for face reconstruction methods development. This project was supported by grants from Riksbankens Jubileumsfond (to AG, MJ, and JS), Knut and Alice Wallenberg foundation (to MJ, JS, AG), the Swedish Research council, no. 2013-1905 (to AG, MJ, and JS) and no. 421-2013-730 (to JA and JS), an European Research Council Starting Grant (to MJ), a Wenner-Gren foundations postdoctoral fellowship (to TG), and Berit Wallenberg foundation (to MF). Sequencing was performed at the National Genomics Infrastructure (NGI) Uppsala and computations were performed at Uppsala Multidisciplinary Center for Advanced Computational Science (UPPMAX).