Ancient DNA Research Articles

Cervidae antlers exploited to manufacture prehistoric tools and hunting implements as a reliable source of ancient DNA

Antler is one of the primary animal raw materials exploited for technical purposes by the hunter-gatherer groups of the Eurasian Upper Palaeolithic (UP) all over the ecological range of deers, and beyond. It was exhaustively employed to produce one of the most critical tools for the survival of the UP societies: hunting weapons. However, antler implements can be made from diverse deer taxa, with different ecological requirements and ethological behaviours. Identifying the antler's origin at a taxonomic level is thus essential in improving our knowledge of humans' functional, practical and symbolic choices, as well as the human-animal interface during Prehistoric times. Nevertheless, palaeogenetics analyses have focused mainly on bone and teeth, with genetic studies of antler generally focused on modern deer conservation. Here we present the results of the first whole mitochondrial genome ancient DNA (aDNA) analysis by means of in-solution hybridisation capture of antlers from pre-Holocene archaeological contexts. We analysed a set of 50 Palaeolithic and Neolithic (c. 34-8ka) antler and osseous objects from South-Western Europe, Central Europe, South-Western Asia and the Caucasus. We successfully obtained aDNA, allowing us to identify the exploited taxa and demonstrate the archaeological relevance of those finds. Moreover, as most of the antlers were sampled using a minimally-invasive method, further analyses (morphometric, technical, genetic, radiometric and more) remain possible on these objects.

A nuclear genome assembly of an extinct flightless bird, the little bush moa.

We present a draft genome of the little bush moa (Anomalopteryx didiformis)-one of approximately nine species of extinct flightless birds from Aotearoa, New Zealand-using ancient DNA recovered from a fossil bone from the South Island. We recover a complete mitochondrial genome at 249.9× depth of coverage and almost 900 megabases of a male moa nuclear genome at ~4 to 5× coverage, with sequence contiguity sufficient to identify more than 85% of avian universal single-copy orthologs. We describe a diverse landscape of transposable elements and satellite repeats, estimate a long-term effective population size of ~240,000, identify a diverse suite of olfactory receptor genes and an opsin repertoire with sensitivity in the ultraviolet range, show that the wingless moa phenotype is likely not attributable to gene loss or pseudogenization, and identify potential function-altering coding sequence variants in moa that could be synthesized for future functional assays. This genomic resource should support further studies of avian evolution and morphological divergence.

Complex genetic landscape revealed by a population in the eastern Tianshan Mountains of Xinjiang between the 8th and 11th centuries CE

AbstractThe eastern Tianshan Mountain (ETS) region in Xinjiang, China, has played a crucial role as a major highway for large‐scale human migrations and technological exchanges across Eurasia throughout history. Understanding the genetic makeup of past populations holds significant importance in comprehending human mobility in this region. Here, we report genome‐wide ancient DNA for the two individuals whose origins remain uncertain, found at the Dahe ancient city site in the ETS region. Our results indicate that although both individuals display a higher genetic affinity to Asian populations, one individual shows some genetic sharing with South Asians, while the other shares more affinity with populations from Northeastern Asia. Radiocarbon dating shows that the two human remains were not deposited simultaneously. Overall, the combined genetic, physical anthropology and radiocarbon evidence strongly suggests that the ETS region, situated at the heart of the Silk Road, has been a crossroads of diverse populations coexisting between the 8th and 11th centuries. Additionally, the genetic ancestry sharing with the Bronze Age Tarim mummies suggested a long‐term residual of indigenous ancestry in Xinjiang and showed that autochthonous deep Asian ancestry had left their genetic legacy into the historical populations of Xinjiang despite a spanning of over three millennia.

Evaluation of a New DNA Extraction Method on Challenging Bone Samples Recovered from a WWII Mass Grave.

Bones and teeth represent a common finding in ancient DNA studies and in forensic casework, even after a long burial. Genetic typing is the gold standard for the personal identification of skeletal remains, but there are two main factors involved in the successful DNA typing of such samples: (1) the set-up of an efficient DNA extraction method; (2) the identification of the most suitable skeletal element for the downstream genetic analyses. In this paper, a protocol based on the processing of 0.5 g of bone powder decalcified using Na2EDTA proved to be suitable for a semi-automated DNA extraction workflow using the Maxwell® FSC DNA IQ™ Casework Kit (Promega, Madison, WI, USA). The performance of this method in terms of DNA recovery and quality was compared with a full demineralisation extraction protocol based on Qiagen technology and kits. No statistically significant differences were scored according to the DNA recovery and DNA degradation index (p-values ≥ 0.176; r ≥ 0.907). This new DNA extraction protocol was applied to 88 bone samples (41 femurs, 19 petrous bones, 12 metacarpals and 16 molars) allegedly belonging to 27 World War II Italian soldiers found in a mass grave on the isle of Cres (Croatia). The results of the qPCR performed by the Quantifiler Human DNA Quantification kit showed values above the lowest Limit of Quantification (lLOQ; 23 pg/µL) for all petrous bones, whereas other bone types showed, in most cases, lower amounts of DNA. Replicate STR-CE analyses showed successful typing (that is, >12 markers) in all tests on the petrous bones, followed by the metacarpals (83.3%), femurs (52.2%) and teeth (20.0%). Full profiles (22/22 autosomal markers) were achieved mainly in the petrous bones (84.2%), followed by the metacarpals (41.7%). Stochastic amplification artefacts such as drop-outs or drop-ins occurred with a frequency of 1.9% in the petrous bones, whereas they were higher when the DNA recovered from other bone elements was amplified (up to 13.9% in the femurs). Overall, the results of this study confirm that petrous bone outperforms other bone elements in terms of the quantity and quality of the recovered DNA; for this reason, if available, it should always be preferred for genetic testing. In addition, our results highlight the need for accurate planning of the DVI operation, which should be carried out by a multi-disciplinary team, and the tricky issue of identifying other suitable skeletal elements for genetic testing. Overall, the results presented in this paper support the need to adopt preanalytical strategies positively related to the successful genetic testing of aged skeletal remains in order to reduce costs and the time of analysis.

The Dogs of Tsenacomoco: Ancient DNA Reveals the Presence of Local Dogs at Jamestown Colony in the Early Seventeenth Century

Abstract Multiple studies have demonstrated that European colonization of the Americas led to the death of nearly all North American dog mitochondrial lineages and replacement with European ones sometime between AD 1492 and the present day. Historical records indicate that colonists imported dogs from Europe to North America, where they became objects of interest and exchange as early as the seventeenth century. However, it is not clear whether the earliest archaeological dogs recovered from colonial contexts were of European, Indigenous, or mixed descent. To clarify the ancestry of dogs from the Jamestown Colony, Virginia, we sequenced ancient mitochondrial DNA from six archaeological dogs from the period 1609–1617. Our analysis shows that the Jamestown dogs have maternal lineages most closely associated with those of ancient Indigenous dogs of North America. Furthermore, these maternal lineages cluster with dogs from Late Woodland, Hopewell, and Virginia Algonquian archaeological sites. Our recovery of Indigenous dog lineages from a European colonial site suggests a complex social history of dogs at the interface of Indigenous and European populations during the early colonial period.

Ancient genomes revealed the complex human interactions of the ancient western Tibetans

The western Tibetan Plateau is the crossroad between the Tibetan Plateau, Central Asia, and South Asia, and it is a potential human migration pathway connecting these regions. However, the population history of the western Tibetan Plateau remains largely unexplored due to the lack of ancient genomes covering a long-time interval from this area. Here, we reported genome-wide data of 65 individuals dated to 3,500-300 years before present (BP) in the Ngari prefecture. The ancient western Tibetan Plateau populations share the majority of their genetic components with the southern Tibetan Plateau populations and have maintained genetic continuity since 3,500 BP while maintaining interactions with populations within and outside the Tibetan Plateau. Within the Tibetan Plateau, the ancient western Tibetan Plateau populations were influenced by the additional expansion from the south to the southwest plateau before 1,800 BP. Outside the Tibetan Plateau, the western Tibetan Plateau populations interacted with both South and Central Asian populations at least 2,000 years ago, and the South Asian-related genetic influence, despite being very limited, was from the Indus Valley Civilization (IVC) migrants in Central Asia instead of the IVC populations from the Indus Valley. In light of the new genetic data, our study revealed the complex population interconnections across and within the Tibetan Plateau.

Phylogenetic insights into the genetic legacies of Hungarian-speaking communities in the Carpathian Basin

This study focuses on exploring the uniparental genetic lineages of Hungarian-speaking minorities residing in rural villages of Baranja (Croatia) and the Zobor region (Slovakia). We aimed to identify ancestral lineages by examining genetic markers distributed across the entire mitogenome and on the Y-chromosome. This allowed us to discern disparities in regional genetic structures within these communities. By integrating our newly acquired genetic data from a total of 168 participants with pre-existing Eurasian and ancient DNA datasets, our goal was to enrich the understanding of the genetic history trajectories of Carpathian Basin populations. Our findings suggest that while population-based analyses may not be sufficiently robust to detect fine-scale uniparental genetic patterns with the sample sizes at hand, phylogenetic analysis of well-characterized Y-chromosomal Short Tandem Repeat (STR) data and entire mitogenome sequences did uncover multiple lineage ties to far-flung regions and eras. While the predominant portions of both paternal and maternal DNA align with the East-Central European spectrum, rarer subhaplogroups and lineages have unveiled ancient ties to both prehistoric and historic populations spanning Europe and Eastern Eurasia. This research augments the expansive field of phylogenetics, offering critical perspectives on the genetic constitution and heritage of the communities in East-Central Europe.

New Canary Islands Roman mediated settlement hypothesis deduced from coalescence ages of curated maternal indigenous lineages

Numerous genetic studies have contributed to reconstructing the human history of the Canary Islands population. The recent use of new ancient DNA targeted enrichment and next-generation sequencing techniques on new Canary Islands samples have greatly improved these molecular results. However, the bulk of the available data is still provided by the classic mitochondrial DNA phylogenetic and phylogeographic studies carried out on the indigenous, historical, and extant human populations of the Canary Islands. In the present study, making use of all the accumulated mitochondrial information, the existence of DNA contamination and archaeological sample misidentification in those samples is evidenced. Following a thorough review of these cases, the new phylogeographic analysis revealed the existence of a heterogeneous indigenous Canarian population, asymmetrically distributed across the various islands, which most likely descended from a unique mainland settlement. These new results and new proposed coalescent ages are compatible with a Roman-mediated arrival driven by the exploitation of the purple dye manufacture in the Canary Islands.

Human ancient DNA analysis in Latin America: Current state and challenges

AbstractAncient DNA analysis has greatly contributed to understanding the population history of several species. In the last 20 years, the field has undergone an important transformation: particularly in our species, thousands of ancient genomes have been analyzed worldwide, providing evidence of population movements and interactions through time. However, several researchers have raised concerns about the way the field is developing and how collaborations are being established. In this work, we describe and evaluate the situation in Latin America, considering both what we have learned about population history through paleogenetics and how it has developed in the region.

Identification of the 18 World War II executed citizens of Adele, Rethymnon, Crete using an ancient DNA approach and low coverage genomes

In the Battle of Crete during the World War II occupation of Greece, the German forces faced substantial civilian resistance. To retribute the numerous German losses, a series of mass executions took place in numerous places in Crete; a common practice reported from Greece and elsewhere. In Adele, a village in the regional unit of Rethymnon, 18 male civilians were executed and buried in a burial pit at the Sarakina site. In this study, the first one conducted for a conflict that occurred in Greece, we identified for humanitarian purposes the 18 skulls of the Sarakina victims, following a request from the local community of Adele. The molecular identification of historical human remains via ancient DNA approaches and low coverage whole genome sequencing has only recently been introduced. Here, we performed genome skimming on the living relatives of the victims, as well as high throughput historical DNA analysis on the skulls to infer the kinship degrees among the victims via genetic relatedness analyses. We also conducted targeted anthropological analysis to successfully complete the identification of all Sarakina victims. We demonstrate that our methodological approach constitutes a potentially highly informative forensic tool to identify war victims. It can hence be applied to analogous studies on degraded DNA, thus, paving the path for systematic war victim identification in Greece and beyond.

The genomes of seven economic Caesalpinioideae trees provide insights into polyploidization history and secondary metabolite biosynthesis

The Caesalpinioideae subfamily contains many well-known trees that are important for economic sustainability and human health, but a lack of genomic resources has hindered their breeding and utilization. Here, we present chromosome-level reference genomes for the two food and industrial trees Gleditsia sinensis (921 Mb) and Biancaea sappan (872 Mb), the three shade and ornamental trees Albizia julibrissin (705 Mb), Delonix regia (580 Mb), and Acacia confusa (566 Mb), and the two pioneer and hedgerow trees Leucaena leucocephala (1338 Mb) and Mimosa bimucronata (641 Mb). Phylogenetic inference shows that the mimosoid clade has a much higher evolutionary rate than the other clades of Caesalpinioideae. Macrosynteny comparison suggests that the fusion and breakage of an unstable chromosome are responsible for the difference in basic chromosome number (13 or 14) for Caesalpinioideae. After an ancient whole-genome duplication (WGD) shared by all Caesalpinioideae species (CWGD, ∼72.0 million years ago [MYA]), there were two recent successive WGD events, LWGD-1 (16.2–19.5 MYA) and LWGD-2 (7.1–9.5 MYA), in L. leucocephala. Thereafter, ∼40% gene loss and genome-size contraction have occurred during the diploidization process in L. leucocephala. To investigate secondary metabolites, we identified all gene copies involved in mimosine metabolism in these species and found that the abundance of mimosine biosynthesis genes in L. leucocephala largely explains its high mimosine production. We also identified the set of all potential genes involved in triterpenoid saponin biosynthesis in G. sinensis, which is more complete than that based on previous transcriptome-derived unigenes. Our results and genomic resources will facilitate biological studies of Caesalpinioideae and promote the utilization of valuable secondary metabolites.

PNPLA3 fatty liver allele was fixed in Neanderthals and segregates neutrally in humans

ObjectiveFat deposition is modulated by environmental factors and genetic predisposition. Genome-wide association studies identified PNPLA3 p.I148M (rs738409) as a common variant that increases risk of developing liver steatosis. When and...

Ancient genome of Empress Ashina reveals the Northeast Asian origin of Göktürk Khanate

Xiao‐Min Yang, Hai‐Liang Meng, Jian‐Lin Zhang, Yao Yu, Edward Allen, Zi‐Yang Xia, Kong‐Yang Zhu, Pan‐Xin Du, Xiao‐Ying Ren, Jian‐Xue Xiong, Xiao‐Yu Lu, Yi Ding, Sheng Han, Wei‐Peng Liu, Li Jin, Chuan‐Chao Wang, and Shao‐Qing Wen. Ancient genome of Empress Ashina reveals the Northeast Asian origin of Göktürk Khanate. – Journal of Systematic and Evolution. Institute of Botany, Chinese Academy of Sciences. 2023. Volume 00 (0) 1-9.

Uncovering Holocene climate fluctuations and ancient conifer populations: Insights from a high-resolution multi-proxy record from Northern Finland

A series of abrupt climate events linked to circum-North Atlantic meltwater forcing have been recognised in Holocene paleoclimate data. To address the paucity of proxy records able to characterise robustly the regional impacts of these events, we retrieved a sub-centennial resolution, well-dated core sequence from Lake Kuutsjärvi, northeast Finland. By analysing a range of paleo-environmental proxies (pollen, plant sedimentary ancient DNA, plant macrofossils, conifer stomata, and non-pollen palynomorphs), and supported with proxy-based paleotemperature and moisture reconstructions, we unravel a well-defined sequence of vegetation and climate dynamics over the early-to-middle Holocene. The birch-dominated pioneer vegetation stage was intersected by two transient tree-cover decrease events at 10.4 and 10.1 thousand years ago (ka), likely representing a two-pronged signal of the 10.3 ka climate event. Our data also show a clear signal of the 8.2 ka climate event, previously not well recorded in the European Arctic, with a collapse of the pine-birch forest and replacement by juniper developing in tight synchrony with Greenland isotopic proxies over 8.4–8.0 ka. Supported by climate modelling, severe winter cooling rather than summer might have been driving vegetation disruptions in the early Holocene. The Kuutsjärvi data indicate an early arrival of Norway spruce (Picea abies) by 9.2 ka (pollen, DNA, and stoma finds), as well as the first evidence for Holocene presence of larch (Larix) in Finland, with pollen finds dating to 9.6–5.9 ka.

Ancient DNA study of canid remains excavated from the Houtaomuga archaeological site, Northeast China

Ancient DNA study of canid remains excavated from the Houtaomuga archaeological site, Northeast China

Advancing responsible genomic analyses of ancient mollusc shells.

The analysis of the DNA entrapped in ancient shells of molluscs has the potential to shed light on the evolution and ecology of this very diverse phylum. Ancient genomics could help reconstruct the responses of molluscs to past climate change, pollution, and human subsistence practices at unprecedented temporal resolutions. Applications are however still in their infancy, partly due to our limited knowledge of DNA preservation in calcium carbonate shells and the need for optimized methods for responsible genomic data generation. To improve ancient shell genomic analyses, we applied high-throughput DNA sequencing to 27 Mytilus mussel shells dated to ~111-6500 years Before Present, and investigated the impact, on DNA recovery, of shell imaging, DNA extraction protocols and shell sub-sampling strategies. First, we detected no quantitative or qualitative deleterious effect of micro-computed tomography for recording shell 3D morphological information prior to sub-sampling. Then, we showed that double-digestion and bleach treatment of shell powder prior to silica-based DNA extraction improves shell DNA recovery, also suggesting that DNA is protected in preservation niches within ancient shells. Finally, all layers that compose Mytilus shells, i.e., the nacreous (aragonite) and prismatic (calcite) carbonate layers, with or without the outer organic layer (periostracum) proved to be valuable DNA reservoirs, with aragonite appearing as the best substrate for genomic analyses. Our work contributes to the understanding of long-term molecular preservation in biominerals and we anticipate that resulting recommendations will be helpful for future efficient and responsible genomic analyses of ancient mollusc shells.

East and Southeast Asian hominin dispersal and evolution: A review

East and Southeast Asia have served as significant habitats for diverse hominins for more than one million years. This has been demonstrated by numerous studies over the past decade that have reported new discoveries, ages, morphological evidence, and ancient biomolecules from fossils. This has revolutionized our understanding of their evolution and dispersal. However, the existing literature lacks a comprehensive overview that combines insights from different scientific disciplines that are needed to address the still-uncertain or contentious aspects of this field. Here, we provide a synthesis of the timing and distribution of the different hominins that lived in East and Southeast Asia. We then review two biomolecular methods, ancient proteomics and sedimentary ancient DNA, which hold great promise for revealing hominin history in this area.

A 1 Ma sedimentary ancient DNA (sedaDNA) record of catchment vegetation changes and the developmental history of tropical Lake Towuti (Sulawesi, Indonesia).

Studying past ecosystems from ancient environmental DNA preserved in lake sediments (sedaDNA) is a rapidly expanding field. This research has mainly involved Holocene sediments from lakes in cool climates, with little known about the suitability of sedaDNA to reconstruct substantially older ecosystems in the warm tropics. Here, we report the successful recovery of chloroplast trnL (UAA) sequences (trnL-P6 loop) from the sedimentary record of Lake Towuti (Sulawesi, Indonesia) to elucidate changes in regional tropical vegetation assemblages during the lake's Late Quaternary paleodepositional history. After the stringent removal of contaminants and sequence artifacts, taxonomic assignment of the remaining genuine trnL-P6 reads showed that native nitrogen-fixing legumes, C3 grasses, and shallow wetland vegetation (Alocasia) were most strongly associated with >1-million-year-old (>1 Ma) peats and silts (114-98.8 m composite depth; mcd), which were deposited in a landscape of active river channels, shallow lakes, and peat-swamps. A statistically significant shift toward partly submerged shoreline vegetation that was likely rooted in anoxic muddy soils (i.e., peatland forest trees and wetland C3 grasses (Oryzaceae) and nutrient-demanding aquatic herbs (presumably Oenanthe javanica)) occurred at 76 mcd (~0.8 Ma), ~0.2 Ma after the transition into a permanent lake. This wetland vegetation was most strongly associated with diatom ooze (46-37 mcd), thought to be deposited during maximum nutrient availability and primary productivity. Herbs (Brassicaceae), trees/shrubs (Fabaceae and Theaceae), and C3 grasses correlated with inorganic parameters, indicating increased drainage of ultramafic sediments and laterite soils from the lakes' catchment, particularly at times of inferred drying. Downcore variability in trnL-P6 from tropical forest trees (Toona), shady ground cover herbs (Zingiberaceae), and tree orchids (Luisia) most strongly correlated with sediments of a predominantly felsic signature considered to be originating from the catchment of the Loeha River draining into Lake Towuti during wetter climate conditions. However, the co-correlation with dry climate-adapted trees (i.e., Castanopsis or Lithocarpus) plus C4 grasses suggests that increased precipitation seasonality also contributed to the increased drainage of felsic Loeha River sediments. This multiproxy approach shows that despite elevated insitu temperatures, tropical lake sediments potentially comprise long-term archives of ancient environmental DNA for reconstructing ecosystems, which warrants further exploration.

Ancient Mycobacterium leprae genome reveals medieval English red squirrels as animal leprosy host

Leprosy, one of the oldest recorded diseases in human history, remains prevalent in Asia, Africa, and South America, with over 200,000 cases every year.1,2 Although ancient DNA (aDNA) approaches on the major causative agent, Mycobacterium leprae, have elucidated the disease's evolutionary history,3,4,5 the role of animal hosts and interspecies transmission in the past remains unexplored. Research has uncovered relationships between medieval strains isolated from archaeological human remains and modern animal hosts such as the red squirrel in England.6,7 However, the time frame, distribution, and direction of transmissions remains unknown. Here, we studied 25 human and 12 squirrel samples from two archaeological sites in Winchester, a medieval English city well known for its leprosarium and connections to the fur trade. We reconstructed four medieval M.leprae genomes, including one from a red squirrel, at a 2.2-fold average coverage. Our analysis revealed a phylogenetic placement of all strains on branch 3 as well as a close relationship between the squirrel strain and one newly reconstructed medieval human strain. In particular, the medieval squirrel strain is more closely related to some medieval human strains from Winchester than to modern red squirrel strains from England, indicating a yet-undetected circulation of M.leprae in non-human hosts in the Middle Ages. Our study represents the first One Health approach for M.leprae in archaeology, which is centered around a medieval animal host strain, and highlights the future capability of such approaches to understand the disease's zoonotic past and current potential.

Population, culture history, and the dynamics of change in European prehistory★

AbstractDespite many attacks on its shortcomings, culture history has remained in practice the dominant framework for describing and interpreting European prehistory. It has gained even more salience in recent years because the new information coming from ancient DNA about the genetic ancestry of individuals in prehistory seems to show that this correlates closely with the cultural affiliation of the archaeological material with which they are found, raising concerns that old and discredited links between biological and cultural identity are being revived. This article argues that exploring the links between cultural and genetic ancestry does not need to fall into these errors if it takes its theory and methods from the discipline of cultural evolution and rejects characterization of the relationship in terms of ‘ethnic groups’. This involves describing the archaeological record in more fine‐grained, less essentialist ways and at the same time linking the archaeological and genetic patterns to histories of the rise and decline of populations and the interactions between them.