Reconstructing Prehistoric African Population Structure.

We assembled genome-wide data from 16 prehistoric Africans. We show that the anciently divergent lineage that comprises the primary ancestry of the southern African San had a wider distribution in the past, contributing approximately two-thirds of the ancestry of Malawi hunter-gatherers ∼8,100-2,500 years ago and approximately one-third of the ancestry of Tanzanian hunter-gatherers ∼1,400 years ago. We document how the spread of farmers from western Africa involved complete replacement of local hunter-gatherers in some regions, and we track the spread of herders by showing that the population of a ∼3,100-year-old pastoralist from Tanzania contributed ancestry to people from northeastern to southern Africa, including a ∼1,200-year-old southern African pastoralist. The deepest diversifications of African lineages were complex, involving either repeated gene flow among geographically disparate groups or a lineage more deeply diverging than that of the San contributing more to some western African populations than to others. We finally leverage ancient genomes to document episodes of natural selection in southern African populations. PAPERCLIP.

Entwined African and Asian genetic roots of medieval peoples of the Swahili coast.

The urban peoples of the Swahili coast traded across eastern Africa and the Indian Ocean and were among the first practitioners of Islam among sub-Saharan people1,2. The extent to which these early interactions between Africans and non-Africans were accompanied by genetic exchange remains unknown. Here we report ancient DNA data for 80 individuals from 6 medieval and early modern (AD 1250-1800) coastal towns and an inland town after AD 1650. More than half of the DNA of many of the individuals from coastal towns originates from primarily female ancestors from Africa, with a large proportion-and occasionally more than half-of the DNA coming from Asian ancestors. The Asian ancestry includes components associated with Persia and India, with 80-90% of the Asian DNA originating from Persian men. Peoples of African and Asian origins began to mix by about AD 1000, coinciding with the large-scale adoption of Islam. Before about AD 1500, the Southwest Asian ancestry was mainly Persian-related, consistent with the narrative of the Kilwa Chronicle, the oldest history told by people of the Swahili coast3. After this time, the sources of DNA became increasingly Arabian, consistent with evidence of growing interactions with southern Arabia4. Subsequent interactions with Asian and African people further changed the ancestry of present-day people of the Swahili coast in relation to the medieval individuals whose DNA we sequenced.

Ancient DNA and deep population structure in sub-Saharan African foragers.

Multiple lines of genetic and archaeological evidence suggest that there were major demographic changes in the terminal Late Pleistocene epoch and early Holocene epoch of sub-Saharan Africa1-4. Inferences about this period are challenging to make because demographic shifts in the past 5,000 years have obscured the structures of more ancient populations3,5. Here we present genome-wide ancient DNA data for six individuals from eastern and south-central Africa spanning the past approximately 18,000 years (doubling the time depth of sub-Saharan African ancient DNA), increase the data quality for 15 previously published ancient individuals and analyse these alongside data from 13 other published ancient individuals. The ancestry of the individuals in our study area can be modelled as a geographically structured mixture of three highly divergent source populations, probably reflecting Pleistocene interactions around 80-20 thousand years ago, including deeply diverged eastern and southern African lineages, plus a previously unappreciated ubiquitous distribution of ancestry that occurs in highest proportion today in central African rainforest hunter-gatherers. Once established, this structure remained highly stable, with limited long-range gene flow. These results provide a new line of genetic evidence in support of hypotheses that have emerged from archaeological analyses but remain contested, suggesting increasing regionalization at the end of the Pleistocene epoch.

A genetic history of the pre-contact Caribbean.

Humans settled the Caribbean about 6,000 years ago, and ceramic use and intensified agriculture mark a shift from the Archaic to the Ceramic Age at around 2,500 years ago1-3. Here we report genome-wide data from 174 ancient individuals from The Bahamas, Haiti and the Dominican Republic (collectively, Hispaniola), Puerto Rico, Curaçao and Venezuela, which we co-analysed with 89 previously published ancient individuals. Stone-tool-using Caribbean people, who first entered the Caribbean during the Archaic Age, derive from a deeply divergent population that is closest to Central and northern South American individuals; contrary to previous work4, we find no support for ancestry contributed by a population related to North American individuals. Archaic-related lineages were >98% replaced by a genetically homogeneous ceramic-using population related to speakers of languages in the Arawak family from northeast South America; these people moved through the Lesser Antilles and into the Greater Antilles at least 1,700 years ago, introducing ancestry that is still present. Ancient Caribbean people avoided close kin unions despite limited mate pools that reflect small effective population sizes, which we estimate to be a minimum of 500-1,500 and a maximum of 1,530-8,150 individuals on the combined islands of Puerto Rico and Hispaniola in the dozens of generations before the individuals who we analysed lived. Census sizes are unlikely to be more than tenfold larger than effective population sizes, so previous pan-Caribbean estimates of hundreds of thousands of people are too large5,6. Confirming a small and interconnected Ceramic Age population7, we detect 19 pairs of cross-island cousins, close relatives buried around 75 km apart in Hispaniola and low genetic differentiation across islands. Genetic continuity across transitions in pottery styles reveals that cultural changes during the Ceramic Age were not driven by migration of genetically differentiated groups from the mainland, but instead reflected interactions within an interconnected Caribbean world1,8.

Ethics of DNA research on human remains: five globally applicable guidelines.

We are a group of archaeologists, anthropologists, curators and geneticists representing diverse global communities and 31 countries. All of us met in a virtual workshop dedicated to ethics in ancient DNA research held in November 2020. There was widespread agreement that globally applicable ethical guidelines are needed, but that recent recommendations grounded in discussion about research on human remains from North America are not always generalizable worldwide. Here we propose the following globally applicable guidelines, taking into consideration diverse contexts. These hold that: (1) researchers must ensure that all regulations were followed in the places where they work and from which the human remains derived; (2) researchers must prepare a detailed plan prior to beginning any study; (3) researchers must minimize damage to human remains; (4) researchers must ensure that data are made available following publication to allow critical re-examination of scientific findings; and (5) researchers must engage with other stakeholders from the beginning of a study and ensure respect and sensitivity to stakeholder perspectives. We commit to adhering to these guidelines and expect they will promote a high ethical standard in DNA research on human remains going forward.

Genomic insights into the formation of human populations in East Asia.

The deep population history of East Asia remains poorly understood owing to a lack of ancient DNA data and sparse sampling of present-day people1,2. Here we report genome-wide data from 166 East Asian individuals dating to between 6000 BC and AD 1000 and 46 present-day groups. Hunter-gatherers from Japan, the Amur River Basin, and people of Neolithic and Iron Age Taiwan and the Tibetan Plateau are linked by a deeply splitting lineage that probably reflects a coastal migration during the Late Pleistocene epoch. We also follow expansions during the subsequent Holocene epoch from four regions. First, hunter-gatherers from Mongolia and the Amur River Basin have ancestry shared by individuals who speak Mongolic and Tungusic languages, but do not carry ancestry characteristic of farmers from the West Liao River region (around 3000 BC), which contradicts theories that the expansion of these farmers spread the Mongolic and Tungusic proto-languages. Second, farmers from the Yellow River Basin (around 3000 BC) probably spread Sino-Tibetan languages, as their ancestry dispersed both to Tibet-where it forms approximately 84% of the gene pool in some groups-and to the Central Plain, where it has contributed around 59-84% to modern Han Chinese groups. Third, people from Taiwan from around 1300 BC to AD 800 derived approximately 75% of their ancestry from a lineage that is widespread in modern individuals who speak Austronesian, Tai-Kadai and Austroasiatic languages, and that we hypothesize derives from farmers of the Yangtze River Valley. Ancient people from Taiwan also derived about 25% of their ancestry from a northern lineage that is related to, but different from, farmers of the Yellow River Basin, which suggests an additional north-to-south expansion. Fourth, ancestry from Yamnaya Steppe pastoralists arrived in western Mongolia after around 3000 BC but was displaced by previously established lineages even while it persisted in western China, as would be expected if this ancestry was associated with the spread of proto-Tocharian Indo-European languages. Two later gene flows affected western Mongolia: migrants after around 2000 BC with Yamnaya and European farmer ancestry, and episodic influences of later groups with ancestry from Turan.

Ethical considerations when co-analyzing ancient DNA and data from private genetic databases.

Ancient DNA studies have begun to explore the possibility of identifying identical DNA segments shared between historical and living people. This research requires access to large genetic datasets to maximize the likelihood of identifying previously unknown, close genetic connections. Direct-to-consumer genetic testing companies, such as 23andMe, Inc., manage by far the largest and most diverse genetic databases that can be used for this purpose. It is therefore important to think carefully about guidelines for carrying out collaborations between researchers and such companies. Such collaborations require consideration of ethical issues, including policies for sharing ancient DNA datasets, and ensuring reproducibility of research findings when access to privately controlled genetic datasets is limited. At the same time, they introduce unique possibilities for returning results to the research participants whose data are analyzed, including those who are identified as close genetic relatives of historical individuals, thereby enabling ancient DNA research to contribute to the restoration of information about ancestral connections that were lost over time, which can be particularly meaningful for families and groups where such history has not been well documented. We explore these issues by describing our experience designing and carrying out a study searching for genetic connections between 18th- and 19th-century enslaved and free African Americans who labored at Catoctin Furnace, Maryland, and 23andMe research participants. We share our experience in the hope of helping future researchers navigate similar ethical considerations, recognizing that our perspective is part of a larger conversation about best ethical practices.

Density separation of petrous bone powders for optimized ancient DNA yields.

Density separation is a process routinely used to segregate minerals, organic matter, and even microplastics, from soils and sediments. Here we apply density separation to archaeological bone powders before DNA extraction to increase endogenous DNA recovery relative to a standard control extraction of the same powders. Using nontoxic heavy liquid solutions, we separated powders from the petrous bones of 10 individuals of similar archaeological preservation into eight density intervals (2.15 to 2.45 g/cm3, in 0.05 increments). We found that the 2.30 to 2.35 g/cm3 and 2.35 to 2.40 g/cm3 intervals yielded up to 5.28-fold more endogenous unique DNA than the corresponding standard extraction (and up to 8.53-fold before duplicate read removal), while maintaining signals of ancient DNA authenticity and not reducing library complexity. Although small 0.05 g/cm3 intervals may maximally optimize yields, a single separation to remove materials with a density above 2.40 g/cm3 yielded up to 2.57-fold more endogenous DNA on average, which enables the simultaneous separation of samples that vary in preservation or in the type of material analyzed. While requiring no new ancient DNA laboratory equipment and fewer than 30 min of extra laboratory work, the implementation of density separation before DNA extraction can substantially boost endogenous DNA yields without decreasing library complexity. Although subsequent studies are required, we present theoretical and practical foundations that may prove useful when applied to other ancient DNA substrates such as teeth, other bones, and sediments.

A minimally destructive protocol for DNA extraction from ancient teeth.

Ancient DNA sampling methods-although optimized for efficient DNA extraction-are destructive, relying on drilling or cutting and powdering (parts of) bones and teeth. As the field of ancient DNA has grown, so have concerns about the impact of destructive sampling of the skeletal remains from which ancient DNA is obtained. Due to a particularly high concentration of endogenous DNA, the cementum of tooth roots is often targeted for ancient DNA sampling, but destructive sampling methods of the cementum often result in the loss of at least one entire root. Here, we present a minimally destructive method for extracting ancient DNA from dental cementum present on the surface of tooth roots. This method does not require destructive drilling or grinding, and, following extraction, the tooth remains safe to handle and suitable for most morphological studies, as well as other biochemical studies, such as radiocarbon dating. We extracted and sequenced ancient DNA from 30 teeth (and nine corresponding petrous bones) using this minimally destructive extraction method in addition to a typical tooth sampling method. We find that the minimally destructive method can provide ancient DNA that is of comparable quality to extracts produced from teeth that have undergone destructive sampling processes. Further, we find that a rigorous cleaning of the tooth surface combining diluted bleach and UV light irradiation seems sufficient to minimize external contaminants usually removed through the physical removal of a superficial layer when sampling through regular powdering methods.

The genomic history of southeastern Europe.

Farming was first introduced to Europe in the mid-seventh millennium bc, and was associated with migrants from Anatolia who settled in the southeast before spreading throughout Europe. Here, to understand the dynamics of this process, we analysed genome-wide ancient DNA data from 225 individuals who lived in southeastern Europe and surrounding regions between 12000 and 500 bc. We document a west-east cline of ancestry in indigenous hunter-gatherers and, in eastern Europe, the early stages in the formation of Bronze Age steppe ancestry. We show that the first farmers of northern and western Europe dispersed through southeastern Europe with limited hunter-gatherer admixture, but that some early groups in the southeast mixed extensively with hunter-gatherers without the sex-biased admixture that prevailed later in the north and west. We also show that southeastern Europe continued to be a nexus between east and west after the arrival of farmers, with intermittent genetic contact with steppe populations occurring up to 2,000 years earlier than the migrations from the steppe that ultimately replaced much of the population of northern Europe.

Human auditory ossicles as an alternative optimal source of ancient DNA.

DNA recovery from ancient human remains has revolutionized our ability to reconstruct the genetic landscape of the past. Ancient DNA research has benefited from the identification of skeletal elements, such as the cochlear part of the osseous inner ear, that provides optimal contexts for DNA preservation; however, the rich genetic information obtained from the cochlea must be counterbalanced against the loss of morphological information caused by its sampling. Motivated by similarities in developmental processes and histological properties between the cochlea and auditory ossicles, we evaluate the ossicles as an alternative source of ancient DNA. We show that ossicles perform comparably to the cochlea in terms of DNA recovery, finding no substantial reduction in data quantity and minimal differences in data quality across preservation conditions. Ossicles can be sampled from intact skulls or disarticulated petrous bones without damage to surrounding bone, and we argue that they should be used when available to reduce damage to human remains. Our results identify another optimal skeletal element for ancient DNA analysis and add to a growing toolkit of sampling methods that help to better preserve skeletal remains for future research while maximizing the likelihood that ancient DNA analysis will produce useable results.

Genomic insights into the peopling of the Southwest Pacific.

The appearance of people associated with the Lapita culture in the South Pacific around 3,000 years ago marked the beginning of the last major human dispersal to unpopulated lands. However, the relationship of these pioneers to the long-established Papuan people of the New Guinea region is unclear. Here we present genome-wide ancient DNA data from three individuals from Vanuatu (about 3,100-2,700 years before present) and one from Tonga (about 2,700-2,300 years before present), and analyse them with data from 778 present-day East Asians and Oceanians. Today, indigenous people of the South Pacific harbour a mixture of ancestry from Papuans and a population of East Asian origin that no longer exists in unmixed form, but is a match to the ancient individuals. Most analyses have interpreted the minimum of twenty-five per cent Papuan ancestry in the region today as evidence that the first humans to reach Remote Oceania, including Polynesia, were derived from population mixtures near New Guinea, before their further expansion into Remote Oceania. However, our finding that the ancient individuals had little to no Papuan ancestry implies that later human population movements spread Papuan ancestry through the South Pacific after the first peopling of the islands.

Genomic insights into the origin of farming in the ancient Near East.

We report genome-wide ancient DNA from 44 ancient Near Easterners ranging in time between ~12,000 and 1,400 bc, from Natufian hunter-gatherers to Bronze Age farmers. We show that the earliest populations of the Near East derived around half their ancestry from a 'Basal Eurasian' lineage that had little if any Neanderthal admixture and that separated from other non-African lineages before their separation from each other. The first farmers of the southern Levant (Israel and Jordan) and Zagros Mountains (Iran) were strongly genetically differentiated, and each descended from local hunter-gatherers. By the time of the Bronze Age, these two populations and Anatolian-related farmers had mixed with each other and with the hunter-gatherers of Europe to greatly reduce genetic differentiation. The impact of the Near Eastern farmers extended beyond the Near East: farmers related to those of Anatolia spread westward into Europe; farmers related to those of the Levant spread southward into East Africa; farmers related to those of Iran spread northward into the Eurasian steppe; and people related to both the early farmers of Iran and to the pastoralists of the Eurasian steppe spread eastward into South Asia.

Genome-wide patterns of selection in 230 ancient Eurasians.

Ancient DNA makes it possible to observe natural selection directly by analysing samples from populations before, during and after adaptation events. Here we report a genome-wide scan for selection using ancient DNA, capitalizing on the largest ancient DNA data set yet assembled: 230 West Eurasians who lived between 6500 and 300 bc, including 163 with newly reported data. The new samples include, to our knowledge, the first genome-wide ancient DNA from Anatolian Neolithic farmers, whose genetic material we obtained by extracting from petrous bones, and who we show were members of the population that was the source of Europe's first farmers. We also report a transect of the steppe region in Samara between 5600 and 300 bc, which allows us to identify admixture into the steppe from at least two external sources. We detect selection at loci associated with diet, pigmentation and immunity, and two independent episodes of selection on height.

Developmental patterns of bilateral asymmetry in ancestral puebloans.

OBJECTIVES: Producing and maintaining a bilaterally symmetric phenotype throughout the lifespan is energetically demanding. Over the course of an individual's life, various intrinsic and external stressors impact the growth trajectory. These perturbations can compromise the allocation of energetic resources to processes that maintain developmental precision, potentially resulting in bilateral asymmetry (BA). Because different stressors are present during the lifespan, BA is a valuable tool for examining the unique factors impacting symmetrical growth and development. This study examines BA in paired long bones across a developmental skeletal series. METHODS: The humeri, radii, femora, and tibiae of 198 individuals from Ancestral Puebloan New Mexico (919-1670 CE) are analyzed to explore BA across development. Individuals are separated into five age categories, and by sex when possible, to explore patterns of BA. RESULTS: Significant BA is found in the bones of the upper limb when the interaction between bone and age is examined. Results suggest that BA in the humerus and radius becomes more right-biased with age. These directional trends are not observed in the lower limbs. Division into age categories illuminates patterns of asymmetry associated with age-related activities and physiological maturity, indicating that BA is differentially affected by varying environmental stressors across development. CONCLUSIONS: Our findings support the hypothesis that BA in long bones is influenced by environmental stressors that impact an individual's ability to produce symmetric morphological traits over the lifespan. Right-biased BA in the upper limb bones indicates that this variation from a symmetric ideal is strongly influenced by handedness resulting from habitual manual activities. Am. J. Hum. Biol. 28:421-430, 2016. © 2015 Wiley Periodicals, Inc.

Medieval DNA from Soqotra points to Eurasian origins of an isolated population at the crossroads of Africa and Arabia.

Soqotra, an island situated at the mouth of the Gulf of Aden in the northwest Indian Ocean between Africa and Arabia, is home to ~60,000 people subsisting through fishing and semi-nomadic pastoralism who speak a Modern South Arabian language. Most of what is known about Soqotri history derives from writings of foreign travellers who provided little detail about local people, and the geographic origins and genetic affinities of early Soqotri people has not yet been investigated directly. Here we report genome-wide data from 39 individuals who lived between ~650 and 1750 CE at six locations across the island and document strong genetic connections between Soqotra and the similarly isolated Hadramawt region of coastal South Arabia that likely reflects a source for the peopling of Soqotra. Medieval Soqotri can be modelled as deriving ~86% of their ancestry from a population such as that found in the Hadramawt today, with the remaining ~14% best proxied by an Iranian-related source with up to 2% ancestry from the Indian sub-continent, possibly reflecting genetic exchanges that occurred along with archaeologically documented trade from these regions. In contrast to all other genotyped populations of the Arabian Peninsula, genome-level analysis of the medieval Soqotri is consistent with no sub-Saharan African admixture dating to the Holocene. The deep ancestry of people from medieval Soqotra and the Hadramawt is also unique in deriving less from early Holocene Levantine farmers and more from groups such as Late Pleistocene hunter-gatherers from the Levant (Natufians) than other mainland Arabians. This attests to migrations by early farmers having less impact in southernmost Arabia and Soqotra and provides compelling evidence that there has not been complete population replacement between the Pleistocene and Holocene throughout the Arabian Peninsula. Medieval Soqotra harboured a small population that showed qualitatively different marriage practices from modern Soqotri, with first-cousin unions occurring significantly less frequently than today.

The genetic legacy of African Americans from Catoctin Furnace.

Few African Americans have been able to trace family lineages back to ancestors who died before the 1870 United States Census, the first in which all Black people were listed by name. We analyzed 27 individuals from Maryland's Catoctin Furnace African American Cemetery (1774-1850), identifying 41,799 genetic relatives among consenting research participants in 23andMe, Inc.'s genetic database. One of the highest concentrations of close relatives is in Maryland, suggesting that descendants of the Catoctin individuals remain in the area. We find that many of the Catoctin individuals derived African ancestry from the Wolof or Kongo groups and European ancestry from Great Britain and Ireland. This study demonstrates the power of joint analysis of historical DNA and large datasets generated through direct-to-consumer ancestry testing.

Stable carbon and nitrogen isotopes identify nuanced dietary changes from the Bronze and Iron Ages on the Great Hungarian Plain.

The Great Hungarian Plain (GHP) served as a geographic funnel for population mobility throughout prehistory. Genomic and isotopic research demonstrates non-linear genetic turnover and technological shifts between the Copper and Iron Ages of the GHP, which influenced the dietary strategies of numerous cultures that intermixed and overlapped through time. Given the complexities of these prehistoric cultural and demographic processes, this study aims to identify and elucidate diachronic and culture-specific dietary signatures. We report on stable carbon and nitrogen isotope ratios from 74 individuals from nineteen sites in the GHP dating to a ~ 3000-year time span between the Early Bronze and Early Iron Ages. The samples broadly indicate a terrestrial C3 diet with nuanced differences amongst populations and through time, suggesting exogenous influences that manifested in subsistence strategies. Slightly elevated δ15N values for Bronze Age samples imply higher reliance on protein than in the Iron Age. Interestingly, the Füzesabony have carbon values typical of C4 vegetation indicating millet consumption, or that of a grain with comparable δ13C ratios, which corroborates evidence from outside the GHP for its early cultivation during the Middle Bronze Age. Finally, our results also suggest locally diverse subsistence economies for GHP Scythians.

Social stratification without genetic differentiation at the site of Kulubnarti in Christian Period Nubia.

Relatively little is known about Nubia's genetic landscape prior to the influence of the Islamic migrations that began in the late 1st millennium CE. Here, we increase the number of ancient individuals with genome-level data from the Nile Valley from three to 69, reporting data for 66 individuals from two cemeteries at the Christian Period (~650-1000 CE) site of Kulubnarti, where multiple lines of evidence suggest social stratification. The Kulubnarti Nubians had ~43% Nilotic-related ancestry (individual variation between ~36-54%) with the remaining ancestry consistent with being  introduced through Egypt and ultimately deriving from an ancestry pool like that found in the Bronze and Iron Age Levant. The Kulubnarti gene pool - shaped over a millennium - harbors disproportionately female-associated West Eurasian-related ancestry. Genetic similarity among individuals from the two cemeteries supports a hypothesis of social division without genetic distinction. Seven pairs of inter-cemetery relatives suggest fluidity between cemetery groups. Present-day Nubians are not directly descended from the Kulubnarti Nubians, attesting to additional genetic input since the Christian Period.

Integrating buccal and occlusal dental microwear with isotope analyses for a complete paleodietary reconstruction of Holocene populations from Hungary.

Dietary reconstruction is used to make inferences about the subsistence strategies of ancient human populations, but it may also serve as a proxy to characterise their diverse cultural and technological manifestations. Dental microwear and stable isotope analyses have been shown to be successful techniques for paleodietary reconstruction of ancient populations but, despite yielding complementary dietary information, these techniques have rarely been combined within the same study. Here we present for the first time a comprehensive approach to interpreting ancient lifeways through the results of buccal and occlusal microwear, and δ13C and δ15N isotope analyses applied to the same individuals of prehistoric populations of Hungary from the Middle Neolithic to the Late Bronze Age periods. This study aimed to (a) assess if the combination of techniques yields a more precise assessment of past dietary and subsistence practices, and (b) contribute to our understanding of the dietary patterns of the prehistoric Hungarian populations. Overall, no correlations between microwear and δ13C and δ15N isotope variables were observed, except for a relationship between nitrogen and the vertical and horizontal index. However, we found that diachronic differences are influenced by the variation within the period. Particularly, we found differences in microwear and isotope variables between Middle Neolithic sites, indicating that there were different dietary practices among those populations. Additionally, microwear results suggest no changes in the abrasiveness of the diet, neither food processing methods, despite higher C4 plant resource consumption shown by carbon isotopic signal. Thus, we demonstrate that the integration of dental microwear and carbon and nitrogen stable isotope methodologies can provide complementary information for making inferences about paleodietary habits.