Homo sapiens reached the higher latitudes of Europe by 45,000 years ago.

The Middle to Upper Palaeolithic transition in Europe is associated with the regional disappearance of Neanderthals and the spread of Homo sapiens. Late Neanderthals persisted in western Europe several millennia after the occurrence of H. sapiens in eastern Europe1. Local hybridization between the two groups occurred2, but not on all occasions3. Archaeological evidence also indicates the presence of several technocomplexes during this transition, complicating our understanding and the association of behavioural adaptations with specific hominin groups4. One such technocomplex for which the makers are unknown is the Lincombian-Ranisian-Jerzmanowician (LRJ), which has been described in northwestern and central Europe5-8. Here we present the morphological and proteomic taxonomic identification, mitochondrial DNA analysis and direct radiocarbon dating of human remains directly associated with an LRJ assemblage at the site Ilsenhöhle in Ranis (Germany). These human remains are among the earliest directly dated Upper Palaeolithic H. sapiens remains in Eurasia. We show that early H. sapiens associated with the LRJ were present in central and northwestern Europe long before the extinction of late Neanderthals in southwestern Europe. Our results strengthen the notion of a patchwork of distinct human populations and technocomplexes present in Europe during this transitional period.

Genome of a middle Holocene hunter-gatherer from Wallacea.

Much remains unknown about the population history of early modern humans in southeast Asia, where the archaeological record is sparse and the tropical climate is inimical to the preservation of ancient human DNA1. So far, only two low-coverage pre-Neolithic human genomes have been sequenced from this region. Both are from mainland Hòabìnhian hunter-gatherer sites: Pha Faen in Laos, dated to 7939-7751 calibrated years before present (yr cal BP; present taken as AD 1950), and Gua Cha in Malaysia (4.4-4.2 kyr cal BP)1. Here we report, to our knowledge, the first ancient human genome from Wallacea, the oceanic island zone between the Sunda Shelf (comprising mainland southeast Asia and the continental islands of western Indonesia) and Pleistocene Sahul (Australia-New Guinea). We extracted DNA from the petrous bone of a young female hunter-gatherer buried 7.3-7.2 kyr cal BP at the limestone cave of Leang Panninge2 in South Sulawesi, Indonesia. Genetic analyses show that this pre-Neolithic forager, who is associated with the 'Toalean' technocomplex3,4, shares most genetic drift and morphological similarities with present-day Papuan and Indigenous Australian groups, yet represents a previously unknown divergent human lineage that branched off around the time of the split between these populations approximately 37,000 years ago5. We also describe Denisovan and deep Asian-related ancestries in the Leang Panninge genome, and infer their large-scale displacement from the region today.

The genome of the offspring of a Neanderthal mother and a Denisovan father.

Neanderthals and Denisovans are extinct groups of hominins that separated from each other more than 390,000 years ago1,2. Here we present the genome of 'Denisova 11', a bone fragment from Denisova Cave (Russia)3 and show that it comes from an individual who had a Neanderthal mother and a Denisovan father. The father, whose genome bears traces of Neanderthal ancestry, came from a population related to a later Denisovan found in the cave4-6. The mother came from a population more closely related to Neanderthals who lived later in Europe2,7 than to an earlier Neanderthal found in Denisova Cave8, suggesting that migrations of Neanderthals between eastern and western Eurasia occurred sometime after 120,000 years ago. The finding of a first-generation Neanderthal-Denisovan offspring among the small number of archaic specimens sequenced to date suggests that mixing between Late Pleistocene hominin groups was common when they met.

Reconstructing the genetic history of late Neanderthals.

Although it has previously been shown that Neanderthals contributed DNA to modern humans, not much is known about the genetic diversity of Neanderthals or the relationship between late Neanderthal populations at the time at which their last interactions with early modern humans occurred and before they eventually disappeared. Our ability to retrieve DNA from a larger number of Neanderthal individuals has been limited by poor preservation of endogenous DNA and contamination of Neanderthal skeletal remains by large amounts of microbial and present-day human DNA. Here we use hypochlorite treatment of as little as 9 mg of bone or tooth powder to generate between 1- and 2.7-fold genomic coverage of five Neanderthals who lived around 39,000 to 47,000 years ago (that is, late Neanderthals), thereby doubling the number of Neanderthals for which genome sequences are available. Genetic similarity among late Neanderthals is well predicted by their geographical location, and comparison to the genome of an older Neanderthal from the Caucasus indicates that a population turnover is likely to have occurred, either in the Caucasus or throughout Europe, towards the end of Neanderthal history. We find that the bulk of Neanderthal gene flow into early modern humans originated from one or more source populations that diverged from the Neanderthals that were studied here at least 70,000 years ago, but after they split from a previously sequenced Neanderthal from Siberia around 150,000 years ago. Although four of the Neanderthals studied here post-date the putative arrival of early modern humans into Europe, we do not detect any recent gene flow from early modern humans in their ancestry.

Population Genetics and Signatures of Selection in Early Neolithic European Farmers.

Human expansion in the course of the Neolithic transition in western Eurasia has been one of the major topics in ancient DNA research in the last 10 years. Multiple studies have shown that the spread of agriculture and animal husbandry from the Near East across Europe was accompanied by large-scale human expansions. Moreover, changes in subsistence and migration associated with the Neolithic transition have been hypothesized to involve genetic adaptation. Here, we present high quality genome-wide data from the Linear Pottery Culture site Derenburg-Meerenstieg II (DER) (N = 32 individuals) in Central Germany. Population genetic analyses show that the DER individuals carried predominantly Anatolian Neolithic-like ancestry and a very limited degree of local hunter-gatherer admixture, similar to other early European farmers. Increasing the Linear Pottery culture cohort size to ∼100 individuals allowed us to perform various frequency- and haplotype-based analyses to investigate signatures of selection associated with changes following the adoption of the Neolithic lifestyle. In addition, we developed a new method called Admixture-informed Maximum-likelihood Estimation for Selection Scans that allowed us test for selection signatures in an admixture-aware fashion. Focusing on the intersection of results from these selection scans, we identified various loci associated with immune function (JAK1, HLA-DQB1) and metabolism (LMF1, LEPR, SORBS1), as well as skin color (SLC24A5, CD82) and folate synthesis (MTHFR, NBPF3). Our findings shed light on the evolutionary pressures, such as infectious disease and changing diet, that were faced by the early farmers of Western Eurasia.

A complete Neandertal mitochondrial genome sequence determined by high-throughput sequencing.

A complete mitochondrial (mt) genome sequence was reconstructed from a 38,000 year-old Neandertal individual with 8341 mtDNA sequences identified among 4.8 Gb of DNA generated from approximately 0.3 g of bone. Analysis of the assembled sequence unequivocally establishes that the Neandertal mtDNA falls outside the variation of extant human mtDNAs, and allows an estimate of the divergence date between the two mtDNA lineages of 660,000 +/- 140,000 years. Of the 13 proteins encoded in the mtDNA, subunit 2 of cytochrome c oxidase of the mitochondrial electron transport chain has experienced the largest number of amino acid substitutions in human ancestors since the separation from Neandertals. There is evidence that purifying selection in the Neandertal mtDNA was reduced compared with other primate lineages, suggesting that the effective population size of Neandertals was small.

A high-coverage Neandertal genome from Chagyrskaya Cave.

We sequenced the genome of a Neandertal from Chagyrskaya Cave in the Altai Mountains, Russia, to 27-fold genomic coverage. We show that this Neandertal was a female and that she was more related to Neandertals in western Eurasia [Prüfer et al., Science 358, 655-658 (2017); Hajdinjak et al., Nature 555, 652-656 (2018)] than to Neandertals who lived earlier in Denisova Cave [Prüfer et al., Nature 505, 43-49 (2014)], which is located about 100 km away. About 12.9% of the Chagyrskaya genome is spanned by homozygous regions that are between 2.5 and 10 centiMorgans (cM) long. This is consistent with the fact that Siberian Neandertals lived in relatively isolated populations of less than 60 individuals. In contrast, a Neandertal from Europe, a Denisovan from the Altai Mountains, and ancient modern humans seem to have lived in populations of larger sizes. The availability of three Neandertal genomes of high quality allows a view of genetic features that were unique to Neandertals and that are likely to have been at high frequency among them. We find that genes highly expressed in the striatum in the basal ganglia of the brain carry more amino-acid-changing substitutions than genes expressed elsewhere in the brain, suggesting that the striatum may have evolved unique functions in Neandertals.

Nuclear DNA sequences from the Middle Pleistocene Sima de los Huesos hominins.

A unique assemblage of 28 hominin individuals, found in Sima de los Huesos in the Sierra de Atapuerca in Spain, has recently been dated to approximately 430,000 years ago. An interesting question is how these Middle Pleistocene hominins were related to those who lived in the Late Pleistocene epoch, in particular to Neanderthals in western Eurasia and to Denisovans, a sister group of Neanderthals so far known only from southern Siberia. While the Sima de los Huesos hominins share some derived morphological features with Neanderthals, the mitochondrial genome retrieved from one individual from Sima de los Huesos is more closely related to the mitochondrial DNA of Denisovans than to that of Neanderthals. However, since the mitochondrial DNA does not reveal the full picture of relationships among populations, we have investigated DNA preservation in several individuals found at Sima de los Huesos. Here we recover nuclear DNA sequences from two specimens, which show that the Sima de los Huesos hominins were related to Neanderthals rather than to Denisovans, indicating that the population divergence between Neanderthals and Denisovans predates 430,000 years ago. A mitochondrial DNA recovered from one of the specimens shares the previously described relationship to Denisovan mitochondrial DNAs, suggesting, among other possibilities, that the mitochondrial DNA gene pool of Neanderthals turned over later in their history.

Present-Day DNA Contamination in Ancient DNA Datasets.

Present-day contamination can lead to false conclusions in ancient DNA studies. A number of methods are available to estimate contamination, which use a variety of signals and are appropriate for different types of data. Here an overview of currently available methods highlighting their strengths and weaknesses is provided, and a classification based on the signals used to estimate contamination is proposed. This overview aims at enabling researchers to choose the most appropriate methods for their dataset. Based on this classification, potential avenues for the further development of methods are discussed.

A genetic analysis of the Gibraltar Neanderthals.

The Forbes' Quarry and Devil's Tower partial crania from Gibraltar are among the first Neanderthal remains ever found. Here, we show that small amounts of ancient DNA are preserved in the petrous bones of the 2 individuals despite unfavorable climatic conditions. However, the endogenous Neanderthal DNA is present among an overwhelming excess of recent human DNA. Using improved DNA library construction methods that enrich for DNA fragments carrying deaminated cytosine residues, we were able to sequence 70 and 0.4 megabase pairs (Mbp) nuclear DNA of the Forbes' Quarry and Devil's Tower specimens, respectively, as well as large parts of the mitochondrial genome of the Forbes' Quarry individual. We confirm that the Forbes' Quarry individual was a female and the Devil's Tower individual a male. We also show that the Forbes' Quarry individual is genetically more similar to the ∼120,000-y-old Neanderthals from Scladina Cave in Belgium (Scladina I-4A) and Hohlenstein-Stadel Cave in Germany, as well as to a ∼60,000- to 70,000-y-old Neanderthal from Russia (Mezmaiskaya 1), than to a ∼49,000-y-old Neanderthal from El Sidrón (El Sidrón 1253) in northern Spain and other younger Neanderthals from Europe and western Asia. This suggests that the Forbes' Quarry fossil predates the latter Neanderthals. The preservation of archaic human DNA in the warm coastal climate of Gibraltar, close to the shores of Africa, raises hopes for the future recovery of archaic human DNA from regions in which climatic conditions are less than optimal for DNA preservation.

An early modern human from Romania with a recent Neanderthal ancestor.

Neanderthals are thought to have disappeared in Europe approximately 39,000-41,000 years ago but they have contributed 1-3% of the DNA of present-day people in Eurasia. Here we analyse DNA from a 37,000-42,000-year-old modern human from Pestera cu Oase, Romania. Although the specimen contains small amounts of human DNA, we use an enrichment strategy to isolate sites that are informative about its relationship to Neanderthals and present-day humans. We find that on the order of 6-9% of the genome of the Oase individual is derived from Neanderthals, more than any other modern human sequenced to date. Three chromosomal segments of Neanderthal ancestry are over 50 centimorgans in size, indicating that this individual had a Neanderthal ancestor as recently as four to six generations back. However, the Oase individual does not share more alleles with later Europeans than with East Asians, suggesting that the Oase population did not contribute substantially to later humans in Europe.

Detecting ancient positive selection in humans using extended lineage sorting.

Natural selection that affected modern humans early in their evolution has likely shaped some of the traits that set present-day humans apart from their closest extinct and living relatives. The ability to detect ancient natural selection in the human genome could provide insights into the molecular basis for these human-specific traits. Here, we introduce a method for detecting ancient selective sweeps by scanning for extended genomic regions where our closest extinct relatives, Neandertals and Denisovans, fall outside of the present-day human variation. Regions that are unusually long indicate the presence of lineages that reached fixation in the human population faster than expected under neutral evolution. Using simulations, we show that the method is able to detect ancient events of positive selection and that it can differentiate those from background selection. Applying our method to the 1000 Genomes data set, we find evidence for ancient selective sweeps favoring regulatory changes and present a list of genomic regions that are predicted to underlie positively selected human specific traits.

The genomic landscape of Neanderthal ancestry in present-day humans.

Genomic studies have shown that Neanderthals interbred with modern humans, and that non-Africans today are the products of this mixture. The antiquity of Neanderthal gene flow into modern humans means that genomic regions that derive from Neanderthals in any one human today are usually less than a hundred kilobases in size. However, Neanderthal haplotypes are also distinctive enough that several studies have been able to detect Neanderthal ancestry at specific loci. We systematically infer Neanderthal haplotypes in the genomes of 1,004 present-day humans. Regions that harbour a high frequency of Neanderthal alleles are enriched for genes affecting keratin filaments, suggesting that Neanderthal alleles may have helped modern humans to adapt to non-African environments. We identify multiple Neanderthal-derived alleles that confer risk for disease, suggesting that Neanderthal alleles continue to shape human biology. An unexpected finding is that regions with reduced Neanderthal ancestry are enriched in genes, implying selection to remove genetic material derived from Neanderthals. Genes that are more highly expressed in testes than in any other tissue are especially reduced in Neanderthal ancestry, and there is an approximately fivefold reduction of Neanderthal ancestry on the X chromosome, which is known from studies of diverse species to be especially dense in male hybrid sterility genes. These results suggest that part of the explanation for genomic regions of reduced Neanderthal ancestry is Neanderthal alleles that caused decreased fertility in males when moved to a modern human genetic background.

Genome sequence of a 45,000-year-old modern human from western Siberia.

We present the high-quality genome sequence of a ∼45,000-year-old modern human male from Siberia. This individual derives from a population that lived before-or simultaneously with-the separation of the populations in western and eastern Eurasia and carries a similar amount of Neanderthal ancestry as present-day Eurasians. However, the genomic segments of Neanderthal ancestry are substantially longer than those observed in present-day individuals, indicating that Neanderthal gene flow into the ancestors of this individual occurred 7,000-13,000 years before he lived. We estimate an autosomal mutation rate of 0.4 × 10(-9) to 0.6 × 10(-9) per site per year, a Y chromosomal mutation rate of 0.7 × 10(-9) to 0.9 × 10(-9) per site per year based on the additional substitutions that have occurred in present-day non-Africans compared to this genome, and a mitochondrial mutation rate of 1.8 × 10(-8) to 3.2 × 10(-8) per site per year based on the age of the bone.

Ancient human genomes suggest three ancestral populations for present-day Europeans.

We sequenced the genomes of a ∼7,000-year-old farmer from Germany and eight ∼8,000-year-old hunter-gatherers from Luxembourg and Sweden. We analysed these and other ancient genomes with 2,345 contemporary humans to show that most present-day Europeans derive from at least three highly differentiated populations: west European hunter-gatherers, who contributed ancestry to all Europeans but not to Near Easterners; ancient north Eurasians related to Upper Palaeolithic Siberians, who contributed to both Europeans and Near Easterners; and early European farmers, who were mainly of Near Eastern origin but also harboured west European hunter-gatherer related ancestry. We model these populations' deep relationships and show that early European farmers had ∼44% ancestry from a 'basal Eurasian' population that split before the diversification of other non-African lineages.

The complete genome sequence of a Neanderthal from the Altai Mountains.

We present a high-quality genome sequence of a Neanderthal woman from Siberia. We show that her parents were related at the level of half-siblings and that mating among close relatives was common among her recent ancestors. We also sequenced the genome of a Neanderthal from the Caucasus to low coverage. An analysis of the relationships and population history of available archaic genomes and 25 present-day human genomes shows that several gene flow events occurred among Neanderthals, Denisovans and early modern humans, possibly including gene flow into Denisovans from an unknown archaic group. Thus, interbreeding, albeit of low magnitude, occurred among many hominin groups in the Late Pleistocene. In addition, the high-quality Neanderthal genome allows us to establish a definitive list of substitutions that became fixed in modern humans after their separation from the ancestors of Neanderthals and Denisovans.

snpAD: an ancient DNA genotype caller.

Motivation: The study of ancient genomes can elucidate the evolutionary past. However, analyses are complicated by base-modifications in ancient DNA molecules that result in errors in DNA sequences. These errors are particularly common near the ends of sequences and pose a challenge for genotype calling. Results: I describe an iterative method that estimates genotype frequencies and errors along sequences to allow for accurate genotype calling from ancient sequences. The implementation of this method, called snpAD, performs well on high-coverage ancient data, as shown by simulations and by subsampling the data of a high-coverage Neandertal genome. Although estimates for low-coverage genomes are less accurate, I am able to derive approximate estimates of heterozygosity from several low-coverage Neandertals. These estimates show that low heterozygosity, compared to modern humans, was common among Neandertals. Availability and implementation: The C++ code of snpAD is freely available at http://bioinf.eva.mpg.de/snpAD/. Supplementary information: Supplementary data are available at Bioinformatics online.

Great ape genetic diversity and population history.

Most great ape genetic variation remains uncharacterized; however, its study is critical for understanding population history, recombination, selection and susceptibility to disease. Here we sequence to high coverage a total of 79 wild- and captive-born individuals representing all six great ape species and seven subspecies and report 88.8 million single nucleotide polymorphisms. Our analysis provides support for genetically distinct populations within each species, signals of gene flow, and the split of common chimpanzees into two distinct groups: Nigeria-Cameroon/western and central/eastern populations. We find extensive inbreeding in almost all wild populations, with eastern gorillas being the most extreme. Inferred effective population sizes have varied radically over time in different lineages and this appears to have a profound effect on the genetic diversity at, or close to, genes in almost all species. We discover and assign 1,982 loss-of-function variants throughout the human and great ape lineages, determining that the rate of gene loss has not been different in the human branch compared to other internal branches in the great ape phylogeny. This comprehensive catalogue of great ape genome diversity provides a framework for understanding evolution and a resource for more effective management of wild and captive great ape populations.

Quantifying and reducing spurious alignments for the analysis of ultra-short ancient DNA sequences.

BACKGROUND: The study of ancient DNA is hampered by degradation, resulting in short DNA fragments. Advances in laboratory methods have made it possible to retrieve short DNA fragments, thereby improving access to DNA preserved in highly degraded, ancient material. However, such material contains large amounts of microbial contamination in addition to DNA fragments from the ancient organism. The resulting mixture of sequences constitutes a challenge for computational analysis, since microbial sequences are hard to distinguish from the ancient sequences of interest, especially when they are short. RESULTS: Here, we develop a method to quantify spurious alignments based on the presence or absence of rare variants. We find that spurious alignments are enriched for mismatches and insertion/deletion differences and lack substitution patterns typical of ancient DNA. The impact of spurious alignments can be reduced by filtering on these features and by imposing a sample-specific minimum length cutoff. We apply this approach to sequences from four ~ 430,000-year-old Sima de los Huesos hominin remains, which contain particularly short DNA fragments, and increase the amount of usable sequence data by 17-150%. This allows us to place a third specimen from the site on the Neandertal lineage. CONCLUSIONS: Our method maximizes the sequence data amenable to genetic analysis from highly degraded ancient material and avoids pitfalls that are associated with the analysis of ultra-short DNA sequences.

The bonobo genome compared with the chimpanzee and human genomes.

Two African apes are the closest living relatives of humans: the chimpanzee (Pan troglodytes) and the bonobo (Pan paniscus). Although they are similar in many respects, bonobos and chimpanzees differ strikingly in key social and sexual behaviours, and for some of these traits they show more similarity with humans than with each other. Here we report the sequencing and assembly of the bonobo genome to study its evolutionary relationship with the chimpanzee and human genomes. We find that more than three per cent of the human genome is more closely related to either the bonobo or the chimpanzee genome than these are to each other. These regions allow various aspects of the ancestry of the two ape species to be reconstructed. In addition, many of the regions that overlap genes may eventually help us understand the genetic basis of phenotypes that humans share with one of the two apes to the exclusion of the other.