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.

Lions and brown bears colonized North America in multiple synchronous waves of dispersal across the Bering Land Bridge.

The Bering Land Bridge connecting North America and Eurasia was periodically exposed and inundated by oscillating sea levels during the Pleistocene glacial cycles. This land connection allowed the intermittent dispersal of animals, including humans, between Western Beringia (far northeast Asia) and Eastern Beringia (northwest North America), changing the faunal community composition of both continents. The Pleistocene glacial cycles also had profound impacts on temperature, precipitation and vegetation, impacting faunal community structure and demography. While these palaeoenvironmental impacts have been studied in many large herbivores from Beringia (e.g., bison, mammoths, horses), the Pleistocene population dynamics of the diverse guild of carnivorans present in the region are less well understood, due to their lower abundances. In this study, we analyse mitochondrial genome data from ancient brown bears (Ursus arctos; n = 103) and lions (Panthera spp.; n = 39), two megafaunal carnivorans that dispersed into North America during the Pleistocene. Our results reveal striking synchronicity in the population dynamics of Beringian lions and brown bears, with multiple waves of dispersal across the Bering Land Bridge coinciding with glacial periods of low sea levels, as well as synchronous local extinctions in Eastern Beringia during Marine Isotope Stage 3. The evolutionary histories of these two taxa underline the crucial biogeographical role of the Bering Land Bridge in the distribution, turnover and maintenance of megafaunal populations in North America.

Early ontogeny of humeral trabecular bone in Neandertals and recent modern humans.

Trabecular bone ontogeny is well known in modern humans and unknown in Neandertals. Yet the bone developmental pattern is useful for interpreting fossils from evolutionary and functional perspectives. Interestingly, microstructure in early ontogeny is supposedly not influenced by high and specific mechanical loading related to the lifestyle of a human group and consequently does not directly depend on the activities of hunter-gatherers. Here, we specifically explored the early growth trajectories of the trabecular bone structure of the humerus and emphasized in particular how bone fraction (bone volume/total volume [BV/TV]) was built up in Neandertals, given the specific modern human bone loss after birth and the use of BV/TV in functional studies. Six Neandertals and 26 recent modern humans ranging from perinates to adolescents were included in this study. Six trabecular parameters were measured within a cubic region of interest extracted from the proximal metaphysis of the humerus. We found that the microstructural changes in Neandertals during early ontogeny (<1 year) fit with modern human growth trajectories for each parameter. The specific bone loss occurring immediately after birth in modern humans also occurred in Neandertals (but not in chimpanzees). However, the early childhood fossil Ferrassie 6 presented unexpectedly high BV/TV, whereas the high BV/TV in the Crouzade I adolescent was predictable. These results suggest that Neandertals and modern humans shared predetermined early growth trajectories and developmental mechanisms. We assume that the close relationship between skeletal characteristics in early ontogeny and adults in modern humans also existed in Neandertals. However, it was difficult to ensure that the high BV/TV in Neandertal early childhood, represented by only one individual, was at the origin of the high BV/TV observed in adults. Consequently, our study does not challenge the mechanical hypothesis that explains the trabecular gracilization of the humerus during the Holocene.

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.

Neonatal postcrania from Mezmaiskaya, Russia, and Le Moustier, France, and the development of Neandertal body form.

Neandertal and modern human adults differ in skeletal features of the cranium and postcranium, and it is clear that many of the cranial differences-although not all of them-are already present at the time of birth. We know less, however, about the developmental origins of the postcranial differences. Here, we address this deficiency with morphometric analyses of the postcrania of the two most complete Neandertal neonates-Mezmaiskaya 1 (from Russia) and Le Moustier 2 (from France)-and a recent human sample. We find that neonatal Neandertals already appear to possess the wide body, long pubis, and robust long bones of adult Neandertals. Taken together, current evidence indicates that skeletal differences between Neandertals and modern humans are largely established by the time of birth.

Revised age of late Neanderthal occupation and the end of the Middle Paleolithic in the northern Caucasus.

Advances in direct radiocarbon dating of Neanderthal and anatomically modern human (AMH) fossils and the development of archaeostratigraphic chronologies now allow refined regional models for Neanderthal-AMH coexistence. In addition, they allow us to explore the issue of late Neanderthal survival in regions of Western Eurasia located within early routes of AMH expansion such as the Caucasus. Here we report the direct radiocarbon ((14)C) dating of a late Neanderthal specimen from a Late Middle Paleolithic (LMP) layer in Mezmaiskaya Cave, northern Caucasus. Additionally, we provide a more accurate chronology for the timing of Neanderthal extinction in the region through a robust series of 16 ultrafiltered bone collagen radiocarbon dates from LMP layers and using Bayesian modeling to produce a boundary probability distribution function corresponding to the end of the LMP at Mezmaiskaya. The direct date of the fossil (39,700 ± 1,100 (14)C BP) is in good agreement with the probability distribution function, indicating at a high level of probability that Neanderthals did not survive at Mezmaiskaya Cave after 39 ka cal BP ("calendrical" age in kiloannum before present, based on IntCal09 calibration curve). This challenges previous claims for late Neanderthal survival in the northern Caucasus. We see striking and largely synchronous chronometric similarities between the Bayesian age modeling for the end of the LMP at Mezmaiskaya and chronometric data from Ortvale Klde for the end of the LMP in the southern Caucasus. Our results confirm the lack of reliably dated Neanderthal fossils younger than ∼ 40 ka cal BP in any other region of Western Eurasia, including the Caucasus.

Archaeological evidence for two culture diverse Neanderthal populations in the North Caucasus and contacts between them.

Neanderthals were widespread during the Middle Palaeolithic (MP) across Europe and Asia, including the Caucasus Mountains. Occupying the border between eastern Europe and West Asia, the Caucasus is important region regarding the Neanderthal occupation of Eurasia. On current radiometric estimates, the MP is represented in the Caucasus between about 260-210 ka and about 40 ka. Archaeological record indicates that several culture diverse MP hominin populations inhabited the Caucasus, but the region complex population history during this period remains poorly understood. In this paper, we identify for the first time the archaeological evidence indicating contacts between two culture diverse MP Neanderthal populations in the North Caucasus and discuss the nature of these contacts. Basing on the lithic assemblages that we excavated at Mezmaiskaya cave in the north-western Caucasus (Kuban River basin) and Saradj-Chuko grotto in the north-central Caucasus (Terek River basin), dating from MIS 5 to MIS 3, and comparative data from other MP sites in the Caucasus, we identify two large cultural regions that existed during the late MP in the North Caucasus. The distinctive toolkits and stone knapping technologies indicate that the MP assemblages from Mezmaiskaya cave and other sites in the west of North Caucasus represent a Caucasian variant of the Eastern Micoquian industry that was wide spread in central and eastern Europe, while the assemblages from Saradj-Chuko Grotto and other sites in the east of North Caucasus closely resemble the Zagros Mousterian industry that was wide spread in the Armenian Highlands, Lesser Caucasus and Zagros Mountains. The archaeological evidence implies that two culture diverse populations of Neanderthals settled the North Caucasus during the Late Pleistocene from two various source regions: from the Armenian Highlands and Lesser Caucasus along the Caspian Sea coast, and from Russian plain along the Sea of Azov coast.

Anatomically modern human in the Châtelperronian hominin collection from the Grotte du Renne (Arcy-sur-Cure, Northeast France).

Around 42,000 years ago, anatomically modern humans appeared in Western Europe to the detriment of indigenous Neanderthal groups. It is during this period that new techno-cultural complexes appear, such as the Châtelperronian that extends from northern Spain to the Paris Basin. The Grotte du Renne (Arcy-sur-Cure) is a key site for discussing the biological identity of its makers. This deposit has yielded several Neanderthal human remains in its Châtelperronian levels. However, the last inventory of the paleoanthropological collection attributed to this techno-complex allowed the identification of an ilium belonging to a neonate (AR-63) whose morphology required a thorough analysis to assess its taxonomic attribution. Using geometric morphometrics, we quantified its morphology and compared it to that of 2 Neanderthals and 32 recent individuals deceased during the perinatal period to explore their morphological variation. Our results indicate a morphological distinction between the ilia of Neanderthals and anatomically modern neonates. Although AR-63 is slightly outside recent variability, it clearly differs from the Neanderthals. We propose that this is due to its belonging to an early modern human lineage whose morphology differs slightly from present-day humans. We also explore different hypotheses about the presence of this anatomically modern neonate ilium among Neanderthal remains.

A uniquely modern human pattern of endocranial development. Insights from a new cranial reconstruction of the Neandertal newborn from Mezmaiskaya.

The globular braincase of modern humans is distinct from all fossil human species, including our closest extinct relatives, the Neandertals. Such adult shape differences must ultimately be rooted in different developmental patterns, but it is unclear at which point during ontogeny these group characteristics emerge. Here we compared internal shape changes of the braincase from birth to adulthood in Neandertals (N = 10), modern humans (N = 62), and chimpanzees (N = 62). Incomplete fossil specimens, including the two Neandertal newborns from Le Moustier 2 and Mezmaiskaya, were reconstructed using reference-based estimation methods. We used 3D geometric morphometrics to statistically compare shapes of virtual endocasts extracted from computed-tomographic scans. Throughout the analysis, we kept track of possible uncertainties due to the missing data values and small fossil sample sizes. We find that some aspects of endocranial development are shared by the three species. However, in the first year of life, modern humans depart from this presumably ancestral pattern of development. Newborn Neandertals and newborn modern humans have elongated braincases, and similar endocranial volumes. During a 'globularization-phase' modern human endocasts change to the globular shape that is characteristic for Homo sapiens. This phase of early development is unique to modern humans, and absent from chimpanzees and Neandertals. Our results support the notion that Neandertals and modern humans reach comparable adult brain sizes via different developmental pathways. The differences between these two human groups are most prominent directly after birth, a critical phase for cognitive development.

Genomic analysis of a novel Neanderthal from Mezmaiskaya Cave provides insights into the genetic relationships of Middle Palaeolithic populations.

The Mezmaiskaya cave is located on the North Caucasus near the border that divides Europe and Asia. Previously, fossil remains for two Neanderthals were reported from Mezmaiskaya Cave. A tooth from the third archaic hominin specimen (Mezmaiskaya 3) was retrieved from layer 3 in Mezmaiskaya Cave. We performed genome sequencing of Mezmaiskaya 3. Analysis of partial nuclear genome sequence revealed that it belongs to a Homo sapiens neanderthalensis female. Based on a high-coverage mitochondrial genome sequence, we demonstrated that the relationships of Mezmaiskaya 3 to Mezmaiskaya 1 and Stajnia S5000 individuals were closer than those to other Neanderthals. Our data demonstrate the close genetic connections between the early Middle Palaeolithic Neanderthals that were replaced by genetically distant later group in the same geographic areas. Based on mitochondrial DNA (mtDNA) data, we suggest that Mezmaiskaya 3 was the latest Neanderthal individual from the early Neanderthal's branches. We proposed a hierarchical nomenclature for the mtDNA haplogroups of Neanderthals. In addition, we retrieved ancestral mtDNA mutations in presumably functional sites fixed in the Neanderthal clades, and also provided the first data showing mtDNA heteroplasmy in Neanderthal specimen.

Neanderthal brain size at birth provides insights into the evolution of human life history.

From birth to adulthood, the human brain expands by a factor of 3.3, compared with 2.5 in chimpanzees [DeSilva J and Lesnik J (2006) Chimpanzee neonatal brain size: Implications for brain growth in Homo erectus. J Hum Evol 51: 207-212]. How the required extra amount of human brain growth is achieved and what its implications are for human life history and cognitive development are still a matter of debate. Likewise, because comparative fossil evidence is scarce, when and how the modern human pattern of brain growth arose during evolution is largely unknown. Virtual reconstructions of a Neanderthal neonate from Mezmaiskaya Cave (Russia) and of two Neanderthal infant skeletons from Dederiyeh Cave (Syria) now provide new comparative insights: Neanderthal brain size at birth was similar to that in recent Homo sapiens and most likely subject to similar obstetric constraints. Neanderthal brain growth rates during early infancy were higher, however. This pattern of growth resulted in larger adult brain sizes but not in earlier completion of brain growth. Because large brains growing at high rates require large, late-maturing, mothers [Leigh SR and Blomquist GE (2007) in Campbell CJ et al. Primates in perspective; pp 396-407], it is likely that Neanderthal life history was similarly slow, or even slower-paced, than in recent H. sapiens.

Early development of the Neanderthal ribcage reveals a different body shape at birth compared to modern humans.

Ontogenetic studies provide clues for understanding important paleobiological aspects of extinct species. When compared to that of modern humans, the adult Neanderthal thorax was shorter, deeper, and wider. This is related to the wide Neanderthal body and is consistent with their hypothetical large requirements for energy and oxygen. Whether these differences were already established at birth or appeared later during development is unknown. To delve into this question, we use virtual reconstruction tools and geometric morphometrics to recover the 3D morphology of the ribcages of four Neanderthal individuals from birth to around 3 years old: Mezmaiskaya 1, Le Moustier 2, Dederiyeh 1, and Roc de Marsal. Our results indicate that the comparatively deep and short ribcage of the Neanderthals was already present at birth, as were other skeletal species-specific traits. This morphology possibly represents the plesiomorphic condition shared with Homo erectus, and it is likely linked to large energetic requirements.

Sedimentary Dosimetry for the Saradj-Chuko Grotto: A Cave in a Lava Tube in the North-Central Caucasus, Russia.

Karst caves host most European Paleolithic sites. Near the Eurasian-Arabian Plate convergence in the Caucasus' Lower Chegem Formation, Saradj-Chuko Grotto (SCG), a lava tube, contains 16 geoarchaeologically distinct horizons yielding modern to laminar obsidian-rich Middle Paleolithic (MP) assemblages. Since electron spin resonance (ESR) can date MP teeth with 2-5% uncertainty, 40 sediment samples were analyzed by neutron activation analysis to measure volumetrically averaged sedimentary dose rates. SCG's rhyolitic ignimbrite walls produce very acidic clay-rich conglomeratic silts that retain 16-24 wt% water today. In Layers 6A-6B, the most prolific MP layers, strongly decalcified bones hinder species identification, but large ungulates inhabited deciduous interglacial forests. Unlike in karst caves, most SCG's layers had sedimentary U concentrations > 4 ppm and Th, > 12 ppm, but Layer 6B2 exceeded 20.8 ppm U, and Layer 7, > 5 ppm Th. Such high concentrations emit dose rates averaging ~ 1.9-3.7 mGy/y, but locally up to 4.1-5.0 mGy/y. Within Layer 6, dose rate variations reflect bone occurrence, necessitating that several samples must be geochemically analyzed around each tooth to ensure age accuracy. Coupled with dentinal dose rates up to 3.7-4.5 mGy/y, SCG's maximum datable ages likely averages ~ 500-800 ka.

The evolutionary history of Neanderthal and Denisovan Y chromosomes.

Ancient DNA has provided new insights into many aspects of human history. However, we lack comprehensive studies of the Y chromosomes of Denisovans and Neanderthals because the majority of specimens that have been sequenced to sufficient coverage are female. Sequencing Y chromosomes from two Denisovans and three Neanderthals shows that the Y chromosomes of Denisovans split around 700 thousand years ago from a lineage shared by Neanderthals and modern human Y chromosomes, which diverged from each other around 370 thousand years ago. The phylogenetic relationships of archaic and modern human Y chromosomes differ from the population relationships inferred from the autosomal genomes and mirror mitochondrial DNA phylogenies, indicating replacement of both the mitochondrial and Y chromosomal gene pools in late Neanderthals. This replacement is plausible if the low effective population size of Neanderthals resulted in an increased genetic load in Neanderthals relative to modern humans.

A high-coverage Neandertal genome from Vindija Cave in Croatia.

To date, the only Neandertal genome that has been sequenced to high quality is from an individual found in Southern Siberia. We sequenced the genome of a female Neandertal from ~50,000 years ago from Vindija Cave, Croatia, to ~30-fold genomic coverage. She carried 1.6 differences per 10,000 base pairs between the two copies of her genome, fewer than present-day humans, suggesting that Neandertal populations were of small size. Our analyses indicate that she was more closely related to the Neandertals that mixed with the ancestors of present-day humans living outside of sub-Saharan Africa than the previously sequenced Neandertal from Siberia, allowing 10 to 20% more Neandertal DNA to be identified in present-day humans, including variants involved in low-density lipoprotein cholesterol concentrations, schizophrenia, and other diseases.

Early cave art and ancient DNA record the origin of European bison.

The two living species of bison (European and American) are among the few terrestrial megafauna to have survived the late Pleistocene extinctions. Despite the extensive bovid fossil record in Eurasia, the evolutionary history of the European bison (or wisent, Bison bonasus) before the Holocene (<11.7 thousand years ago (kya)) remains a mystery. We use complete ancient mitochondrial genomes and genome-wide nuclear DNA surveys to reveal that the wisent is the product of hybridization between the extinct steppe bison (Bison priscus) and ancestors of modern cattle (aurochs, Bos primigenius) before 120 kya, and contains up to 10% aurochs genomic ancestry. Although undetected within the fossil record, ancestors of the wisent have alternated ecological dominance with steppe bison in association with major environmental shifts since at least 55 kya. Early cave artists recorded distinct morphological forms consistent with these replacement events, around the Last Glacial Maximum (LGM, ∼21-18 kya).

ESR at Treugol'naya Cave, Northern Caucasus Mt., Russia: dating Russia's oldest archaeological site and paleoclimatic change in Oxygen Isotope Stage 11.

At 1510 m asl, Treugol'naya Cave, Russia, is the highest cave showing evidence for human occupation in eastern Europe. Layers 4-7 in the 4.5-m-thick sequence yielded many artifacts representing Lower Paleolithic pebble and flake tool industries. Abundant faunal remains include extinct Middle Pleistocene species. Palynological, paleomagnetic, and microsedimentological analyses indicate that several climatic changes of different magnitudes occurred in the sequence. To determine absolute ages for Treugol'naya, 32 independent subsamples from nine ungulate teeth collected from the Lower Paleolithic layers were dated by standard and isochron electron spin resonance (ESR) analyses. Isochron analyses indicate that the teeth experienced no significant U leaching or secondary uptake, and that linear uptake (LU) provides accurate ages. Layers 4b through 5b dated to 365+/-12-406+/-15 ka. Therefore, hominids visited the site periodically throughout Oxygen Isotope Stage (OIS) 11, indicating that they utilized resources at elevations >1000 m at least seasonally by 400 ka. ESR, paleomagnetic, palynological and paleontological analyses all indicate that the Lower Paleolithic Layers 4-5 correlate with OIS 11. The thickness of Layers 4-5 (more than 1.5 m) makes this one of the thickest OIS 11 terrestrial deposits known.

A draft sequence of the Neandertal genome.

Neandertals, the closest evolutionary relatives of present-day humans, lived in large parts of Europe and western Asia before disappearing 30,000 years ago. We present a draft sequence of the Neandertal genome composed of more than 4 billion nucleotides from three individuals. Comparisons of the Neandertal genome to the genomes of five present-day humans from different parts of the world identify a number of genomic regions that may have been affected by positive selection in ancestral modern humans, including genes involved in metabolism and in cognitive and skeletal development. We show that Neandertals shared more genetic variants with present-day humans in Eurasia than with present-day humans in sub-Saharan Africa, suggesting that gene flow from Neandertals into the ancestors of non-Africans occurred before the divergence of Eurasian groups from each other.

Targeted retrieval and analysis of five Neandertal mtDNA genomes.

Analysis of Neandertal DNA holds great potential for investigating the population history of this group of hominins, but progress has been limited due to the rarity of samples and damaged state of the DNA. We present a method of targeted ancient DNA sequence retrieval that greatly reduces sample destruction and sequencing demands and use this method to reconstruct the complete mitochondrial DNA (mtDNA) genomes of five Neandertals from across their geographic range. We find that mtDNA genetic diversity in Neandertals that lived 38,000 to 70,000 years ago was approximately one-third of that in contemporary modern humans. Together with analyses of mtDNA protein evolution, these data suggest that the long-term effective population size of Neandertals was smaller than that of modern humans and extant great apes.