New techniques for the recovery of small amounts of mature enamel proteins

Currently there are no non-destructive techniques to obtain protein from the dental enamel, the most mineralized tissue in mammals and most resistant to diagenesis, which provides a window to the developing period by means of incremental markings containing proteins. To recover protein, dissolution of powdered enamel is required. Here we tested whether samples obtained by micro-etching of the enamel surface were adequate for protein analysis by MALDI-TOF/TOF mass spectrometry and identification in protein databases. The micro-etch techniques were effective in generating adequate samples for mass spectrometry (from 3 to 13.4 μm superficial enamel), being also highly conservative, since they rendered masses of enamel ranging from 0.1 to 0.4 mg. Using these techniques the separation of proteins by SDS-PAGE was not necessary, and the whole procedure was easier. Results showed successful identification of specific enamel proteins after whole crown superficial etching with 11% EDTA in the case of immature porcine samples, and with 10% HCl in the case of mature human enamel. X- and Y-isoforms of amelogenin, ameloblastin, and enamelin peptides were identified. The new techniques described here allowed the successful recovery of enamel proteins, opening new avenues for the use of enamel protein information in fossil/archeological material, where sometimes little protein is left.     

The Cradle of Thought: Growth,Learning, Play and Attachment in Neanderthal Children

Childhood is a core stage in development, essential in the acquisition of social, practical and cultural skills. However, this area receives limited attention in archaeological debate, especially in early prehistory. We here consider Neanderthal childhood, exploring the experience of Neanderthal children using biological, cultural and social evidence. We conclude that Neanderthal childhood experience was subtly different from that of their modern human counterparts, orientated around a greater focus on social relationships within their group. Neanderthal children, as reflected in the burial record, may have played a particularly significant role in their society, especially in the domain of symbolic expression. A consideration of childhood informs broader debates surrounding the subtle differences between Neanderthals and modern humans.     

Fish ’n chips: ZooMS peptide mass fingerprinting in a 96 well plate format to identify fish bone fragments

Fish are a large, highly diverse, and anthropologically important group of vertebrates. However, fish bones are underrepresented in the archaeological literature because they are less stable than those of other taxa and identification of bone to species is often difficult or impossible. We explore a new identification system, ZooMS (Zooarchaeology by Mass Spectrometry), which is based upon protein barcoding. As proteins can be cleaved enzymatically and analyzed by mass spectrometry in a repeatable way, protein barcoding is used widely in microbiological contexts for quick and inexpensive protein identification; mass spectra reflect the differences in protein sequence and can therefore be reproducibly linked to a particular protein or protein fragment. ZooMS uses peptide fingerprinting of bone collagen as a method for rapid identification of archaeological bone. This has involved the identification of masses related to peptides of known sequence. For mammals, sufficient sequence information is available for this approach but for groups, such the teleost fish, species are highly diverse and there are few available collagen sequences. Here we report a preliminary investigation into the identification of fish species by peptide mass fingerprinting that does not require sequence information. Collagen mass spectra are used to identify eight species of bony fishes (Osteichthyes) from four orders: Clupeiformes, Salmoniformes, Gadiformes, and Perciformes. The method is applied to both modern and archaeological fish remains and offers the capacity to identify traditionally unidentifiable fish fragments, thus increasing the Number of Identified Specimens (NISP) and providing invaluable information in specialized contexts.     

In situ oxygen isotope micro-analysis of faunal material and human teeth using a SHRIMP II: a new tool for palaeo-ecology and archaeology

A Sensitive High Resolution Ion MicroProbe (SHRIMP II) has been used to make high spatial resolution in situ micro-analyses of oxygen isotopes in fish otoliths, and teeth from fossil herbivores and a Neanderthal. Large intra-tooth variations in the oxygen isotopic composition (up to 9‰) were observed in the enamel of herbivores from the Neanderthal fossil site of Payre, consistent with preservation of seasonal cyclicity. The range of isotopic compositions observed in Neanderthal tooth enamel was much smaller (∼3‰), possibly the result of a longer enamel maturation time averaging out variability. An archaeological otolith from a Preceramic site in Northern Peru exhibited marked changes in δ¹⁸O over life, due either to the fish occasionally migrating from the sea to a lower salinity habitat, or to short-lived rises in sea water temperature. A fish otolith from Australia's Willandra Lakes World Heritage Area showed clear seasonal variations, but also a general trend towards isotopically heavier and more saline water, as indicated by higher δ¹⁸O and Sr/Ca values resulting from increased evaporation. The results of these case studies are compared to results of oxygen isotope analysis using more conventional methods and demonstrate the ability of the SHRIMP II to provide precise high spatial resolution in-situ oxygen isotope analyses of a variety of biogenic materials. This approach has major advantages over conventional methods. It can provide rapid, micro-scale isotopic analyses of sub-permil precision without the need for chemical preparation of the sample.     

Laser ablation depth profiling of U-series and Sr isotopes in human fossils

We have explored laser depth profiling to obtain data sets for U-series as well as Sr analyses. Laser probing with an 81 μm spot size allows for the exploration of low uranium domains of up to 400 μm below the outer surface in tooth enamel. These low U domains will contain Sr isotope compositions of the individual, that are least affected by diagenetic Sr overprints. The small holes drilled for U surveying are not visible to the naked eye. Using larger spot sizes of around 233 μm, laser drilling can be used to obtain reliable U-series isotope data to a depth of approximately 1000 μm in enamel and around 1300 μm in bone. Furthermore, meaningful ⁸⁷Sr/⁸⁶Sr isotope data can also be obtained with this spot size. Using our sampling strategy, the overall damage to a human tooth is minute, as demonstrated on a Neanderthal tooth from Moula-Guercy. We expect that laser ablation depth profiling will become routine for gaining insights into the age of human fossils and the migrations of ancient humans.     

DATING OF THE HOMINID (HOMO NEANDERTHALENSIS) REMAINS ACCUMULATION FROM EL SIDRÓN CAVE (PILOÑA,ASTURIAS, NORTH SPAIN): AN EXAMPLE OF A MULTI‐METHODOLOGICAL APPROACH TO THE DATING OF UPPER PLEISTOCENE SITES

The age of Neanderthal remains and associated sediments from El Sidrón cave has been obtained through different dating methods (¹⁴CAMS, U/TH, OSL, ESR and AAR) and samples (charcoal debris, bone, tooth dentine, stalagmitic flowstone, carbonate‐rich sediments, sedimentary quartz grains, tooth enamel and land snail shells). Detrital Th contamination rendered Th/U dating analyses of flowstone unreliable. Recent ¹⁴C contamination produced spurious age‐values from charcoal samples as well as from inadequately pretreated tooth samples. Most consistent ¹⁴C dates are grouped into two series: one between 35 and 40 ka and the other between 48 and 49 ka. Most ESR and AAR samples yielded concordant ages, ranging between 39 and 45 ka; OSL dating results permitted adequate bracketing of the sedimentary layer that contained the human remains. Our results emphasize the value of multi‐dating approaches for the establishment of reliable chronologies of human remains.     

Population relationships of later Pleistocene hominids: A multivariate study of available crania

The Neanderthal populations of the Upper Pleistocene have been regarded by a number of anthropologists as direct ancestors to modern man. Results of multivariate analyses conducted in this study suggest that early Neanderthal crania were morphologically more like modern Homo sapiens than were the later Neanderthals. The implications of these results are discussed in the light of archaeological evidence and comparative studies of Pleistocene crania from outside Europe.     

A novel and non-destructive approach for ZooMS analysis: ammonium bicarbonate buffer extraction

Bone collagen is found throughout most of the archaeological record. Under experimental conditions, collagen is apparently preserved as an intact molecule, with amino acid compositions and isotopic profiles only changing when almost all of the protein is lost. The ubiquity of collagen in archaeological bone has lead to the development of the use of collagen peptide mass fingerprints for the identification of bone fragments—Zooarchaeology by Mass Spectrometry (ZooMS). We report a novel, but a simple method for the partial extraction of collagen for ZooMS that uses ammonium bicarbonate buffer but avoids demineralisation. We compared conventional acid demineralisation with ammonium bicarbonate buffer extraction to test ZooMS in a range of modern and archaeological bone samples. The sensitivity of the current generation of mass spectrometers is high enough for the non-destructive buffer method to extract sufficient collagen for ZooMS. We envisage that a particular advantage of this method is that it leaves worked bone artefacts effectively undamaged post-treatment, suitable for subsequent analysis or museum storage or display. Furthermore, it may have potential as a screening tool to aid curators in the selection of material for more advanced molecular analysis—such as DNA sequencing.     

A radiocarbon chronology for the complete Middle to Upper Palaeolithic transitional sequence of Les Cottés (France)

The Middle to Upper Palaeolithic transition is the key period for our understanding of Neanderthal and modern human interactions in Europe. The site of Les Cottés in south-west France is one of the rare sites with a complete and well defined sequence covering this transition period. We undertook an extensive radiocarbon dating program on mammal bone which allows us to propose a chronological framework of five distinct phases dating from the Mousterian to the Early Aurignacian at this site. We found that the Mousterian and Châtelperronian industries are separated from the overlying Protoaurignacian by a gap of approximately 1000 calendar years. Based on a comparison with Upper Paleolithic sites in Europe we see an overlap in the ages of Châtelperronian industries and Aurignacian lithic assemblages, which are usually associated with Anatomical Modern Humans, which is consistent with an acculturation at distance model for these late Neanderthals. The Proto and Early Aurignacian appear contemporaneous indicating that this transition was rapid in this region. Anatomically Modern Humans are present at the site of Les Cottés at least at 39,500 cal BP roughly coincident with the onset of the cold phase Heinrich 4.     

The Characterization of Biologically Available Strontium Isotope Ratios for the Study of Prehistoric Migration

Strontium isotope analysis of bone and tooth enamel from prehistoric human skeletons is an important new technique used to address questions regarding migration. Two problems arise in such investigations: (1) levels of strontium isotope ratios in local bedrock, soil, water, plants and animals are variable; and (2) a range of values in human bone and enamel data make it difficult to distinguish some migrants from locals. Analysis of the bones of small animals provides a robust measure of local strontium isotope ratios and a reliable, if conservative, means for determining confidence limits for distinguishing migrants. Data from various geographical areas are presented here in a discussion of variability in strontium isotope values. Examples are provided using modern and prehistoric materials. We conclude with the recommendation that studies involving strontium isotope analysis should incorporate small animal samples for comparative purposes whenever possible.     

Lead isotope ratios: Tracking the migration of European‐Americans to Grafton,Illinois in the 19th century

Lead isotope analysis was conducted using inductively coupled plasma mass spectrometer (ICP‐MS) instruments on local soil samples and human premolar tooth enamel from a 19th century population from Grafton, Illinois, USA. The goal of the study was to determine if lead isotope analysis could be used to infer place of birth and patterns of 19th century migration into the city of Grafton. Five soil core samples from a location near Grafton, Illinois, five grave soil samples from the city cemetery and the tooth enamel of 19 human premolars were analysed. The results of the soil core analysis indicated that the lead isotopic signature of Grafton differs significantly from isotope ratios of other geographic areas associated with recorded places of birth of 19th century Jersey County residents. Elemental and isotope analysis of the soil samples indicated that diagenesis was not a factor in the analysis of lead isotopic signatures of enamel. From the lead isotope analysis of human premolars, the geographic origin of 13 of the remaining 15 individuals could be inferred. The inferred geographic origin was supplemented by an analysis of 1860 mortality and census records and demonstrated the utility of using lead isotope analysis in bio‐archaeological investigations. Copyright © 2010 John Wiley & Sons, Ltd.     

On Neanderthal subsistence strategies and land use: A regional focus on the Rhone Valley area in southeastern France

This study of Middle Palaeolithic assemblages from the Rhone Valley, in the South-East of France, increases our understanding of Neanderthal subsistence strategies and modes of territorial organisation by comparing a wide corpus of human occupations in limited chronological and geographical frameworks. The Neanderthal occupation modes may be examined using sites located in a reduced area, linking medium-altitude territories (Massif Central and the Alpine foothills) to the Rhone corridor. Through the combined analysis of the occupation levels of ten sites, all dated to between Marine Isotopic Stage (MIS) 7 and the beginning of MIS 3, we identify three types of occupation durations: (1) type 1 as long-term residential camps, (2) type 2 as short-term regular hunting camps and (3) type 3 as brief stopover camps. We discuss this variability of habitat types according to various parameters: site age, technical behaviour, environmental conditions, and site localisation and occupation seasons. The aim is to discern the underlying motivations behind Neanderthal group mobility. One of the main features of the Rhone Valley area is the great homogeneity of behaviours reflected within the sequences. This homogeneity, linked to the variability of the site occupations, supports the hypothesis of Neanderthal groups anticipating their land use requirements, and furthermore suggests that another type of circulating model was used in this area.     

非洲与现代人的起源

德国慕尼黑和美国宾夕法尼亚两个实验室的科学研究小组,在没有研究一根尼安德特人骨头的情况下,可能一劳永逸地解决了尼人问题。这两个实验室的专家称,他们在1856年发现的尼人骨架上提取了尼人的...     

Hunted or Scavenged Neanderthals? Taphonomic Approach to Hominin Fossils with Carnivore Damage

Carnivore damage on Neanderthal fossils is a much more common taphonomic modification than previously thought. Its presence could have different explanations, including predatory attacks or scavenging scenarios, which are both situations with important implications concerning Neanderthal behaviour. In the present paper, we analyse several Neanderthal hominin fossils from a taphonomic and forensic perspective in order to infer the nature of the modifications observed on the bone surfaces. Fossils displaying carnivore modifications from Spain, Germany, Belgium and Greece are evaluated from a taphonomic perspective for the first time in a significant sample of hominin specimens. Our results show that the materials analysed have been modified by small to large carnivores and that both attacks and strictly carnivore scavenging events can be inferred. This study also points out the importance of developing taphonomic approaches to the analysis of hominin bone surfaces, which can contribute significantly to knowledge of several aspects of Neanderthal behaviour. Copyright © 2017 John Wiley & Sons, Ltd.     

Preservation assessment of Miocene–Pliocene tooth enamel from Tugen Hills (Kenyan Rift Valley) through FTIR,chemical and stable-isotope analyses

We investigated fossil tooth enamel of mammals and crocodiles from two Mio-Pliocene East-African formations (Lukeino and Mabaget Fms) using infrared spectroscopy and chemical and stable-isotope analyses. Infrared spectra indicate that the fossil enamel contains biological apatite (bioapatite), without significant secondary carbonate contaminations. Several empirical infrared indexes were used to analyze the crystal–chemical characteristics of enamel. Fossil enamel has less organic matter, water and structural carbonate of apatite than modern enamel with which it was compared. Fossil apatite has a better crystallinity than bioapatite. The calcium/phosphorus mass ratio and the fluorine content of fossil apatite show intermediate values between bioapatite and geological fluorapatite. The samples also display significant crystal-chemical variations, depending on the vertebrate group (mammals vs. reptiles) and the taphonomic context (Lukeino Fm vs. Mabaget Fm). In spite of these changes, no relationship was observed between the chemical contents (carbonate and fluorine) and the stable-isotope ratios of carbonate (δ¹³C and δ¹⁸O) in fossil enamel. Preservation of the palaeoenvironmental signals is strongly supported by the fact that the stable-isotope composition of the three investigated fossil mammalian taxa (deinotheres, equids and hippos) is consistent with their ecological features. For instance, typical C₃- and C₄-plant isotope compositions are reflected in the deinotheres and equids, respectively, and amphibious hippos display lower δ¹⁸O values than terrestrial herbivores, as expected.     

Stable isotopes and human diet in central western Argentina

Central western Argentina is the southern boundary of prehispanic maize in the archaeological record. This paper explores the stable isotope information from human bone (δ¹³C, from collagen and δ¹³C from apatite) and tooth (δ¹³C from enamel) samples in order to characterize the temporal and spatial trends of these isotopes and to understand them in terms of human diet. We analyzed 104 human individuals from the last 6000 years. Using the Kellner and Schoeninger (2007) model the results shows a high variation in human diet with few human individuals with a notably high significance of C₄ plants (probably maize) as an energy source and these samples have a chronology around ca. 1000 years BP. On the other hand a high variation in an individual’s dietary life history is shown by differences between δ¹³C collagen, apatite and enamel.     

Strontium isotopes and human mobility in prehistoric Denmark

The principles behind the strontium isotopic system are an important tool for archaeologists tracing human migration and patterns of movement in prehistory. However, there are several scientific challenges of analytical nature, as well as those which relate to unknown parameters inherent to the interpretation of such data. One prerequisite is the knowledge of the range of strontium isotopic ratios that best characterize the bioavailable fractions of a particular area of interest. The study reported here attempts to establish a baseline for strontium isotope signatures valuable for Denmark (excluding the island of Bornholm) and particularly for the use in archaeological investigations. We present strontium isotope ratios of bones and teeth from modern mice contained in owl pellets, of snail shells, and of archaeological fauna samples. We compare these ratios with median strontium isotope signatures characterizing human enamel populations from archaeological sites within Denmark. The fauna samples reported here range from ⁸⁷Sr/⁸⁶Sr = 0.70717 to 0.71185 with an average of 0.70919, and human enamel defines a range from ⁸⁷Sr/⁸⁶Sr = 0.7086 to 0.7110 with an average of 0.7098. In both datasets, we observe a small difference between the baseline values for the western (Jutland) and eastern (Funen, Zealand, and the southern islands) parts of Denmark. We therefore propose two slightly different baseline ranges with a partial overlap for the isotopic signatures of bioavailable strontium fractions within Denmark, namely a range of ⁸⁷Sr/⁸⁶Sr = 0.7078–0.7098 for the western area and a range of ⁸⁷Sr/⁸⁶Sr = 0.7089–0.7108 for the eastern parts.     

EXAMINING CHILDHOOD DIETS AT KAMINALJUYU,GUATEMALA, THROUGH STABLE ISOTOPIC ANALYSIS OF SEQUENTIAL ENAMEL MICROSAMPLES*

Stable carbon and oxygen isotope ratios of tooth enamel provide a means to examine nutritional changes during childhood. To date, such studies have used large enamel samples, often spanning the developmental period of the tooth. This paper reports the results of small samples drilled from first molars and premolars of human teeth from Kaminaljuyu, a Classic period city in highland Guatemala. Carbon isotopes show considerable increase between cuspal and cervical enamel for both teeth, indicating an increase in maize consumption during childhood. Oxygen isotope trends are more variable, and suggest some seasonal fluctuation. While the oxygen isotope data confirm the identification of foreign skeletons among the samples, they illustrate a need for caution when sampling teeth due to variable δ¹⁸O composition of enamel within a tooth.     

Comments on Richards et al., Journal of Archaeological Science 35, 2008 “Strontium isotope evidence of Neanderthal mobility at the site of Lakonis,Greece using laser-ablation PIMMS”

We present an evaluation of the laser ablation Sr isotope data reported by Richards et al. [Richards, M., Harvati, K., Grimes, V., Colin Smith, C., Smith, T., Jean-Jacques Hublin, J.J., Karkanas, P., and Panagopoulou, E., 2008. Strontium isotope evidence of Neanderthal mobility at the site of Lakonis, Greece using laser ablation PIMMS. Journal of Archaeological Science 35, 1251–1256] for a Neanderthal tooth recovered from a site in Greece. Based on an alternative and analytically more robust method of correcting for isobaric interferences present during the analysis the tooth appears to be isotopically homogenous and within uncertainty of the value for modern seawater. If this is the case then contrary to the migration model proposed by Richards et al. [Richards, M., Harvati, K., Grimes, V., Colin Smith, C., Smith, T., Jean-Jacques Hublin, J.J., Karkanas, P., and Panagopoulou, E., 2008. Strontium isotope evidence of Neanderthal mobility at the site of Lakonis, Greece using laser ablation PIMMS. Journal of Archaeological Science 35, 1251–1256] the Neanderthal individual may have actually been a coastal dweller and lived within the vicinity of the find site.