The role of birds at Çatalhöyük revealed by the analysis of eggshell

The exceptional eggshell assemblage from Çatalhöyük was studied using an integrated approach combining morphology (by optical and scanning electron microscopy) and palaeoproteomics (by mass spectrometry). We provide taxonomic classification for 90 fragments, of which only 11 remain undetermined. The striking predominance of Anseriformes (probably including greylag geese, as well as ducks and swans) in all types of deposits examined, including middens and burial fills, suggests that these eggs were exploited as food and, at the same time, had a special significance for the inhabitants of the site. We detected the presence of crane eggshell in contexts associated with both the world of the living and the world of the dead (consistent with the well-known importance of this bird at Çatalhöyük), as well as the possible but infrequent occurrence of bustards and herons. Overall, we suggest that eggshell analysis, together with osteological data and the archaeological context, can provide the basis for a nuanced understanding of the relationship between humans and birds in the past

Threads of memory: Reviving the ornament of a dead child at the Neolithic village of Ba`ja (Jordan)

In 2018, a well-constructed cist-type grave was discovered at Ba`ja, a Neolithic village (7,400–6,800 BCE) in Southern Jordan. Underneath multiple grave layers, an 8-year-old child was buried in a fetal position. Over 2,500 beads were found on the chest and neck, along with a double perforated stone pendant and a delicately engraved mother-of-pearl ring discovered among the concentration of beads. The first was found behind the neck, and the second on the chest. The meticulous documentation of the bead distribution indicated that the assemblage was a composite ornament that had gradually collapsed, partly due to the burying position. Our aim was to challenge time degradation and to reimagine the initial composition in order to best explore the significance of this symbolic category of material culture, not as mere group of beads, but as an ornamental creation with further aesthetic, artisanal and socioeconomic implications. The reconstruction results exceeded our expectations as it revealed an imposing multi-row necklace of complex structure and attractive design. Through multiple lines of evidence, we suggest that the necklace was created at Ba`ja, although significant parts of beads were made from exotic shells and stones, including fossil amber, an unprecedented material never attested before for this period. The retrieval of such an ornament from life and its attribution to a young dead child highlights the significant social status of this individual. Beyond the symbolic functions related to identity, the necklace is believed to have played a key role in performing the inhumation rituals, understood as a public event gathering families, relatives, and people from other villages. In this sense, the necklace is not seen as belonging completely to the realm of death but rather to the world of the living, materializing a collective memory and shared moments of emotions and social cohesion

Threads of memory: Reviving the ornament of a dead child at the Neolithic village of Ba'ja (Jordan)

In 2018, a well-constructed cist-type grave was discovered at Ba'ja, a Neolithic village (7,400-6,800 BCE) in Southern Jordan. Underneath multiple grave layers, an 8-year-old child was buried in a fetal position. Over 2,500 beads were found on the chest and neck, along with a double perforated stone pendant and a delicately engraved mother-of-pearl ring discovered among the concentration of beads. The first was found behind the neck, and the second on the chest. The meticulous documentation of the bead distribution indicated that the assemblage was a composite ornament that had gradually collapsed, partly due to the burying position. Our aim was to challenge time degradation and to reimagine the initial composition in order to best explore the significance of this symbolic category of material culture, not as mere group of beads, but as an ornamental creation with further aesthetic, artisanal and socioeconomic implications. The reconstruction results exceeded our expectations as it revealed an imposing multi-row necklace of complex structure and attractive design. Through multiple lines of evidence, we suggest that the necklace was created at Ba'ja, although significant parts of beads were made from exotic shells and stones, including fossil amber, an unprecedented material never attested before for this period. The retrieval of such an ornament from life and its attribution to a young dead child highlights the significant social status of this individual. Beyond the symbolic functions related to identity, the necklace is believed to have played a key role in performing the inhumation rituals, understood as a public event gathering families, relatives, and people from other villages. In this sense, the necklace is not seen as belonging completely to the realm of death but rather to the world of the living, materializing a collective memory and shared moments of emotions and social cohesion.ArchaeologyHub.CSIC 2022 Internal Research Grant (HA) https://archaeologyhub.csic.es/ - H2020 Marie Sklodowska-Curie Actions, grant number 846097 (HA) https://cordis.europa.eu/project/id/846097 - German Research Foundation (BO 1599/14-1; BO 1599/16-1) (MB, HGG) https://www.dfg.de/en/ - Franz-and Eva Rutzen Stiftung Foundation (MB) https://www.deutsches-stiftungszentrum.de/stiftungen/franz-und-eva-rutzen-stiftung - Junta de Andalucía (Consejería de Economía, Conocimiento, Empresas y Universidad), under contract P20_01080 (CPO) https://www.juntadeandalucia.es/organismos/universidadinvestigacioneinnovacion.htm

'Palaeoshellomics' reveals the use of freshwater mother-of-pearl in prehistory

The extensive use of mollusc shell as a versatile raw material is testament to its importance in prehistoric times. The consistent choice of certain species for different purposes, including the making of ornaments, is a direct representation of how humans viewed and exploited their environment. The necessary taxonomic information, however, is often impossible to obtain from objects that are small, heavily worked or degraded. Here we propose a novel biogeochemical approach to track the biological origin of prehistoric mollusc shell. We conducted an in-depth study of archaeological ornaments using microstructural, geochemical and biomolecular analyses, including 'palaeoshellomics', the first application of palaeoproteomics to mollusc shells (and indeed to any invertebrate calcified tissue). We reveal the consistent use of locally-sourced freshwater mother-of-pearl for the standardized manufacture of 'double-buttons'. This craft is found throughout Europe between 4200-3800 BCE, highlighting the ornament-makers' profound knowledge of the biogeosphere and the existence of cross-cultural traditions

Approches protéomiques en archéologie et héritage culturel : caractérisation de protéines anciennes préservées dans des coquilles de mollusques

Archaeological mollusc shell artifacts and ecofacts are valuable sources to study past cultures and provide insights on how people exploited their environments. Mollusc shells were often used as raw materials to make personal ornaments and are abundantly found in archaeological sites. However, minute, heavily worked and/or fragmented shell ornaments are rarely identified at different taxonomic levels, due to limited availability of analytical approaches to determine taxon-specific (diagnostic) features. In recent years, proteomics, which exploits the high sensitivity of modern mass spectrometry techniques, has been successfully applied to the study of a variety of bioarchaeological remains, opening a new research field referred to as ‘palaeoproteomics’. While modern mollusc shells represent key study models in biomineralization research to investigate the molecular mechanisms of biocalcification and mineral deposition, palaeoproteomics on archaeological shells has never been carried out before. The challenges are mainly due to the intrinsic physico-chemical features of their substrates (including low protein content), and the paucity of protein sequences for reference.Therefore, this work presents a comprehensive proteomic-based investigation of archaeological mollusc shells, grounded in biomineralization research, and the first application of palaeoproteomics to shells. The main objective of this PhD was to investigate three key concepts: whether shell proteins carry taxonomic information, could be preserved over archaeological times scales and could be used as molecular barcodes for the taxonomic determination of ancient shell artifacts.High performance liquid chromatography coupled to tandem mass spectrometry (HPLC-MS/MS) was used for an in-depth proteomics characterisation of a selected model, Spondylus gaederopus, demonstrating that its shell-associated proteins are very different from other models and likely represent lineage-specific sequences. Peptide mass fingerprinting (PMF) by MALDI-TOF mass spectrometry was employed to obtain sequence information from thirty-four different molluscan taxa. Intracrystalline shell proteins displayed clearly distinct PMFs indicating that they may encode taxonomic information and could be used as molecular barcodes to identify archaeological mollusc shells. Accelerated aging experiments were performed to mimic the diagenesis of Spondylus shell and the stability of intracrystalline shell proteins was investigated by a combination of immunochemistry and quantitative TMT proteomics. We concluded that Spondylus represents a favourable system for protein preservation, as evidenced by the thermal stability of shell peptide sequences. However, the observed degradation patterns pinpointed the complexity of the whole diagenesis process, which does not entirely follow expected trajectories from theoretical models.Finally, a set of archaeological shell ornaments, recovered from different Mesolithic and Neolithic European prehistoric sites, were studied by proteomics coupled with structural, biomolecular and geochemical analyses. The studies showed that PMF by MALDI-TOF approach was not able to characterise these archaeological samples, which are very degraded or and/or have low protein contents, and the analysis of ancient shell proteins requires higher sensitivity, as offered by HPLC-MS/MS mass spectrometry. Indeed, palaeoproteomic analysis by HPLC-MS/MS of small “double-button” ornaments identified that they were made of freshwater mother-of-pearl shells, resolving the long-standing debate over their biological origin and provenance. The research carried out in this thesis highlights the immense potential of proteomics-based approaches to study archaeological and palaeontological shell remains.Les coquilles de mollusques ont très souvent été employées par les populations préhistoriques, comme source de matière première, à la confection d'ornements vestimentaires ou de parures. Ces objets, largement transformés, constituent un témoignage inestimable pour comprendre les cultures passées et la façon dont des populations disparues ont exploité leur environnement naturel. Abondamment retrouvés dans les sites archéologiques, les restes coquilliers sont le plus souvent minuscules, modifiés ou bien retrouvés à l'état de fragments, à un point tel qu'il est impossible de leur assigner une appartenance taxonomique.Depuis quelques années, les approches protéomiques, basées sur la sensibilité des techniques de spectrométrie de masse, ont été utilisées pour l'étude de restes bio-archéologiques, ouvrant ainsi un champ disciplinaire nouveau, la paléoprotéomique. Alors que la protéomique est classiquement employée sur des coquilles modernes pour en comprendre les mécanismes moléculaires de biominéralisation, la paléoprotéomique de fragments de coquilles archéologiques n'a jamais été réalisée jusqu'à présent, pour plusieurs raisons: faibles quantités d'échantillons (et, par conséquent, de protéines), données de séquences très parcellaires sur les protéines coquillières de mollusques.Ce travail de thèse représente donc la première tentative d'analyse des protéines coquillières d'échantillons archéologiques par une approche protéomique, telle que réalisée dans le domaine de la biominéralisation. Trois concepts-clés sont mis en exergue: les protéines de coquille contiennent de l'information taxonomique; elles peuvent être préservées à des échelles de temps archéologiques; elles peuvent être utilisées comme "code-barre" moléculaire pour identifier les taxons dont sont issus les restes archéologiques.En premier lieu, l'approche protéomique (HPLC-MS/MS) a été testée sur la coquille d'un bivalve-modèle, Spondylus gaederopus, pour en caractériser son contenu protéique. Nos données indiquent que le répertoire protéique coquillier de cette espèce diffère de celui de formes voisines et comprend des séquences "taxon-spécifiques". En second lieu, des "empreintes de masse peptidique" (PMF en anglais) par MALDI-TOF ont été réalisées sur les coquilles récentes de 34 espèces de mollusques, couvrant un large spectre de taxons trouvés sur des sites archéologiques. Les spectres PMF obtenus ont une signature particulière, contiennent une information taxonomique, un "code-barre", applicable à des échantillons archéologiques. En troisième lieu, des expériences de vieillissement accéléré ont simulé la diagenèse de coquilles de Spondylus. La stabilité des protéines coquillières a été étudiée en combinant immunochimie et protéomique quantitative. Les données démontrent que Spondylus est un bon modèle, favorable à la préservation des protéines de coquilles. Certains processus chimiques de clivage peptidique ont pu être analysés finement. Les patrons de dégradation des protéines sont cependant complexes, une complexité non prédite par les modèles théoriques. Enfin, des ornements archéologiques, provenant de divers sites européens mésolithiques et néolithiques ont été étudiés par protéomique couplée à des analyses microstructurales et géochimiques. La réalisation d'empreintes de masse peptidique (PMF) n'a pas permis de lever le voile sur l'identité taxonomique des échantillons, sans doute du fait des très faibles quantités de protéines restantes, ces échantillons requérant des analyses plus complètes par HPLC-MS/MS. Par contre, les données obtenues sur de minuscules boutons-doubles ont clairement montré que ces derniers ont été fabriqués à partir de nacre de moules d'eau douce (unionidés) largement réparties en Europe

Shell palaeoproteomics: first application of peptide mass fingerprinting for the rapid identification of mollusc shells in archaeology.

10 pagesInternational audienceMolluscs were one of the most widely-used natural resources in the past, and their shells are abundant among archaeological findings. However, our knowledge of the variety of shells that were circulating in prehistoric times (and thus their socio-economic and cultural value) is scarce due to the difficulty of achieving taxonomic determination of fragmented and/or worked remains. This study aims to obtain molecular barcodes based on peptide mass fingerprints (PMFs) of intracrystalline proteins, in order to obtain shell identification. Palaeoproteomic applications on shells are challenging, due to low concentration of molluscan proteins and an incomplete understanding of their sequences. We explore different approaches for protein extraction from small-size samples (<20 mg), followed by MALDI-TOF-MS analysis. The SP3 (single-pot, solid-phase) sample preparation method was found to be the most successful in retrieving the intracrystalline protein fraction from seven molluscan shell taxa, which belong to different phylogenetic groups, possess distinct microstructures and are relevant for archaeology. Furthermore, all the shells analysed, including a 7000-year-old specimen of the freshwater bivalve Pseudunio, yielded good-quality distinctive spectra, demonstrating that PMFs can be used for shell taxon determination. Our work suggests good potential for large-scale screening of archaeological molluscan remains. SIGNIFICANCE: We characterise for the first time the peptide mass fingerprints of the intracrystalline shell protein fraction isolated from different molluscan taxa. We demonstrate that these proteins yield distinctive PMFs, even for shells that are phylogenetically related and/or that display similar microstructures. Furthermore, we extend the range of sample preparation approaches for "shellomics" by testing the SP3 method, which proved to be well-suited to shell protein extraction from small-size and protein-poor samples. This work thus lays the foundations for future large-scale applications for the identification of mollusc shells and other invertebrate remains from the archaeological and palaeontological records

The role of birds at Çatalhöyük revealed by the analysis of eggshell.

11 pagesInternational audienceThe exceptional eggshell assemblage from Çatalhöyük was studied using an integrated approach combining morphology (by optical and scanning electron microscopy) and palaeoproteomics (by mass spectrometry). We provide taxonomic classification for 90 fragments, of which only 11 remain undetermined. The striking predominance of Anseriformes (probably including greylag geese, as well as ducks and swans) in all types of deposits examined, particularly middens and burial fills, suggests that these eggs were exploited as food and, at the same time, had a special significance for the inhabitants of the site. We detected the presence of crane eggshell in contexts associated with both the world of the living and the world of the dead (consistent with the well-known importance of this bird at Çatalhöyük), as well as the possible but infrequent occurrence of bustards and herons. Overall, we suggest that eggshell analysis, together with osteological data and the archaeological context, can provide the basis for a nuanced understanding of the relationship between humans and birds in the past

The role of birds at Çatalhöyük revealed by the analysis of eggshell

The exceptional eggshell assemblage from Çatalhöyük was studied using an integrated approach combining morphology (by optical and scanning electron microscopy) and palaeoproteomics (by mass spectrometry). We provide taxonomic classification for 90 fragments, of which only 11 remain undetermined. The striking predominance of Anseriformes (probably including greylag geese, as well as ducks and swans) in all types of deposits examined, including middens and burial fills, suggests that these eggs were exploited as food and, at the same time, had a special significance for the inhabitants of the site. We detected the presence of crane eggshell in contexts associated with both the world of the living and the world of the dead (consistent with the well-known importance of this bird at Çatalhöyük), as well as the possible but infrequent occurrence of bustards and herons. Overall, we suggest that eggshell analysis, together with osteological data and the archaeological context, can provide the basis for a nuanced understanding of the relationship between humans and birds in the past