Inter- and intra-observer variation in phytolith morphometry.

BACKGROUND: Archaeobotanists and palaeoecologists extensively use geometric morphometrics to identify plant opal phytoliths. Particularly when applied to assemblages of phytoliths from concentrations retrieved from closed contexts, morphometric data from archaeological phytoliths compared with similar data from reference material may allow taxonomic attribution. Observer variation is one aspect of phytolith morphometry that has received little attention but may be an important source of error, and hence cause of potential misidentification of plant remains. SCOPE: To investigate inter- and intra-observer variation in phytolith morphometry, eight researchers (observers) from different laboratories measured 50 samples each from three phytolith morphotypes, Bilobate, Bulliform flabellate and Elongate dendritic, three times, under the auspices of the International Committee for Phytolith Morphometrics (ICPM). METHODS: Data for 17 size and shape variables were collected for each phytolith by manually digitising a phytolith outline (mask) from a photograph, followed by measurement of the mask with open-source morphometric software. KEY RESULTS: Inter-observer variation ranged from 0 to 23% difference from the mean of all observers. Intra-observer variation ranged from 0 to 9% difference from the mean of individual observers per week. Inter- and intra-observer variation was generally higher among inexperienced researchers. CONCLUSIONS: Scaling errors were a major cause of variation and occurred more with less experienced researchers, which is likely related to familiarity with data collection. The results indicate that inter- and intra-observer variation can be substantially reduced by providing clear instructions for and training with the equipment, photo capturing, software, data collection and data cleaning. In this paper, the ICPM provides recommendations to minimise variation.Advances in automatic data collection may eventually reduce inter- and intra-observer variation, but until this is common practice, the ICPM recommends that phytolith morphometric analyses adhere to standardised guidelines to assure that measured phytolith variables are accurate, consistent and comparable between different researchers and laboratories.

Exotic foods reveal contact between South Asia and the Near East during the second millennium BCE.

Although the key role of long-distance trade in the transformation of cuisines worldwide has been well-documented since at least the Roman era, the prehistory of the Eurasian food trade is less visible. In order to shed light on the transformation of Eastern Mediterranean cuisines during the Bronze Age and Early Iron Age, we analyzed microremains and proteins preserved in the dental calculus of individuals who lived during the second millennium BCE in the Southern Levant. Our results provide clear evidence for the consumption of expected staple foods, such as cereals (Triticeae), sesame (Sesamum), and dates (Phoenix). We additionally report evidence for the consumption of soybean (Glycine), probable banana (Musa), and turmeric (Curcuma), which pushes back the earliest evidence of these foods in the Mediterranean by centuries (turmeric) or even millennia (soybean). We find that, from the early second millennium onwards, at least some people in the Eastern Mediterranean had access to food from distant locations, including South Asia, and such goods were likely consumed as oils, dried fruits, and spices. These insights force us to rethink the complexity and intensity of Indo-Mediterranean trade during the Bronze Age as well as the degree of globalization in early Eastern Mediterranean cuisine.

The evolution and changing ecology of the African hominid oral microbiome.

The oral microbiome plays key roles in human biology, health, and disease, but little is known about the global diversity, variation, or evolution of this microbial community. To better understand the evolution and changing ecology of the human oral microbiome, we analyzed 124 dental biofilm metagenomes from humans, including Neanderthals and Late Pleistocene to present-day modern humans, chimpanzees, and gorillas, as well as New World howler monkeys for comparison. We find that a core microbiome of primarily biofilm structural taxa has been maintained throughout African hominid evolution, and these microbial groups are also shared with howler monkeys, suggesting that they have been important oral members since before the catarrhine-platyrrhine split ca. 40 Mya. However, community structure and individual microbial phylogenies do not closely reflect host relationships, and the dental biofilms of Homo and chimpanzees are distinguished by major taxonomic and functional differences. Reconstructing oral metagenomes from up to 100 thousand years ago, we show that the microbial profiles of both Neanderthals and modern humans are highly similar, sharing functional adaptations in nutrient metabolism. These include an apparent Homo-specific acquisition of salivary amylase-binding capability by oral streptococci, suggesting microbial coadaptation with host diet. We additionally find evidence of shared genetic diversity in the oral bacteria of Neanderthal and Upper Paleolithic modern humans that is not observed in later modern human populations. Differences in the oral microbiomes of African hominids provide insights into human evolution, the ancestral state of the human microbiome, and a temporal framework for understanding microbial health and disease.

Dietary evidence from Central Asian Neanderthals: A combined isotope and plant microremains approach at Chagyrskaya Cave (Altai, Russia).

Neanderthals are known primarily from their habitation of Western Eurasia, but they also populated large expanses of Northern Asia for thousands of years. Owing to a sparse archaeological record, relatively little is known about these eastern Neanderthal populations. Unlike in their western range, there are limited zooarchaeological and paleobotanical studies that inform us about the nature of their subsistence. Here, we perform a combined analysis of carbon and nitrogen stable isotopes on bone collagen and microbotanical remains in dental calculus to reconstruct the diet of eastern Neanderthals at Chagyrskaya Cave in the Altai Mountains of Southern Siberia, Russia. Stable isotopes identify one individual as possessing a high trophic level due to the hunting of large- and medium-sized ungulates, while the analysis of dental calculus also indicates the presence of plants in the diet of this individual and others from the site. These findings indicate eastern Neanderthals may have had broadly similar subsistence patterns to those elsewhere in their range.

New insights on Neolithic food and mobility patterns in Mediterranean coastal populations.

OBJECTIVES: The aims of this research are to explore the diet, mobility, social organization, and environmental exploitation patterns of early Mediterranean farmers, particularly the role of marine and plant resources in these foodways. In addition, this work strives to document possible gendered patterns of behavior linked to the neolithization of this ecologically rich area. To achieve this, a set of multiproxy analyses (isotopic analyses, dental calculus, microremains analysis, ancient DNA) were performed on an exceptional deposit (n = 61) of human remains from the Les Bréguières site (France), dating to the transition of the sixth to the fifth millennium BCE. MATERIALS AND METHODS: The samples used in this study were excavated from the Les Bréguières site (Mougins, Alpes-Maritimes, France), located along the southeastern Mediterranean coastline of France. Stable isotope analyses (C, N) on bone collagen (17 coxal bones, 35 craniofacial elements) were performed as a means to infer protein intake during tissue development. Sulfur isotope ratios were used as indicators of geographical and environmental points of origin. The study of ancient dental calculus helped document the consumption of plants. Strontium isotope analysis on tooth enamel (n = 56) was conducted to infer human provenance and territorial mobility. Finally, ancient DNA analysis was performed to study maternal versus paternal diversity within this Neolithic group (n = 30). RESULTS: Stable isotope ratios for human bones range from -20.3 to -18.1‰ for C, from 8.9 to 11.1‰ for N and from 6.4 to 15‰ for S. Domestic animal data range from -22.0 to -20.2‰ for C, from 4.1 to 6.9‰ for N, and from 10.2 to 12.5‰ for S. Human enamel 87 Sr/86 Sr range from 0.7081 to 0.7102, slightly wider than the animal range (between 0.7087 and 0.7096). Starch and phytolith microremains were recovered as well as other types of remains (e.g., hairs, diatoms, fungal spores). Starch grains include Triticeae type and phytolith includes dicotyledons and monocot types as panicoid grasses. Mitochondrial DNA characterized eight different maternal lineages: H1, H3, HV (5.26%), J (10.53%), J1, K, T (5.2%), and U5 (10.53%) but no sample yielded reproducible Y chromosome SNPs, preventing paternal lineage characterization. DISCUSSION: Carbon and nitrogen stable isotope ratios indicate a consumption of protein by humans mainly focused on terrestrial animals and possible exploitation of marine resources for one male and one undetermined adult. Sulfur stable isotope ratios allowed distinguishing groups with different geographical origins, including two females possibly more exposed to the sea spray effect. While strontium isotope data do not indicate different origins for the individuals, mitochondrial lineage diversity from petrous bone DNA suggests the burial includes genetically differentiated groups or a group practicing patrilocality. Moreover, the diversity of plant microremains recorded in dental calculus provide the first evidence that the groups of Les Bréguières consumed a wide breadth of plant foods (as cereals and wild taxa) that required access to diverse environments. This transdisciplinary research paves the way for new perspectives and highlights the relevance for novel research of contexts (whether recently discovered or in museum collections) excavated near shorelines, due to the richness of the biodiversity and the wide range of edible resources available.

Dental calculus indicates widespread plant use within the stable Neanderthal dietary niche.

The ecology of Neanderthals is a pressing question in the study of hominin evolution. Diet appears to have played a prominent role in their adaptation to Eurasia. Based on isotope and zooarchaeological studies, Neanderthal diet has been reconstructed as heavily meat-based and generally similar across different environments. This image persists, despite recent studies suggesting more plant use and more variation. However, we have only a fragmentary picture of their dietary ecology, and how it may have varied among habitats, because we lack broad and environmentally representative information about their use of plants and other foods. To address the problem, we examined the plant microremains in Neanderthal dental calculus from five archaeological sites representing a variety of environments from the northern Balkans, and the western, central and eastern Mediterranean. The recovered microremains revealed the consumption of a variety of non-animal foods, including starchy plants. Using a modeling approach, we explored the relationships among microremains and environment, while controlling for chronology. In the process, we compared the effectiveness of various diversity metrics and their shortcomings for studying microbotanical remains, which are often morphologically redundant for identification. We developed Minimum Botanical Units as a new way of estimating how many plant types or parts are present in a microbotanical sample. In contrast to some previous work, we found no evidence that plant use is confined to the southern-most areas of Neanderthal distribution. Although interpreting the ecogeographic variation is limited by the incomplete preservation of dietary microremains, it is clear that plant exploitation was a widespread and deeply rooted Neanderthal subsistence strategy, even if they were predominately game hunters. Given the limited dietary variation across Neanderthal range in time and space in both plant and animal food exploitation, we argue that vegetal consumption was a feature of a generally static dietary niche.

Earliest floral grave lining from 13,700-11,700-y-old Natufian burials at Raqefet Cave, Mt. Carmel, Israel.

Flowering plants possess mechanisms that stimulate positive emotional and social responses in humans. It is difficult to establish when people started to use flowers in public and ceremonial events because of the scarcity of relevant evidence in the archaeological record. We report on uniquely preserved 13,700-11,700-y-old grave linings made of flowers, suggesting that such use began much earlier than previously thought. The only potentially older instance is the questionable use of flowers in the Shanidar IV Neanderthal grave. The earliest cemeteries (ca. 15,000-11,500 y ago) in the Levant are known from Natufian sites in northern Israel, where dozens of burials reflect a wide range of inhumation practices. The newly discovered flower linings were found in four Natufian graves at the burial site of Raqefet Cave, Mt. Carmel, Israel. Large identified plant impressions in the graves include stems of sage and other Lamiaceae (Labiatae; mint family) or Scrophulariaceae (figwort family) species; accompanied by a plethora of phytoliths, they provide the earliest direct evidence now known for such preparation and decoration of graves. Some of the plant species attest to spring burials with a strong emphasis on colorful and aromatic flowers. Cave floor chiseling to accommodate the desired grave location and depth is also evident at the site. Thus, grave preparation was a sophisticated planned process, embedded with social and spiritual meanings reflecting a complex preagricultural society undergoing profound changes at the end of the Pleistocene.

Synchrotron radiation-based phase-contrast microtomography of human dental calculus allows nondestructive analysis of inclusions: implications for archeological samples.

Purpose: Dental calculus forms on teeth during the life of an individual and its investigation can yield information about diet, health status, and environmental pollution. Currently, the analytical techniques used to visualize the internal structure of human dental calculus and entrapped inclusions are limited and require destructive sampling, which cannot always be justified. Approach: We used propagation phase-contrast synchrotron radiation micro-computed tomography (PPC-SR- µ CT ) to non-destructively examine the internal organization of dental calculus, including its microstructure and entrapped inclusions, on both modern and archeological samples. Results: The virtual histological exploration of the samples shows that PPC-SR- µ CT is a powerful approach to visualize the internal organization of dental calculus. We identified several important features, including previously undetected negative imprints of enamel and dentine growth markers (perikymata and periradicular bands, respectively), the non-contiguous structure of calculus layers with multiple voids, and entrapped plant remains. Conclusions: PPC-SR- µ CT is an effective technique to explore dental calculus structural organization, and is especially powerful for enabling the identification of inclusions. The non-destructive nature of synchrotron tomography helps protect samples for future research. However, the irregular layers and frequent voids reveal a high heterogeneity and variability within calculus, with implications for research focusing on inclusions.

Ancient proteins provide evidence of dairy consumption in eastern Africa.

Consuming the milk of other species is a unique adaptation of Homo sapiens, with implications for health, birth spacing and evolution. Key questions nonetheless remain regarding the origins of dairying and its relationship to the genetically-determined ability to drink milk into adulthood through lactase persistence (LP). As a major centre of LP diversity, Africa is of significant interest to the evolution of dairying. Here we report proteomic evidence for milk consumption in ancient Africa. Using liquid chromatography tandem mass spectrometry (LC-MS/MS) we identify dairy proteins in human dental calculus from northeastern Africa, directly demonstrating milk consumption at least six millennia ago. Our findings indicate that pastoralist groups were drinking milk as soon as herding spread into eastern Africa, at a time when the genetic adaptation for milk digestion was absent or rare. Our study links LP status in specific ancient individuals with direct evidence for their consumption of dairy products.

Archaeometric evidence for the earliest exploitation of lignite from the bronze age Eastern Mediterranean.

This paper presents the earliest evidence for the exploitation of lignite (brown coal) in Europe and sheds new light on the use of combustion fuel sources in the 2nd millennium BCE Eastern Mediterranean. We applied Thermal Desorption/Pyrolysis-Gas Chromatography-Mass Spectrometry and Polarizing Microscopy to the dental calculus of 67 individuals and we identified clear evidence for combustion markers embedded within this calculus. In contrast to the scant evidence for combustion markers within the calculus samples from Egypt, all other individuals show the inhalation of smoke from fires burning wood identified as Pinaceae, in addition to hardwood, such as oak and olive, and/or dung. Importantly, individuals from the Palatial Period at the Mycenaean citadel of Tiryns and the Cretan harbour site of Chania also show the inhalation of fire-smoke from lignite, consistent with the chemical signature of sources in the northwestern Peloponnese and Western Crete respectively. This first evidence for lignite exploitation was likely connected to and at the same time enabled Late Bronze Age Aegean metal and pottery production, significantly by both male and female individuals.

CoproID predicts the source of coprolites and paleofeces using microbiome composition and host DNA content.

Shotgun metagenomics applied to archaeological feces (paleofeces) can bring new insights into the composition and functions of human and animal gut microbiota from the past. However, paleofeces often undergo physical distortions in archaeological sediments, making their source species difficult to identify on the basis of fecal morphology or microscopic features alone. Here we present a reproducible and scalable pipeline using both host and microbial DNA to infer the host source of fecal material. We apply this pipeline to newly sequenced archaeological specimens and show that we are able to distinguish morphologically similar human and canine paleofeces, as well as non-fecal sediments, from a range of archaeological contexts.

Ancient genomes reveal complex patterns of population movement, interaction, and replacement in sub-Saharan Africa.

Africa hosts the greatest human genetic diversity globally, but legacies of ancient population interactions and dispersals across the continent remain understudied. Here, we report genome-wide data from 20 ancient sub-Saharan African individuals, including the first reported ancient DNA from the DRC, Uganda, and Botswana. These data demonstrate the contraction of diverse, once contiguous hunter-gatherer populations, and suggest the resistance to interaction with incoming pastoralists of delayed-return foragers in aquatic environments. We refine models for the spread of food producers into eastern and southern Africa, demonstrating more complex trajectories of admixture than previously suggested. In Botswana, we show that Bantu ancestry post-dates admixture between pastoralists and foragers, suggesting an earlier spread of pastoralism than farming to southern Africa. Our findings demonstrate how processes of migration and admixture have markedly reshaped the genetic map of sub-Saharan Africa in the past few millennia and highlight the utility of combined archaeological and archaeogenetic approaches.

Dental calculus evidence of Taï Forest Chimpanzee plant consumption and life history transitions.

Dental calculus (calcified dental plaque) is a source of multiple types of data on life history. Recent research has targeted the plant microremains preserved in this mineralised deposit as a source of dietary and health information for recent and past populations. However, it is unclear to what extent we can interpret behaviour from microremains. Few studies to date have directly compared the microremain record from dental calculus to dietary records, and none with long-term observation dietary records, thus limiting how we can interpret diet, food acquisition and behaviour. Here we present a high-resolution analysis of calculus microremains from wild chimpanzees (Pan troglodytes verus) of Taï National Park, Côte d'Ivoire. We test microremain assemblages against more than two decades of field behavioural observations to establish the ability of calculus to capture the composition of diet. Our results show that some microremain classes accumulate as long-lived dietary markers. Phytolith abundance in calculus can reflect the proportions of plants in the diet, yet this pattern is not true for starches. We also report microremains can record information about other dietary behaviours, such as the age of weaning and learned food processing techniques like nut-cracking.