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    The soil in our microbial DNA informs about environmental interfaces across host and subsistence modalities. Schnorr Stephanie L Philosophical transactions of the Royal Society of London. Series B, Biological sciences In this study, I use microbiome datasets from global soil samples and diverse hosts to learn whether soil microbial taxa are found in host microbiomes, and whether these observations fit the narrative that environmental interaction influences human microbiomes. A major motivation for conducting host-associated microbiome research is to contribute towards understanding how the environment may influence host physiology. The microbial molecular network is considered a key vector by which environmental traits may be transmitted to the host. Research on human evolution seeks evidence that can inform about the living experiences of human ancestors. This objective is substantially enhanced by recent work on ancient biomolecules from preserved microbial tissues, such as dental calculus, faecal sediments and whole coprolites. A challenge yet is to distinguish authentic biomolecules from environmental contaminants deposited contemporaneously, primarily from soil. However, we do not have sound expectations about the soil microbial elements arriving to host-associated microbiomes in a modern context. One assumption in human microbiome research is that proximity to the natural environment should affect biodiversity or impart genetic elements. I present evidence supporting the assumption that environmental soil taxa are found among host-associated gut taxa, which can recapitulate the surrounding host habitat ecotype. Soil taxa found in gut microbiomes relate to a set of universal 'core' taxa for all soil ecotypes, demonstrating that widespread host organisms may experience a consistent pattern of external environmental cues, perhaps critical for development. Observed differentiation of soil feature diversity, abundance and composition among human communities, great apes and invertebrate hosts also indicates that lifestyle patterns are inferable from an environmental signal that is retrievable from gut microbiome amplicon data. This article is part of the theme issue 'Insights into health and disease from ancient biomolecules'. 10.1098/rstb.2019.0577
    Intestinal helminths as a biomolecular complex in archaeological research. Flammer Patrik G,Smith Adrian L Philosophical transactions of the Royal Society of London. Series B, Biological sciences Enteric helminths are common parasites in many parts of the world and in the past were much more widespread both geographically and socially. Many enteric helminths are relatively long-lived in the human host, often benign or of low pathogenicity while producing large numbers of environmentally resistant eggs voided in the faeces or found associated with individual remains (skeletons and mummies). The combination of helminth characters offers opportunities to the field of historical pathogen research that are quite different to that of some of the more intensively studied high impact pathogens. Historically, a wealth of studies has employed microscopic techniques to diagnose infection using the morphology of the helminth eggs. More recently, various ancient DNA (aDNA) approaches have been applied in the archaeoparasitological context and these are revolutionizing the field, allowing much more specific diagnosis as well as interrogating the epidemiology of helminths. These advances have enhanced the potential for the field to provide unique information on past populations including using diseases to consider many aspects of life (e.g. sanitation, hygiene, diet, culinary practices and other aspects of society). Here, we consider the impact of helminth archaeoparasitology and more specifically the impact and potential for application of aDNA technologies as a part of the archaeologists' toolkit. This article is part of the theme issue 'Insights into health and disease from ancient biomolecules'. 10.1098/rstb.2019.0570
    Ancient RNA virus epidemics through the lens of recent adaptation in human genomes. Enard David,Petrov Dmitri A Philosophical transactions of the Royal Society of London. Series B, Biological sciences Over the course of the last several million years of evolution, humans probably have been plagued by hundreds or perhaps thousands of epidemics. Little is known about such ancient epidemics and a deep evolutionary perspective on current pathogenic threats is lacking. The study of past epidemics has typically been limited in temporal scope to recorded history, and in physical scope to pathogens that left sufficient DNA behind, such as during the Great Plague. Host genomes, however, offer an indirect way to detect ancient epidemics beyond the current temporal and physical limits. Arms races with pathogens have shaped the genomes of the hosts by driving a large number of adaptations at many genes, and these signals can be used to detect and further characterize ancient epidemics. Here, we detect the genomic footprints left by ancient viral epidemics that took place in the past approximately 50 000 years in the 26 human populations represented in the 1000 Genomes Project. By using the enrichment in signals of adaptation at approximately 4500 host loci that interact with specific types of viruses, we provide evidence that RNA viruses have driven a particularly large number of adaptive events across diverse human populations. These results suggest that different types of viruses may have exerted different selective pressures during human evolution. Knowledge of these past selective pressures will provide a deeper evolutionary perspective on current pathogenic threats. This article is part of the theme issue 'Insights into health and disease from ancient biomolecules'. 10.1098/rstb.2019.0575
    Evolutionary history of in the Pacific Islands. Blevins Kelly E,Crane Adele E,Lum Christopher,Furuta Kanako,Fox Keolu,Stone Anne C Philosophical transactions of the Royal Society of London. Series B, Biological sciences As one of the oldest known human diseases, leprosy or Hansen's disease remains a public health concern around the world with over 200 000 new cases in 2018. Most human leprosy cases are caused by , but a small number of cases are now known to be caused by , a sister taxon of . The global pattern of genomic variation in is not well defined. Particularly, in the Pacific Islands, the origins of leprosy are disputed. Historically, it has been argued that leprosy arrived on the islands during nineteenth century colonialism, but some oral traditions and palaeopathological evidence suggest an older introduction. To address this, as well as investigate patterns of pathogen exchange across the Pacific Islands, we extracted DNA from 39 formalin-fixed paraffin-embedded biopsy blocks dating to 1992-2016. Using whole-genome enrichment and next-generation sequencing, we produced nine genomes dating to 1998-2015 and ranging from 4-63× depth of coverage. Phylogenetic analyses indicate that these strains belong to basal lineages within the phylogeny, specifically falling in branches 0 and 5. The phylogeographical patterning and evolutionary dating analysis of these strains support a pre-modern introduction of into the Pacific Islands. This article is part of the theme issue 'Insights into health and disease from ancient biomolecules'. 10.1098/rstb.2019.0582
    Variola virus genome sequenced from an eighteenth-century museum specimen supports the recent origin of smallpox. Ferrari Giada,Neukamm Judith,Baalsrud Helle T,Breidenstein Abagail M,Ravinet Mark,Phillips Carina,Rühli Frank,Bouwman Abigail,Schuenemann Verena J Philosophical transactions of the Royal Society of London. Series B, Biological sciences Smallpox, caused by the variola virus (VARV), was a highly virulent disease with high mortality rates causing a major threat for global human health until its successful eradication in 1980. Despite previously published historic and modern VARV genomes, its past dissemination and diversity remain debated. To understand the evolutionary history of VARV with respect to historic and modern VARV genetic variation in Europe, we sequenced a VARV genome from a well-described eighteenth-century case from England (specimen P328). In our phylogenetic analysis, the new genome falls between the modern strains and another historic strain from Lithuania, supporting previous claims of larger diversity in early modern Europe compared to the twentieth century. Our analyses also resolve a previous controversy regarding the common ancestor between modern and historic strains by confirming a later date around the seventeenth century. Overall, our results point to the benefit of historic genomes for better resolution of past VARV diversity and highlight the value of such historic genomes from around the world to further understand the evolutionary history of smallpox as well as related diseases. This article is part of the theme issue 'Insights into health and disease from ancient biomolecules'. 10.1098/rstb.2019.0572
    New ancient Eastern European genomes illuminate the dispersal of plague in Europe. Morozova Irina,Kasianov Artem,Bruskin Sergey,Neukamm Judith,Molak Martyna,Batieva Elena,Pudło Aleksandra,Rühli Frank J,Schuenemann Verena J Philosophical transactions of the Royal Society of London. Series B, Biological sciences , the causative agent of plague, has been prevalent among humans for at least 5000 years, being accountable for several devastating epidemics in history, including the Black Death. Analyses of the genetic diversity of ancient strains of have shed light on the mechanisms of evolution and the spread of plague in Europe. However, many questions regarding the origins of the pathogen and its long persistence in Europe are still unresolved, especially during the late medieval time period. To address this, we present four newly assembled genomes from Eastern Europe (Poland and Southern Russia), dating from the fifteenth to eighteenth century AD. The analysis of polymorphisms in these genomes and their phylogenetic relationships with other ancient and modern strains may suggest several independent introductions of plague into Eastern Europe or its persistence in different reservoirs. Furthermore, with the reconstruction of a partial genome from rat skeletal remains found in a Polish ossuary, we were able to identify a potential animal reservoir in late medieval Europe. Overall, our results add new information concerning transmission and its evolutionary history in Eastern Europe. This article is part of the theme issue 'Insights into health and disease from ancient biomolecules'. 10.1098/rstb.2019.0569
    'TB or not TB': the conundrum of pre-European contact tuberculosis in the Pacific. McDonald S K,Matisoo-Smith E A,Buckley H R,Walter R K,Aung H L,Collins C J,Cook G M,Kardailsky O,Krause J,Knapp M Philosophical transactions of the Royal Society of London. Series B, Biological sciences Tuberculosis (TB) is a major global health threat, infecting one-third of the world's population. Despite this prominence, the age, origin and spread of the disease have been topics of contentious debate. Molecular studies suggest that , the most common strain of TB infecting humans today, originated in Africa and from there spread into Europe and Asia. The strains most commonly found across the Pacific and the Americas today are most closely related to European strains, supporting a hypothesis that the disease only reached these regions relatively recently via European sailors or settlers. However, this hypothesis is inconsistent with palaeopathological evidence of TB-like lesions in human remains from across the Pacific that predate European contact. Similarly, genetic evidence from pre-European South American mummies challenges the notion of a European introduction of the disease into the Pacific. Here, we review the complex evidence for the age and origin of TB in the Pacific, and discuss key gaps in our knowledge and how these may be addressed. This article is part of the theme issue 'Insights into health and disease from ancient biomolecules'. 10.1098/rstb.2019.0583
    Estimating molecular preservation of the intestinal microbiome via metagenomic analyses of latrine sediments from two medieval cities. Sabin Susanna,Yeh Hui-Yuan,Pluskowski Aleks,Clamer Christa,Mitchell Piers D,Bos Kirsten I Philosophical transactions of the Royal Society of London. Series B, Biological sciences Ancient latrine sediments, which contain the concentrated collective biological waste of past whole human communities, have the potential to be excellent proxies for human gastrointestinal health on the population level. A rich body of literature explores their use to detect the presence of gut-associated eukaryotic parasites through microscopy, immunoassays and genetics. Despite this interest, a lack of studies have explored the whole genetic content of ancient latrine sediments through consideration not only of gut-associated parasites, but also of core community gut microbiome signals that remain from the group that used the latrine. Here, we present a metagenomic analysis of bulk sediment from medieval latrines in Riga (Latvia) and Jerusalem. Our analyses reveal survival of microbial DNA representative of intestinal flora as well as numerous parasites. These data are compared against parasite taxon identifications obtained via microscopy and ELISA techniques. Together, these findings provide a first glimpse into the rich prokaryotic and eukaryotic intestinal flora of pre-industrial agricultural populations, which may give a better context for interpreting the health of modern microbiomes. This article is part of the theme issue 'Insights into health and disease from ancient biomolecules'. 10.1098/rstb.2019.0576
    The dental proteome of Homo antecessor. Welker Frido,Ramos-Madrigal Jazmín,Gutenbrunner Petra,Mackie Meaghan,Tiwary Shivani,Rakownikow Jersie-Christensen Rosa,Chiva Cristina,Dickinson Marc R,Kuhlwilm Martin,de Manuel Marc,Gelabert Pere,Martinón-Torres María,Margvelashvili Ann,Arsuaga Juan Luis,Carbonell Eudald,Marques-Bonet Tomas,Penkman Kirsty,Sabidó Eduard,Cox Jürgen,Olsen Jesper V,Lordkipanidze David,Racimo Fernando,Lalueza-Fox Carles,Bermúdez de Castro José María,Willerslev Eske,Cappellini Enrico Nature The phylogenetic relationships between hominins of the Early Pleistocene epoch in Eurasia, such as Homo antecessor, and hominins that appear later in the fossil record during the Middle Pleistocene epoch, such as Homo sapiens, are highly debated. For the oldest remains, the molecular study of these relationships is hindered by the degradation of ancient DNA. However, recent research has demonstrated that the analysis of ancient proteins can address this challenge. Here we present the dental enamel proteomes of H. antecessor from Atapuerca (Spain) and Homo erectus from Dmanisi (Georgia), two key fossil assemblages that have a central role in models of Pleistocene hominin morphology, dispersal and divergence. We provide evidence that H. antecessor is a close sister lineage to subsequent Middle and Late Pleistocene hominins, including modern humans, Neanderthals and Denisovans. This placement implies that the modern-like face of H. antecessor-that is, similar to that of modern humans-may have a considerably deep ancestry in the genus Homo, and that the cranial morphology of Neanderthals represents a derived form. By recovering AMELY-specific peptide sequences, we also conclude that the H. antecessor molar fragment from Atapuerca that we analysed belonged to a male individual. Finally, these H. antecessor and H. erectus fossils preserve evidence of enamel proteome phosphorylation and proteolytic digestion that occurred in vivo during tooth formation. Our results provide important insights into the evolutionary relationships between H. antecessor and other hominin groups, and pave the way for future studies using enamel proteomes to investigate hominin biology across the existence of the genus Homo. 10.1038/s41586-020-2153-8
    Bronze Age population dynamics and the rise of dairy pastoralism on the eastern Eurasian steppe. Jeong Choongwon,Wilkin Shevan,Amgalantugs Tsend,Bouwman Abigail S,Taylor William Timothy Treal,Hagan Richard W,Bromage Sabri,Tsolmon Soninkhishig,Trachsel Christian,Grossmann Jonas,Littleton Judith,Makarewicz Cheryl A,Krigbaum John,Burri Marta,Scott Ashley,Davaasambuu Ganmaa,Wright Joshua,Irmer Franziska,Myagmar Erdene,Boivin Nicole,Robbeets Martine,Rühli Frank J,Krause Johannes,Frohlich Bruno,Hendy Jessica,Warinner Christina Proceedings of the National Academy of Sciences of the United States of America Recent paleogenomic studies have shown that migrations of Western steppe herders (WSH) beginning in the Eneolithic (ca. 3300-2700 BCE) profoundly transformed the genes and cultures of Europe and central Asia. Compared with Europe, however, the eastern extent of this WSH expansion is not well defined. Here we present genomic and proteomic data from 22 directly dated Late Bronze Age burials putatively associated with early pastoralism in northern Mongolia (ca. 1380-975 BCE). Genome-wide analysis reveals that they are largely descended from a population represented by Early Bronze Age hunter-gatherers in the Baikal region, with only a limited contribution (∼7%) of WSH ancestry. At the same time, however, mass spectrometry analysis of dental calculus provides direct protein evidence of bovine, sheep, and goat milk consumption in seven of nine individuals. No individuals showed molecular evidence of lactase persistence, and only one individual exhibited evidence of >10% WSH ancestry, despite the presence of WSH populations in the nearby Altai-Sayan region for more than a millennium. Unlike the spread of Neolithic farming in Europe and the expansion of Bronze Age pastoralism on the Western steppe, our results indicate that ruminant dairy pastoralism was adopted on the Eastern steppe by local hunter-gatherers through a process of cultural transmission and minimal genetic exchange with outside groups. 10.1073/pnas.1813608115
    Middle Holocene plant cultivation on the Atlantic Forest coast of Brazil? Pezo-Lanfranco Luis,Eggers Sabine,Petronilho Cecilia,Toso Alice,da Rocha Bandeira Dione,Von Tersch Matthew,Dos Santos Adriana M P,Ramos da Costa Beatriz,Meyer Roberta,Colonese André Carlo Royal Society open science This work provides robust oral pathology and stable isotope evidence on Bayesian mixing model for an unexpectedly high consumption of carbohydrates by a Middle Holocene coastal population of the Atlantic Forest of South America, an area traditionally viewed as peripheral to early centres of food production on the continent. A diversified economy with substantial consumption of plant resources was in place at the shellmound (or ) of Morro do Ouro, in Babitonga Bay, and supported a dense population at 4500 cal BP. This dietary composition is unique when compared with that of other contemporary and later groups in the region, including peoples who used ceramics and domesticated crops. The results corroborate independent dietary evidence, such as stone tool artefacts for plant processing and plant microremains in dental calculus of the same individuals, and suggest plant cultivation possibly took place in this region at the same time as the development of early agriculture in Amazonia and the La Plata Basin. Our study situates the Atlantic Forest coast of Brazil on the map of early plant management in the Neotropics. 10.1098/rsos.180432
    The past, present and future of ancient bacterial DNA. Arning Nicolas,Wilson Daniel J Microbial genomics Groundbreaking studies conducted in the mid-1980s demonstrated the possibility of sequencing ancient DNA (aDNA), which has allowed us to answer fundamental questions about the human past. Microbiologists were thus given a powerful tool to glimpse directly into inscrutable bacterial history, hitherto inaccessible due to a poor fossil record. Initially plagued by concerns regarding contamination, the field has grown alongside technical progress, with the advent of high-throughput sequencing being a breakthrough in sequence output and authentication. Albeit burdened with challenges unique to the analysis of bacteria, a growing number of viable sources for aDNA has opened multiple avenues of microbial research. Ancient pathogens have been extracted from bones, dental pulp, mummies and historical medical specimens and have answered focal historical questions such as identifying the aetiological agent of the black death as . Furthermore, ancient human microbiomes from fossilized faeces, mummies and dental plaque have shown shifts in human commensals through the Neolithic demographic transition and industrial revolution, whereas environmental isolates stemming from permafrost samples have revealed signs of ancient antimicrobial resistance. Culminating in an ever-growing repertoire of ancient genomes, the quickly expanding body of bacterial aDNA studies has also enabled comparisons of ancient genomes to their extant counterparts, illuminating the evolutionary history of bacteria. In this review we summarize the present avenues of research and contextualize them in the past of the field whilst also pointing towards questions still to be answered. 10.1099/mgen.0.000384
    Identification of ancient viruses from metagenomic data of the Jomon people. Nishimura Luca,Sugimoto Ryota,Inoue Jun,Nakaoka Hirofumi,Kanzawa-Kiriyama Hideaki,Shinoda Ken-Ichi,Inoue Ituro Journal of human genetics Ancient DNA studies provide genomic information about the origins, population structures, and physical characteristics of ancient humans that cannot be solely examined by archeological studies. The DNAs extracted from ancient human bones, teeth, or tissues are often contaminated with coexisting bacterial and viral genomes that contain DNA from ancient microbes infecting those of ancient humans. Information on ancient viral genomes is useful in making inferences about the viral evolution. Here, we have utilized metagenomic sequencing data from the dental pulp of five Jomon individuals, who lived on the Japanese archipelago more than 3000 years ago; this is to detect ancient viral genomes. We conducted de novo assembly of the non-human reads where we have obtained 277,387 contigs that were longer than 1000 bp. These contigs were subjected to homology searches against a collection of modern viral genome sequences. We were able to detect eleven putative ancient viral genomes. Among them, we reconstructed the complete sequence of the Siphovirus contig89 (CT89) viral genome. The Jomon CT89-like sequence was determined to contain 59 open reading frames, among which five genes known to encode phage proteins were under strong purifying selection. The host of CT89 was predicted to be Schaalia meyeri, a bacterium residing in the human oral cavity. Finally, the CT89 phylogenetic tree showed two clusters, from both of which the Jomon sequence was separated. Our results suggest that metagenomic information from the dental pulp of the Jomon people is essential in retrieving ancient viral genomes used to examine their evolution. 10.1038/s10038-020-00841-6
    Selection of Appropriate Metagenome Taxonomic Classifiers for Ancient Microbiome Research. Velsko Irina M,Frantz Laurent A F,Herbig Alexander,Larson Greger,Warinner Christina mSystems Metagenomics enables the study of complex microbial communities from myriad sources, including the remains of oral and gut microbiota preserved in archaeological dental calculus and paleofeces, respectively. While accurate taxonomic assignment is essential to this process, DNA damage characteristic of ancient samples (e.g., reduction in fragment size and cytosine deamination) may reduce the accuracy of read taxonomic assignment. Using a set of -generated metagenomic data sets, we investigated how the addition of ancient DNA (aDNA) damage patterns influences microbial taxonomic assignment by five widely used profilers: QIIME/UCLUST, MetaPhlAn2, MIDAS, CLARK-S, and MALT. -generated data sets were designed to mimic dental plaque, consisting of 40, 100, and 200 microbial species/strains, both with and without simulated aDNA damage patterns. Following taxonomic assignment, the profiles were evaluated for species presence/absence, relative abundance, alpha diversity, beta diversity, and specific taxonomic assignment biases. Unifrac metrics indicated that both MIDAS and MetaPhlAn2 reconstructed the most accurate community structure. QIIME/UCLUST, CLARK-S, and MALT had the highest number of inaccurate taxonomic assignments; false-positive rates were highest by CLARK-S and QIIME/UCLUST. Filtering out species present at <0.1% abundance greatly increased the accuracy of CLARK-S and MALT. All programs except CLARK-S failed to detect some species from the input file that were in their databases. The addition of ancient DNA damage resulted in minimal differences in species detection and relative abundance between simulated ancient and modern data sets for most programs. Overall, taxonomic profiling biases are program specific rather than damage dependent, and the choice of taxonomic classification program should be tailored to specific research questions. Ancient biomolecules from oral and gut microbiome samples have been shown to be preserved in the archaeological record. Studying ancient microbiome communities using metagenomic techniques offers a unique opportunity to reconstruct the evolutionary trajectories of microbial communities through time. DNA accumulates specific damage over time, which could potentially affect taxonomic classification and our ability to accurately reconstruct community assemblages. It is therefore necessary to assess whether ancient DNA (aDNA) damage patterns affect metagenomic taxonomic profiling. Here, we assessed biases in community structure, diversity, species detection, and relative abundance estimates by five popular metagenomic taxonomic classification programs using -generated data sets with and without aDNA damage. Damage patterns had minimal impact on the taxonomic profiles produced by each program, while false-positive rates and biases were intrinsic to each program. Therefore, the most appropriate classification program is one that minimizes the biases related to the questions being addressed. 10.1128/mSystems.00080-18
    Ancient dental pulp: Masterpiece tissue for paleomicrobiology. Mai Ba Hoang Anh,Drancourt Michel,Aboudharam Gérard Molecular genetics & genomic medicine INTRODUCTION:Dental pulp with special structure has become a good reference sample in paleomicrobiology-related blood-borne diseases, many pathogens were detected by different methods based on the diagnosis of nucleic acids and proteins. OBJECTIVES:This review aims to propose the preparation process from ancient teeth collection to organic molecule extraction of dental pulp and summary, analyze the methods that have been applied to detect septicemic pathogens through ancient dental pulps during the past 20 years following the first detection of an ancient microbe. METHODS:The papers used in this review with two main objectives were obtained from PubMed and Google scholar with combining keywords: "ancient," "dental pulp," "teeth," "anatomy," "structure," "collection," "preservation," "selection," "photography," "radiography," "contamination," "decontamination," "DNA," "protein," "extraction," "bone," "paleomicrobiology," "bacteria," "virus," "pathogen," "molecular biology," "proteomics," "PCR," "MALDI-TOF," "LC/MS," "ELISA," "immunology," "immunochromatography," "genome," "microbiome," "metagenomics." RESULTS:The analysis of ancient dental pulp should have a careful preparation process with many different steps to give highly accurate results, each step complies with the rules in archaeology and paleomicrobiology. After the collection of organic molecules from dental pulp, they were investigated for pathogen identification based on the analysis of DNA and protein. Actually, DNA approach takes a principal role in diagnosis while the protein approach is more and more used. A total of seven techniques was used and ten bacteria (Yersinia pestis, Bartonella quintana, Salmonella enterica serovar Typhi, Salmonella enterica serovar Paratyphi C, Mycobacterium leprae, Mycobacterium tuberculosis, Rickettsia prowazeki, Staphylococcus aureus, Borrelia recurrentis, Bartonella henselae) and one virus (Anelloviridae) were identified. Y. pestis had the most published in quantity and all methods were investigated for this pathogen, S. aureus and B. recurrentis were identified by three different methods and others only by one. The combining methods interestingly increase the positive rate with ELISA, PCR and iPCR in Yersinia pestis diagnosis. Twenty-seven ancient genomes of Y. pestis and one ancient genome of B. recurrentis were reconstructed. Comparing to the ancient bone, ancient teeth showed more advantage in septicemic diagnosis. Beside pathogen identification, ancient pulp help to distinguish species. CONCLUSIONS:Dental pulp with specific tissue is a suitable sample for detection of the blood infection in the past through DNA and protein identification with the correct preparation process, furthermore, it helps to more understand the pathogens of historic diseases and epidemics. 10.1002/mgg3.1202
    Oral health status in historic population: Macroscopic and metagenomic evidence. Willmann Claire,Mata Xavier,Hanghoej Kristian,Tonasso Laure,Tisseyre Lenka,Jeziorski Céline,Cabot Elodie,Chevet Pierre,Crubézy Eric,Orlando Ludovic,Esclassan Rémi,Thèves Catherine PloS one Recent developments in High-Throughput DNA sequencing (HTS) technologies and ancient DNA (aDNA) research have opened access to the characterization of the microbial communities within past populations. Most studies have, however, relied on the analysis of dental calculus as one particular material type particularly prone to the molecular preservation of ancient microbial biofilms and potential of entire teeth for microbial characterization, both of healthy communities and pathogens in ancient individuals, remains overlooked. In this study, we used shotgun sequencing to characterize the bacterial composition from historical subjects showing macroscopic evidence of oral pathologies. We first carried out a macroscopic analysis aimed at identifying carious or periodontal diseases in subjects belonging to a French rural population of the 18th century AD. We next examined radiographically six subjects showing specific, characteristic dental pathologies and applied HTS shotgun sequencing to characterize the microbial communities present in and on the dental material. The presence of Streptococcus mutans and also Rothia dentocariosa, Actinomyces viscosus, Porphyromonas gingivalis, Tannerella forsythia, Pseudoramibacter alactolyticus, Olsenella uli and Parvimonas micra was confirmed through the presence of typical signatures of post-mortem DNA damage at an average depth-of-coverage ranging from 0.5 to 7X, with a minimum of 35% (from 35 to 93%) of the positions in the genome covered at least once. Each sampled tooth showed a specific bacterial signature associated with carious or periodontal pathologies. This work demonstrates that from a healthy independent tooth, without visible macroscopic pathology, we can identify a signature of specific pathogens and deduce the oral health status of an individual. 10.1371/journal.pone.0196482
    Exploring oral paleopathology in the Central Andes: A review. Gagnon Celeste Marie International journal of paleopathology This targeted review of oral paleopathology in the Central Andes explores research that focuses on a set of interrelated, multifactorial processes: dental caries, macrowear, alveolar abscess, antemortem tooth loss (AMTL), periodontal disease, and the presence of dental calculus. These conditions help characterize oral health because they result from the culturally mediated interaction of individuals' oral cavity with their external environment. To better understand how osteologists working in the Central Andes have interpreted the frequencies of these conditions, I review the etiology of each, as well as discuss the important issues in their analyses. I then highlight studies that integrate of a number of oral paleopathological conditions, that examine associations between oral conditions and other skeletal indicators of health, or that use multivariate statistical techniques to analyze conditions. In the Central Andes, these proxies for oral health have generally focused on several key research themes including the introduction of domesticated foods may have occurred earlier than expected, but that populations may have maintained mixed subsistence strategies for a significant period. Researchers have also identified that changes accompanying Inca imperialism were likely not as detrimental to local populations as was Spanish colonialism. Finally, the long-practiced, culturally important, activity of chewing coca has been shown to create an identifiable pattern of oral paleopathological conditions. 10.1016/j.ijpp.2019.10.003
    Assessing alignment-based taxonomic classification of ancient microbial DNA. Eisenhofer Raphael,Weyrich Laura Susan PeerJ The field of palaeomicrobiology-the study of ancient microorganisms-is rapidly growing due to recent methodological and technological advancements. It is now possible to obtain vast quantities of DNA data from ancient specimens in a high-throughput manner and use this information to investigate the dynamics and evolution of past microbial communities. However, we still know very little about how the characteristics of ancient DNA influence our ability to accurately assign microbial taxonomies (i.e. identify species) within ancient metagenomic samples. Here, we use both simulated and published metagenomic data sets to investigate how ancient DNA characteristics affect alignment-based taxonomic classification. We find that nucleotide-to-nucleotide, rather than nucleotide-to-protein, alignments are preferable when assigning taxonomies to short DNA fragment lengths routinely identified within ancient specimens (<60 bp). We determine that deamination (a form of ancient DNA damage) and random sequence substitutions corresponding to ∼100,000 years of genomic divergence minimally impact alignment-based classification. We also test four different reference databases and find that database choice can significantly bias the results of alignment-based taxonomic classification in ancient metagenomic studies. Finally, we perform a reanalysis of previously published ancient dental calculus data, increasing the number of microbial DNA sequences assigned taxonomically by an average of 64.2-fold and identifying microbial species previously unidentified in the original study. Overall, this study enhances our understanding of how ancient DNA characteristics influence alignment-based taxonomic classification of ancient microorganisms and provides recommendations for future palaeomicrobiological studies. 10.7717/peerj.6594
    Isotopic and microbotanical insights into Iron Age agricultural reliance in the Central African rainforest. Bleasdale Madeleine,Wotzka Hans-Peter,Eichhorn Barbara,Mercader Julio,Styring Amy,Zech Jana,Soto María,Inwood Jamie,Clarke Siobhán,Marzo Sara,Fiedler Bianca,Linseele Veerle,Boivin Nicole,Roberts Patrick Communications biology The emergence of agriculture in Central Africa has previously been associated with the migration of Bantu-speaking populations during an anthropogenic or climate-driven 'opening' of the rainforest. However, such models are based on assumptions of environmental requirements of key crops (e.g. Pennisetum glaucum) and direct insights into human dietary reliance remain absent. Here, we utilise stable isotope analysis (δC, δN, δO) of human and animal remains and charred food remains, as well as plant microparticles from dental calculus, to assess the importance of incoming crops in the Congo Basin. Our data, spanning the early Iron Age to recent history, reveals variation in the adoption of cereals, with a persistent focus on forest and freshwater resources in some areas. These data provide new dietary evidence and document the longevity of mosaic subsistence strategies in the region. 10.1038/s42003-020-01324-2
    Multidisciplinary investigations of the diets of two post-medieval populations from London using stable isotopes and microdebris analysis. Bleasdale Madeleine,Ponce Paola,Radini Anita,Wilson Andrew S,Doherty Sean,Daley Patrick,Brown Chloe,Spindler Luke,Sibun Lucy,Speller Camilla,Alexander Michelle M Archaeological and anthropological sciences This paper presents the first multi-tissue study of diet in post-medieval London using both the stable light isotope analysis of carbon and nitrogen and analysis of microdebris in dental calculus. Dietary intake was explored over short and long timescales. Bulk bone collagen was analysed from humans from the Queen's Chapel of the Savoy (QCS) ( = 66) and the St Barnabas/St Mary Abbots (SB) ( = 25). Incremental dentine analysis was performed on the second molar of individual QCS1123 to explore childhood dietary intake. Bulk hair samples ( = 4) were sampled from adults from QCS, and dental calculus was analysed from four other individuals using microscopy. In addition, bone collagen from a total of 46 animals from QCS ( = 11) and the additional site of Prescot Street ( = 35) was analysed, providing the first animal dietary baseline for post-medieval London. Overall, isotopic results suggest a largely C-based terrestrial diet for both populations, with the exception of QCS1123 who exhibited values consistent with the consumption of C food sources throughout childhood and adulthood. The differences exhibited in δN across both populations likely reflect variations in diet due to social class and occupation, with individuals from SB likely representing wealthier individuals consuming larger quantities of animal and marine fish protein. Microdebris analysis results were limited but indicate the consumption of domestic cereals. This paper demonstrates the utility of a multidisciplinary approach to investigate diet across long and short timescales to further our understanding of variations in social status and mobility. 10.1007/s12520-019-00910-8
    Dairy pastoralism sustained eastern Eurasian steppe populations for 5,000 years. Wilkin Shevan,Ventresca Miller Alicia,Taylor William T T,Miller Bryan K,Hagan Richard W,Bleasdale Madeleine,Scott Ashley,Gankhuyg Sumiya,Ramsøe Abigail,Uliziibayar S,Trachsel Christian,Nanni Paolo,Grossmann Jonas,Orlando Ludovic,Horton Mark,Stockhammer Philipp W,Myagmar Erdene,Boivin Nicole,Warinner Christina,Hendy Jessica Nature ecology & evolution Dairy pastoralism is integral to contemporary and past lifeways on the eastern Eurasian steppe, facilitating survival in agriculturally challenging environments. While previous research has indicated that ruminant dairy pastoralism was practiced in the region by circa 1300 BC, the origin, extent and diversity of this custom remain poorly understood. Here, we analyse ancient proteins from human dental calculus recovered from geographically diverse locations across Mongolia and spanning 5,000 years. We present the earliest evidence for dairy consumption on the eastern Eurasian steppe by circa 3000 BC and the later emergence of horse milking at circa 1200 BC, concurrent with the first evidence for horse riding. We argue that ruminant dairying contributed to the demographic success of Bronze Age Mongolian populations and that the origins of traditional horse dairy products in eastern Eurasia are closely tied to the regional emergence of mounted herding societies during the late second millennium BC. 10.1038/s41559-020-1120-y
    Ancient DNA Reconstructs the Genetic Legacies of Precontact Puerto Rico Communities. Nieves-Colón Maria A,Pestle William J,Reynolds Austin W,Llamas Bastien,de la Fuente Constanza,Fowler Kathleen,Skerry Katherine M,Crespo-Torres Edwin,Bustamante Carlos D,Stone Anne C Molecular biology and evolution Indigenous peoples have occupied the island of Puerto Rico since at least 3000 BC. Due to the demographic shifts that occurred after European contact, the origin(s) of these ancient populations, and their genetic relationship to present-day islanders, are unclear. We use ancient DNA to characterize the population history and genetic legacies of precontact Indigenous communities from Puerto Rico. Bone, tooth, and dental calculus samples were collected from 124 individuals from three precontact archaeological sites: Tibes, Punta Candelero, and Paso del Indio. Despite poor DNA preservation, we used target enrichment and high-throughput sequencing to obtain complete mitochondrial genomes (mtDNA) from 45 individuals and autosomal genotypes from two individuals. We found a high proportion of Native American mtDNA haplogroups A2 and C1 in the precontact Puerto Rico sample (40% and 44%, respectively). This distribution, as well as the haplotypes represented, supports a primarily Amazonian South American origin for these populations and mirrors the Native American mtDNA diversity patterns found in present-day islanders. Three mtDNA haplotypes from precontact Puerto Rico persist among Puerto Ricans and other Caribbean islanders, indicating that present-day populations are reservoirs of precontact mtDNA diversity. Lastly, we find similarity in autosomal ancestry patterns between precontact individuals from Puerto Rico and the Bahamas, suggesting a shared component of Indigenous Caribbean ancestry with close affinity to South American populations. Our findings contribute to a more complete reconstruction of precontact Caribbean population history and explore the role of Indigenous peoples in shaping the biocultural diversity of present-day Puerto Ricans and other Caribbean islanders. 10.1093/molbev/msz267
    2000-year-old pathogen genomes reconstructed from metagenomic analysis of Egyptian mummified individuals. Neukamm Judith,Pfrengle Saskia,Molak Martyna,Seitz Alexander,Francken Michael,Eppenberger Partick,Avanzi Charlotte,Reiter Ella,Urban Christian,Welte Beatrix,Stockhammer Philipp W,Teßmann Barbara,Herbig Alexander,Harvati Katerina,Nieselt Kay,Krause Johannes,Schuenemann Verena J BMC biology BACKGROUND:Recent advances in sequencing have facilitated large-scale analyses of the metagenomic composition of different samples, including the environmental microbiome of air, water, and soil, as well as the microbiome of living humans and other animals. Analyses of the microbiome of ancient human samples may provide insights into human health and disease, as well as pathogen evolution, but the field is still in its very early stages and considered highly challenging. RESULTS:The metagenomic and pathogen content of Egyptian mummified individuals from different time periods was investigated via genetic analysis of the microbial composition of various tissues. The analysis of the dental calculus' microbiome identified Red Complex bacteria, which are correlated with periodontal diseases. From bone and soft tissue, genomes of two ancient pathogens, a 2200-year-old Mycobacterium leprae strain and a 2000-year-old human hepatitis B virus, were successfully reconstructed. CONCLUSIONS:The results show the reliability of metagenomic studies on Egyptian mummified individuals and the potential to use them as a source for the extraction of ancient pathogen DNA. 10.1186/s12915-020-00839-8
    Paleogenomic insights into the red complex bacteria in Pre-Hispanic and Colonial individuals from Mexico. Bravo-Lopez Miriam,Villa-Islas Viridiana,Rocha Arriaga Carolina,Villaseñor-Altamirano Ana B,Guzmán-Solís Axel,Sandoval-Velasco Marcela,Wesp Julie K,Alcantara Keitlyn,López-Corral Aurelio,Gómez-Valdés Jorge,Mejía Elizabeth,Herrera Alberto,Meraz-Moreno Alejandro,Moreno-Cabrera Maria de la Luz,Moreno-Estrada Andrés,Nieves-Colón Maria A,Olvera Joel,Pérez-Pérez Julia,Iversen Katrine Højholt,Rasmussen Simon,Sandoval Karla,Zepeda Gabriela,Ávila-Arcos María C Philosophical transactions of the Royal Society of London. Series B, Biological sciences The 'red complex' is an aggregate of three oral bacteria (, and ) responsible for severe clinical manifestation of periodontal disease. Here, we report the first direct evidence of ancient DNA in dentin and dental calculus samples from archaeological skeletal remains that span from the Pre-Hispanic to the Colonial period in Mexico. We recovered twelve partial ancient genomes and observed a distinct phylogenetic placement of samples, suggesting that the strains present in Pre-Hispanic individuals likely arrived with the first human migrations to the Americas and that new strains were introduced with the arrival of European and African populations in the sixteenth century. We also identified instances of the differential presence of genes between periods in the ancient genomes, with certain genes present in Pre-Hispanic individuals and absent in Colonial individuals, and . This study highlights the potential for studying ancient genomes to unveil past social interactions through analysis of disease transmission. Our results illustrate the long-standing relationship between this oral pathogen and its human host, while also unveiling key evidence to understand its evolutionary history in Pre-Hispanic and Colonial Mexico. This article is part of the theme issue 'Insights into health and disease from ancient biomolecules'. 10.1098/rstb.2019.0580
    Interbreeding between Neanderthals and modern humans: Remarks and methodological dangers of a dental calculus microbiome analysis. Charlier Philippe,Gaultier Frédérick,Héry-Arnaud Geneviève Journal of human evolution 10.1016/j.jhevol.2018.06.007
    Living off the land: Terrestrial-based diet and dairying in the farming communities of the Neolithic Balkans. Stojanovski Darko,Živaljević Ivana,Dimitrijević Vesna,Dunne Julie,Evershed Richard P,Balasse Marie,Dowle Adam,Hendy Jessica,McGrath Krista,Fischer Roman,Speller Camilla,Jovanović Jelena,Casanova Emmanuelle,Knowles Timothy,Balj Lidija,Naumov Goce,Putica Anđelka,Starović Andrej,Stefanović Sofija PloS one The application of biomolecular techniques to archaeological materials from the Balkans is providing valuable new information on the prehistory of the region. This is especially relevant for the study of the neolithisation process in SE Europe, which gradually affected the rest of the continent. Here, to answer questions regarding diet and subsistence practices in early farming societies in the central Balkans, we combine organic residue analyses of archaeological pottery, taxonomic and isotopic study of domestic animal remains and biomolecular analyses of human dental calculus. The results from the analyses of the lipid residues from pottery suggest that milk was processed in ceramic vessels. Dairy products were shown to be part of the subsistence strategies of the earliest Neolithic communities in the region but were of varying importance in different areas of the Balkan. Conversely, milk proteins were not detected within the dental calculus. The molecular and isotopic identification of meat, dairy, plants and beeswax in the pottery lipids also provided insights into the diversity of diet in these early Neolithic communities, mainly based on terrestrial resources. We also present the first compound-specific radiocarbon dates for the region, obtained directly from absorbed organic residues extracted from pottery, identified as dairy lipids. 10.1371/journal.pone.0237608
    Exploring late Paleolithic and Mesolithic diet in the Eastern Alpine region of Italy through multiple proxies. Oxilia Gregorio,Bortolini Eugenio,Badino Federica,Bernardini Federico,Gazzoni Valentina,Lugli Federico,Romandini Matteo,Radini Anita,Terlato Gabriele,Marciani Giulia,Silvestrini Sara,Menghi Sartorio Jessica C,Thun Hohenstein Ursula,Fiorenza Luca,Kullmer Ottmar,Tuniz Claudio,Moggi Cecchi Jacopo,Talamo Sahra,Fontana Federica,Peresani Marco,Benazzi Stefano,Cristiani Emanuela American journal of physical anthropology OBJECTIVES:The analysis of prehistoric human dietary habits is key for understanding the effects of paleoenvironmental changes on the evolution of cultural and social human behaviors. In this study, we compare results from zooarchaeological, stable isotope and dental calculus analyses as well as lower second molar macrowear patterns to gain a broader understanding of the diet of three individuals who lived between the end of the Late Pleistocene and the Early Holocene (ca., 17-8 ky cal BP) in the Eastern Alpine region of Italy. MATERIALS AND METHODS:We analyze individuals buried at the sites of Riparo Tagliente (Verona), Riparo Villabruna, and Mondeval de Sora (Belluno). The three burials provide a unique dataset for diachronically exploring the influence of climatic changes on human subsistence strategies. RESULTS:Isotopic results indicate that all individuals likely relied on both terrestrial and freshwater animal proteins. Even though dental calculus analysis was, in part, hindered by the amount of mineral deposit available on the teeth, tooth macrowear study suggests that the dietary habits of the individuals included plant foods. Moreover, differences in macrowear patterns of lower second molars have been documented between Neanderthals and modern humans in the present sample, due to a prevalence of Buccal wear among the former as opposed to higher values of Lingual wear in modern human teeth. DISCUSSION:Isotopic analyses have emphasized the contribution of animal proteins in the diet of the three foragers from the Eastern Alpine region. The possible intake of carbohydrate-rich plant foods, suggested by the retrieval of plant remains in dental calculus, is supported by the signal of macrowear analysis. Moreover, the latter method indicates that the distribution of macrowear in lower second molars (M s) allows us to discriminate between Neanderthals and modern humans within the present reference sample. Overall, our results show these three prehistoric hunter-gatherers were well adapted to the environment in which they lived exploiting many natural resources. 10.1002/ajpa.24128
    New insights on Neolithic food and mobility patterns in Mediterranean coastal populations. Goude Gwenaëlle,Salazar-García Domingo C,Power Robert C,Rivollat Maïté,Gourichon Lionel,Deguilloux Marie-France,Pemonge Marie-Hélène,Bouby Laurent,Binder Didier American journal of physical anthropology 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 Sr/ 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. 10.1002/ajpa.24089
    Metagenomics of the modern and historical human oral microbiome with phylogenetic studies on and . Achtman Mark,Zhou Zhemin Philosophical transactions of the Royal Society of London. Series B, Biological sciences We have recently developed bioinformatic tools to accurately assign metagenomic sequence reads to microbial taxa: SPARSE for probabilistic, taxonomic classification of sequence reads; EToKi for assembling and polishing genomes from short-read sequences; and GrapeTree, a graphic visualizer of genetic distances between large numbers of genomes. Together, these methods support comparative analyses of genomes from ancient skeletons and modern humans. Here, we illustrate these capabilities with 784 samples from historical dental calculus, modern saliva and modern dental plaque. The analyses revealed 1591 microbial species within the oral microbiome. We anticipated that the oral complexes of Socransky ., which were defined in 1998, would predominate among taxa whose frequencies differed by source. However, although some species discriminated between sources, we could not confirm the existence of the complexes. The results also illustrate further functionality of our pipelines with two species that are associated with dental caries, and . They were rare in historical dental calculus but common in modern plaque, and even more common in saliva. Reconstructed draft genomes of these two species from metagenomic samples in which they were abundant were combined with modern public genomes to provide a detailed overview of their core genomic diversity. This article is part of the theme issue 'Insights into health and disease from ancient biomolecules'. 10.1098/rstb.2019.0573
    A multidisciplinary approach for investigating dietary and medicinal habits of the Medieval population of Santa Severa (7th-15th centuries, Rome, Italy). Gismondi Angelo,Baldoni Marica,Gnes Micaela,Scorrano Gabriele,D'Agostino Alessia,Di Marco Gabriele,Calabria Giulietta,Petrucci Michela,Müldner Gundula,Von Tersch Matthew,Nardi Alessandra,Enei Flavio,Canini Antonella,Rickards Olga,Alexander Michelle,Martínez-Labarga Cristina PloS one A multidisciplinary approach, combining stable isotope analysis from bone proteins and investigations on dental calculus using DNA analysis, light microscopy, and gas chromatography coupled with mass spectrometry, was applied to reconstruct dietary and medicinal habits of the individuals recovered in the cemetery of the Castle of Santa Severa (7th-15th centuries CE; Rome, Italy). Stable isotope analysis was performed on 120 humans, 41 faunal specimens and 8 charred seeds. Dental calculus analyses were carried out on 94 samples. Overall, isotope data indicated an omnivorous diet based on C3-terrestrial protein, although some individuals possessed carbon values indicative of C4 plant consumption. In terms of animal protein, the diet was probably based on cattle, sheep, pig and chicken products, as witnessed by the archaeozoological findings. Evidence from calculus suggested the consumption of C3 cereals, Fabaceae, Fagaceae, milk and dairy products. Secondary metabolites of herbs and wine were also detected. The detection of marine fish ancient DNA, as well as of ω3 fatty acids in calculus, hypothesized the consumption of marine foodstuffs for this coastal population, despite the lack of a clear marine isotopic signal and the presence of polyunsaturated fatty acids in plant tissues. Moreover, the knowledge of ethnopharmacological tradition and the application of medicinal plants (e.g. Punica granatum L., Ephedra sp. L.) were also identified. The detection of artemisinin, known to have antimalarial properties, led to hypothesize the presence of malaria in the area. Altogether, the combined application of microscopy and biomolecular techniques provided an innovative reconstruction of Medieval lifeways in Central Italy. 10.1371/journal.pone.0227433
    Oral microbiome diversity in chimpanzees from Gombe National Park. Ozga Andrew T,Gilby Ian,Nockerts Rebecca S,Wilson Michael L,Pusey Anne,Stone Anne C Scientific reports Historic calcified dental plaque (dental calculus) can provide a unique perspective into the health status of past human populations but currently no studies have focused on the oral microbial ecosystem of other primates, including our closest relatives, within the hominids. Here we use ancient DNA extraction methods, shotgun library preparation, and next generation Illumina sequencing to examine oral microbiota from 19 dental calculus samples recovered from wild chimpanzees (Pan troglodytes schweinfurthii) who died in Gombe National Park, Tanzania. The resulting sequences were trimmed for quality, analyzed using MALT, MEGAN, and alignment scripts, and integrated with previously published dental calculus microbiome data. We report significant differences in oral microbiome phyla between chimpanzees and anatomically modern humans (AMH), with chimpanzees possessing a greater abundance of Bacteroidetes and Fusobacteria, and AMH showing higher Firmicutes and Proteobacteria. Our results suggest that by using an enterotype clustering method, results cluster largely based on host species. These clusters are driven by Porphyromonas and Fusobacterium genera in chimpanzees and Haemophilus and Streptococcus in AMH. Additionally, we compare a nearly complete Porphyromonas gingivalis genome to previously published genomes recovered from human gingiva to gain perspective on evolutionary relationships across host species. Finally, using shotgun sequence data we assessed indicators of diet from DNA in calculus and suggest exercising caution when making assertions related to host lifestyle. These results showcase core differences between host species and stress the importance of continued sequencing of nonhuman primate microbiomes in order to fully understand the complexity of their oral ecologies. 10.1038/s41598-019-53802-1
    Investigating the demographic history of Japan using ancient oral microbiota. Eisenhofer Raphael,Kanzawa-Kiriyama Hideaki,Shinoda Ken-Ichi,Weyrich Laura S Philosophical transactions of the Royal Society of London. Series B, Biological sciences While microbial communities in the human body (microbiota) are now commonly associated with health and disease in industrialised populations, we know very little about how these communities co-evolved and changed with humans throughout history and deep prehistory. We can now examine these communities by sequencing ancient DNA preserved within calcified dental plaque (calculus), providing insights into the origins of disease and their links to human history. Here, we examine ancient DNA preserved within dental calculus samples and their associations with two major cultural periods in Japan: the Jomon period hunter-gatherers approximately 3000 years before present (BP) and the Edo period agriculturalists 400-150 BP. We investigate how human oral microbiomes have changed in Japan through time and explore the presence of microorganisms associated with oral diseases (e.g. periodontal disease, dental caries) in ancient Japanese populations. Finally, we explore oral microbial strain diversity and its potential links to ancient demography in ancient Japan by performing phylogenomic analysis of a widely conserved oral species- oral taxon 439. This research represents, to our knowledge, the first study of ancient oral microbiomes from Japan and demonstrates that the analysis of ancient dental calculus can provide key information about the origin of non-infectious disease and its deep roots with human demography. This article is part of the theme issue 'Insights into health and disease from ancient biomolecules'. 10.1098/rstb.2019.0578
    Dental calculus indicates widespread plant use within the stable Neanderthal dietary niche. Power Robert C,Salazar-García Domingo C,Rubini Mauro,Darlas Andrea,Harvati Katerina,Walker Michael,Hublin Jean-Jacques,Henry Amanda G Journal of human evolution 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. 10.1016/j.jhevol.2018.02.009
    A probable prehistoric case of meningococcal disease from San Francisco Bay: Next generation sequencing of Neisseria meningitidis from dental calculus and osteological evidence. Eerkens Jelmer W,Nichols Ruth V,Murray Gemma G R,Perez Katherine,Murga Engel,Kaijankoski Phil,Rosenthal Jeffrey S,Engbring Laurel,Shapiro Beth International journal of paleopathology Next Generation Sequencing (NGS) of ancient dental calculus samples from a prehistoric site in San Francisco Bay, CA-SCL-919, reveals a wide range of potentially pathogenic bacteria. One older adult woman, in particular, had high levels of Neisseria meningitidis and low levels of Haemophilus influenzae, species that were not observed in the calculus from three other individuals. Combined with the presence of incipient endocranial lesions and pronounced meningeal grooves, we interpret this as an ancient case of meningococcal disease. This disease afflicts millions around the globe today, but little is known about its (pre)history. With additional sampling, we suggest NGS of calculus offers an exciting new window into the evolutionary history of these bacterial species and their interactions with humans. 10.1016/j.ijpp.2018.05.001
    Dental calculus and isotopes provide direct evidence of fish and plant consumption in Mesolithic Mediterranean. Cristiani Emanuela,Radini Anita,Borić Dušan,Robson Harry K,Caricola Isabella,Carra Marialetizia,Mutri Giuseppina,Oxilia Gregorio,Zupancich Andrea,Šlaus Mario,Vujević Dario Scientific reports In this contribution we dismantle the perceived role of marine resources and plant foods in the subsistence economy of Holocene foragers of the Central Mediterranean using a combination of dental calculus and stable isotope analyses. The discovery of fish scales and flesh fragments, starch granules and other plant and animal micro-debris in the dental calculus of a Mesolithic forager dated to the end of the 8th millenium BC and buried in the Vlakno Cave on Dugi Otok Island in the Croatian Archipelago demonstrates that marine resources were regularly consumed by the individual together with a variety of plant foods. Since previous stable isotope data in the Eastern Adriatic and the Mediterranean region emphasises that terrestrial-based resources contributed mainly to Mesolithic diets in the Mediterranean Basin, our results provide an alternative view of the dietary habits of Mesolithic foragers in the Mediterranean region based on a combination of novel methodologies and data. 10.1038/s41598-018-26045-9
    Tooth grooves, occlusal striations, dental calculus, and evidence for fiber processing in an Italian eneolithic/bronze age cemetery. Sperduti Alessandra,Giuliani Maria Rita,Guida Giuseppe,Petrone Pier Paolo,Rossi Paola Francesca,Vaccaro Serena,Frayer David W,Bondioli Luca American journal of physical anthropology OBJECTIVES:We conducted a systematic macroscopic and microscopic examination of occlusal and para-occlusal wear in a large dental sample (n = 3,014) from 217 individuals dated to the Early Bronze age site of Gricignano d'Aversa, Italy. We used macroscopic and microscopic techniques to document nondietary occlusal and para-occlusal wear and to analyze calculus inclusions in some of the teeth. In combining an analysis of the wear with the calculus inclusions we linked the specific wear to the likely fiber that was involved in producing it. MATERIALS AND METHODS:Teeth and their high resolution epoxy casts were analyzed through SEM and reflected light microscopes. Nineteen individuals (fifteen with activity induced dental modifications and four as a control sample) were examined for the presence of calculus inclusions. RESULTS:Activity induced dental modifications (AIDMs), notches, grooves and micro-striations, were found in the 62.2% of the adult females, in 21.2% of the adults of unknown sex and in a single male. We found the full spectrum of dental manipulations from very minor nonocclusal wear in some young individuals to severe attrition at the other extreme. The width of the striations and grooves, mostly on the upper incisors, suggests a craft activity involving fibers and thread production and manipulation. From the dental calculus of two females with grooves and striations, we extracted three fragments of fibers, identified as hemp (Cannabis, sp.). Previously from Gricignano woven hemp fibers were found on both surfaces of a metal blade associated with a male burial. DISCUSSION:This study found the co-occurrence of tooth AIDMs and the actual fibers preserved in the dental calculus. As more work is done analyzing dental calculus in a variety of humans, it is apparent that this biological material holds rich resources documenting non-dietary habits. 10.1002/ajpa.23619
    Medieval women's early involvement in manuscript production suggested by lapis lazuli identification in dental calculus. Radini A,Tromp M,Beach A,Tong E,Speller C,McCormick M,Dudgeon J V,Collins M J,Rühli F,Kröger R,Warinner C Science advances During the European Middle Ages, the opening of long-distance Asian trade routes introduced exotic goods, including ultramarine, a brilliant blue pigment produced from lapis lazuli stone mined only in Afghanistan. Rare and as expensive as gold, this pigment transformed the European color palette, but little is known about its early trade or use. Here, we report the discovery of lapis lazuli pigment preserved in the dental calculus of a religious woman in Germany radiocarbon-dated to the 11th or early 12th century. The early use of this pigment by a religious woman challenges widespread assumptions about its limited availability in medieval Europe and the gendered production of illuminated texts. 10.1126/sciadv.aau7126
    Commensal and Pathogenic Members of the Dental Calculus Microbiome of Badia Pozzeveri Individuals from the 11th to 19th Centuries. Santiago-Rodriguez Tasha M,Fornaciari Antonio,Fornaciari Gino,Luciani Stefania,Marota Isolina,Vercellotti Giuseppe,Toranzos Gary A,Giuffra Valentina,Cano Raul J Genes The concept of the human oral microbiome was applied to understand health and disease, lifestyles, and dietary habits throughout part of human history. In the present study, we augment the understanding of ancient oral microbiomes by characterizing human dental calculus samples recovered from the ancient Abbey of Badia Pozzeveri (central Italy), with differences in socioeconomic status, time period, burial type, and sex. Samples dating from the Middle Ages (11th century) to the Industrial Revolution era (19th century) were characterized using high-throughput sequencing of the 16S ribosomal RNA (rRNA) gene V4 region. Consistent with previous studies, individuals from Badia Pozzeveri possessed commensal oral bacteria that resembled modern oral microbiomes. These results suggest that members of the oral microbiome are ubiquitous despite differences in geographical regions, time period, sex, and socioeconomic status. The presence of fecal bacteria could be in agreement with poor hygiene practices, consistent with the time period. Respiratory tract, nosocomial, and other rare pathogens detected in the dental calculus samples are intriguing and could suggest subject-specific comorbidities that could be reflected in the oral microbiome. 10.3390/genes10040299
    Calculus and survivorship in medieval London: The association between dental disease and a demographic measure of general health. Yaussy Samantha L,DeWitte Sharon N American journal of physical anthropology OBJECTIVES:Dental plaque is associated with a variety of systemic diseases and mortality risks in living populations. However, bioarchaeologists have not fully investigated the mortality risks associated with plaque (or its mineralized form, calculus) in the past. This study examines the relationship between survivorship and calculus in a medieval skeletal sample. MATERIALS AND METHODS:Our sample (n = 1,098) from four medieval London cemeteries, c. 1000-1540 CE, includes people who died under attritional (normal) and catastrophic (famine and plague) conditions. The associations between age and the presence of dental calculus on the permanent left first mandibular molar are assessed using binary logistic regression and Kaplan-Meier survival analysis. RESULTS:The regression results indicate a significant negative relationship between age and calculus presence for individuals of all ages who died under normal mortality conditions and for adults who died under both normal and catastrophic conditions. Survival analysis reveals decreased survivorship for people of all ages with calculus under normal mortality conditions. Similarly, during conditions of catastrophic mortality, adult males with calculus suffered reduced survivorship compared to males without it, though there was no difference in survivorship between adult females with and without calculus. CONCLUSIONS:These results suggest that, as in modern populations, calculus accumulation in the inhabitants of medieval London reflects a greater risk of premature death. The evaluation of calculus, a potential measure of underlying frailty, in the context of a demographic measure of general health suggests that it might provide insights into health in past populations. 10.1002/ajpa.23772
    Quantitative metaproteomics of medieval dental calculus reveals individual oral health status. Jersie-Christensen Rosa R,Lanigan Liam T,Lyon David,Mackie Meaghan,Belstrøm Daniel,Kelstrup Christian D,Fotakis Anna K,Willerslev Eske,Lynnerup Niels,Jensen Lars J,Cappellini Enrico,Olsen Jesper V Nature communications The composition of ancient oral microbiomes has recently become accessible owing to advanced biomolecular methods such as metagenomics and metaproteomics, but the utility of metaproteomics for such analyses is less explored. Here, we use quantitative metaproteomics to characterize the dental calculus associated with the remains of 21 humans retrieved during the archeological excavation of the medieval (ca. 1100-1450 CE) cemetery of Tjærby, Denmark. We identify 3671 protein groups, covering 220 bacterial species and 81 genera across all medieval samples. The metaproteome profiles of bacterial and human proteins suggest two distinct groups of archeological remains corresponding to health-predisposed and oral disease-susceptible individuals, which is supported by comparison to the calculus metaproteomes of healthy living individuals. Notably, the groupings identified by metaproteomics are not apparent from the bioarchaeological analysis, illustrating that quantitative metaproteomics has the potential to provide additional levels of molecular information about the oral health status of individuals from archeological contexts. 10.1038/s41467-018-07148-3
    Pleistocene dental calculus: Recovering information on Paleolithic food items, medicines, paleoenvironment and microbes. Hardy Karen,Buckley Stephen,Copeland Les Evolutionary anthropology Dental calculus is now widely used to recover information on items ingested in the past. It is particularly valuable in the earlier Paleolithic, where recovered data may represent the only evidence for plant use. Several recovery methods are used and each one produces different results. Biomolecular markers and genetic material recovered from dental calculus is providing new data on identifiable dietary and medicinal items and human microbial communities. The recovery of microfossils, in particular, starch granules, has triggered a new awareness of the role of plants in the diet throughout the Paleolithic. However, the minute amount of material recovered has little relationship with food eaten during a person's life, while salivary amylase breaks down cooked starch. Therefore, broader dietary interpretations and detection of cooked food are problematic. The study of ancient dental calculus holds great potential to recover information about past lives, within realistic parameters. 10.1002/evan.21718
    Cleaning protocol of archaeological dental calculus: A methodological proposal for vegetable microremains analysis. Tavarone Aldana,de Los Milagros Colobig María,Passeggi Esteban,Fabra Mariana American journal of physical anthropology OBJECTIVES:One of the major drawbacks involves the contamination produced during the sampling of dental calculus samples due to their manipulation with non-sterilized latex gloves containing maize starch. These gloves have been commonly used for the sampling, cleaning and conservation of archaeological materials. The objective of the present work is to propose a protocol for the cleaning of dental calculus samples that have been contaminated, allowing the removal of such material from the calculus surface. The application of this protocol guarantees the exogenous elements elimination without causing the loss of the archaeological material, since as it is a nondestructive and/or invasive cleaning method, it remains contained within the dental calculus which protects and guarantees its preservation. MATERIALS AND METHODS:Fifty-seven fragments of dental calculus were recovered from several archaeological sites in the province of Córdoba, Argentina, dated by AMS between 4058 and 387 14C years BP. A sample comprising 22 fragments larger than 2 mm, which represent 38.59% of the total, were subjected to four cleaning stages through distilled and centrifugal water. As a control test, 10 fragments of human dental tartar were taken, known to have not been previously handled with powdered latex gloves. RESULTS AND DISCUSSION:The results show significant differences between the four cleaning stages, representing an important reduction of modern starch grains adhered to the surface of archaeological samples of human dental calculus. 10.1002/ajpa.23630
    Proteomic evidence of dietary sources in ancient dental calculus. Hendy Jessica,Warinner Christina,Bouwman Abigail,Collins Matthew J,Fiddyment Sarah,Fischer Roman,Hagan Richard,Hofman Courtney A,Holst Malin,Chaves Eros,Klaus Lauren,Larson Greger,Mackie Meaghan,McGrath Krista,Mundorff Amy Z,Radini Anita,Rao Huiyun,Trachsel Christian,Velsko Irina M,Speller Camilla F Proceedings. Biological sciences Archaeological dental calculus has emerged as a rich source of ancient biomolecules, including proteins. Previous analyses of proteins extracted from ancient dental calculus revealed the presence of the dietary milk protein β-lactoglobulin, providing direct evidence of dairy consumption in the archaeological record. However, the potential for calculus to preserve other food-related proteins has not yet been systematically explored. Here we analyse shotgun metaproteomic data from 100 archaeological dental calculus samples ranging from the Iron Age to the post-medieval period (eighth century BC to nineteenth century AD) in England, as well as 14 dental calculus samples from contemporary dental patients and recently deceased individuals, to characterize the range and extent of dietary proteins preserved in dental calculus. In addition to milk proteins, we detect proteomic evidence of foodstuffs such as cereals and plant products, as well as the digestive enzyme salivary amylase. We discuss the importance of optimized protein extraction methods, data analysis approaches and authentication strategies in the identification of dietary proteins from archaeological dental calculus. This study demonstrates that proteomic approaches can robustly identify foodstuffs in the archaeological record that are typically under-represented due to their poor macroscopic preservation. 10.1098/rspb.2018.0977
    The efficacy of whole human genome capture on ancient dental calculus and dentin. Ziesemer Kirsten A,Ramos-Madrigal Jazmín,Mann Allison E,Brandt Bernd W,Sankaranarayanan Krithivasan,Ozga Andrew T,Hoogland Menno,Hofman Courtney A,Salazar-García Domingo C,Frohlich Bruno,Milner George R,Stone Anne C,Aldenderfer Mark,Lewis Cecil M,Hofman Corinne L,Warinner Christina,Schroeder Hannes American journal of physical anthropology OBJECTIVES:Dental calculus is among the richest known sources of ancient DNA in the archaeological record. Although most DNA within calculus is microbial, it has been shown to contain sufficient human DNA for the targeted retrieval of whole mitochondrial genomes. Here, we explore whether calculus is also a viable substrate for whole human genome recovery using targeted enrichment techniques. MATERIALS AND METHODS:Total DNA extracted from 24 paired archaeological human dentin and calculus samples was subjected to whole human genome enrichment using in-solution hybridization capture and high-throughput sequencing. RESULTS:Total DNA from calculus exceeded that of dentin in all cases, and although the proportion of human DNA was generally lower in calculus, the absolute human DNA content of calculus and dentin was not significantly different. Whole genome enrichment resulted in up to four-fold enrichment of the human endogenous DNA content for both dentin and dental calculus libraries, albeit with some loss in complexity. Recovering more on-target reads for the same sequencing effort generally improved the quality of downstream analyses, such as sex and ancestry estimation. For nonhuman DNA, comparison of phylum-level microbial community structure revealed few differences between precapture and postcapture libraries, indicating that off-target sequences in human genome-enriched calculus libraries may still be useful for oral microbiome reconstruction. DISCUSSION:While ancient human dental calculus does contain endogenous human DNA sequences, their relative proportion is low when compared with other skeletal tissues. Whole genome enrichment can help increase the proportion of recovered human reads, but in this instance enrichment efficiency was relatively low when compared with other forms of capture. We conclude that further optimization is necessary before the method can be routinely applied to archaeological samples. 10.1002/ajpa.23763
    Exploitation and utilization of tropical rainforests indicated in dental calculus of ancient Oceanic Lapita culture colonists. Tromp Monica,Matisoo-Smith Elizabeth,Kinaston Rebecca,Bedford Stuart,Spriggs Matthew,Buckley Hallie Nature human behaviour Remote Oceania, which largely consists of islands covered in tropical forests, was the last region on earth to be successfully colonized by humans, beginning 3,000 years ago. We examined human dental calculus from burials in an ancient Lapita culture cemetery to gain insight into the early settlement of this previously untouched tropical environment, specifically on the island of Efate in Vanuatu. Dental calculus is an ideal material to analyse questions of human and plant interactions due to the ingestion of plant-derived microparticles that become incorporated into the calculus as it forms throughout a person's life. Most of the microparticles identified here are from tree and shrub resources, including a ~2,900 calibrated (cal) BP example of banana in Remote Oceania, providing direct evidence for the importance of forests and arboriculture during the settlement of Remote Oceania. 10.1038/s41562-019-0808-y
    Investigating Plant Micro-Remains Embedded in Dental Calculus of the Phoenician Inhabitants of Motya (Sicily, Italy). D'Agostino Alessia,Canini Antonella,Di Marco Gabriele,Nigro Lorenzo,Spagnoli Federica,Gismondi Angelo Plants (Basel, Switzerland) Plant records reveal remarkable evidence about past environments and human cultures. Exploiting dental calculus analysis and using a combined approach of microscopy and gas chromatography mass spectrometry, our research outlines dietary ecology and phytomedicinal practices of the ancient community of Motya (Sicily, eight to sixth century BC), one of the most important Phoenician settlements in the Mediterranean basin. Micro-remains suggest use or consumption of Triticeae cereals, and animal-derived sources (e.g., milk and aquatic birds). Markers of grape (or wine), herbs, and rhizomes, endemic of Mediterranean latitudes and the East, provide insight into the subsistence of this colony, in terms of foodstuffs and phytotherapeutic products. The application of resins and wood of Gymnosperms for social and cultural purposes is hypothesized through the identification of Pinaceae secondary metabolites and pollen grains. The information hidden in dental calculus discloses the strong human-plant interaction in Motya's Phoenician community, in terms of cultural traditions and land use. 10.3390/plants9101395
    Differential preservation of endogenous human and microbial DNA in dental calculus and dentin. Mann Allison E,Sabin Susanna,Ziesemer Kirsten,Vågene Åshild J,Schroeder Hannes,Ozga Andrew T,Sankaranarayanan Krithivasan,Hofman Courtney A,Fellows Yates James A,Salazar-García Domingo C,Frohlich Bruno,Aldenderfer Mark,Hoogland Menno,Read Christopher,Milner George R,Stone Anne C,Lewis Cecil M,Krause Johannes,Hofman Corinne,Bos Kirsten I,Warinner Christina Scientific reports Dental calculus (calcified dental plaque) is prevalent in archaeological skeletal collections and is a rich source of oral microbiome and host-derived ancient biomolecules. Recently, it has been proposed that dental calculus may provide a more robust environment for DNA preservation than other skeletal remains, but this has not been systematically tested. In this study, shotgun-sequenced data from paired dental calculus and dentin samples from 48 globally distributed individuals are compared using a metagenomic approach. Overall, we find DNA from dental calculus is consistently more abundant and less contaminated than DNA from dentin. The majority of DNA in dental calculus is microbial and originates from the oral microbiome; however, a small but consistent proportion of DNA (mean 0.08 ± 0.08%, range 0.007-0.47%) derives from the host genome. Host DNA content within dentin is variable (mean 13.70 ± 18.62%, range 0.003-70.14%), and for a subset of dentin samples (15.21%), oral bacteria contribute > 20% of total DNA. Human DNA in dental calculus is highly fragmented, and is consistently shorter than both microbial DNA in dental calculus and human DNA in paired dentin samples. Finally, we find that microbial DNA fragmentation patterns are associated with guanine-cytosine (GC) content, but not aspects of cellular structure. 10.1038/s41598-018-28091-9
    Metagenomic analysis of dental calculus in ancient Egyptian baboons. Ottoni Claudio,Guellil Meriam,Ozga Andrew T,Stone Anne C,Kersten Oliver,Bramanti Barbara,Porcier Stéphanie,Van Neer Wim Scientific reports Dental calculus, or mineralized plaque, represents a record of ancient biomolecules and food residues. Recently, ancient metagenomics made it possible to unlock the wealth of microbial and dietary information of dental calculus to reconstruct oral microbiomes and lifestyle of humans from the past. Although most studies have so far focused on ancient humans, dental calculus is known to form in a wide range of animals, potentially informing on how human-animal interactions changed the animals' oral ecology. Here, we characterise the oral microbiome of six ancient Egyptian baboons held in captivity during the late Pharaonic era (9-6 centuries BC) and of two historical baboons from a zoo via shotgun metagenomics. We demonstrate that these captive baboons possessed a distinctive oral microbiome when compared to ancient and modern humans, Neanderthals and a wild chimpanzee. These results may reflect the omnivorous dietary behaviour of baboons, even though health, food provisioning and other factors associated with human management, may have changed the baboons' oral microbiome. We anticipate our study to be a starting point for more extensive studies on ancient animal oral microbiomes to examine the extent to which domestication and human management in the past affected the diet, health and lifestyle of target animals. 10.1038/s41598-019-56074-x
    Dental Calculus as a Tool to Study the Evolution of the Mammalian Oral Microbiome. Brealey Jaelle C,Leitão Henrique G,van der Valk Tom,Xu Wenbo,Bougiouri Katia,Dalén Love,Guschanski Katerina Molecular biology and evolution Dental calculus, the calcified form of the mammalian oral microbial plaque biofilm, is a rich source of oral microbiome, host, and dietary biomolecules and is well preserved in museum and archaeological specimens. Despite its wide presence in mammals, to date, dental calculus has primarily been used to study primate microbiome evolution. We establish dental calculus as a valuable tool for the study of nonhuman host microbiome evolution, by using shotgun metagenomics to characterize the taxonomic and functional composition of the oral microbiome in species as diverse as gorillas, bears, and reindeer. We detect oral pathogens in individuals with evidence of oral disease, assemble near-complete bacterial genomes from historical specimens, characterize antibiotic resistance genes, reconstruct components of the host diet, and recover host genetic profiles. Our work demonstrates that metagenomic analyses of dental calculus can be performed on a diverse range of mammalian species, which will allow the study of oral microbiome and pathogen evolution from a comparative perspective. As dental calculus is readily preserved through time, it can also facilitate the quantification of the impact of anthropogenic changes on wildlife and the environment. 10.1093/molbev/msaa135
    Functional diversity of microbial ecologies estimated from ancient human coprolites and dental calculus. Jacobson David K,Honap Tanvi P,Monroe Cara,Lund Justin,Houk Brett A,Novotny Anna C,Robin Cynthia,Marini Elisabetta,Lewis Cecil M Philosophical transactions of the Royal Society of London. Series B, Biological sciences Human microbiome studies are increasingly incorporating macroecological approaches, such as community assembly, network analysis and functional redundancy to more fully characterize the microbiome. Such analyses have not been applied to ancient human microbiomes, preventing insights into human microbiome evolution. We address this issue by analysing published ancient microbiome datasets: coprolites from Rio Zape ( = 7; 700 CE Mexico) and historic dental calculus ( = 44; 1770-1855 CE, UK), as well as two novel dental calculus datasets: Maya ( = 7; 170 BCE-885 CE, Belize) and Nuragic Sardinians ( = 11; 1400-850 BCE, Italy). Periodontitis-associated bacteria (, and ) were identified as keystone taxa in the dental calculus datasets. Coprolite keystone taxa included known short-chain fatty acid producers () and potentially disease-associated bacteria (, . Overlap in ecological profiles between ancient and modern microbiomes was indicated by similarity in functional response diversity profiles between contemporary hunter-gatherers and ancient coprolites, as well as parallels between ancient Maya, historic UK, and modern Spanish dental calculus; however, the ancient Nuragic dental calculus shows a distinct ecological structure. We detected key ecological signatures from ancient microbiome data, paving the way to expand understanding of human microbiome evolution. This article is part of the theme issue 'Insights into health and disease from ancient biomolecules'. 10.1098/rstb.2019.0586
    Ancient DNA analysis of food remains in human dental calculus from the Edo period, Japan. Sawafuji Rikai,Saso Aiko,Suda Wataru,Hattori Masahira,Ueda Shintaroh PloS one Although there are many methods for reconstructing diets of the past, detailed taxon identification is still challenging, and most plants hardly remain at a site. In this study, we applied DNA metabarcoding to dental calculus of premodern Japan for the taxonomic identification of food items. DNA was extracted from 13 human dental calculi from the Unko-in site (18th-19th century) of the Edo period, Japan. Polymerase chain reaction (PCR) and sequencing were performed using a primer set specific to the genus Oryza because rice (Oryza sativa) was a staple food and this was the only member of this genus present in Japan at that time. DNA metabarcoding targeting plants, animals (meat and fish), and fungi were also carried out to investigate dietary diversity. We detected amplified products of the genus Oryza from more than half of the samples using PCR and Sanger sequencing. DNA metabarcoding enabled us to identify taxa of plants and fungi, although taxa of animals were not detected, except human. Most of the plant taxonomic groups (family/genus level) are present in Japan and include candidate species consumed as food at that time, as confirmed by historical literature. The other groups featured in the lifestyle of Edo people, such as for medicinal purposes and tobacco. The results indicate that plant DNA analysis from calculus provides information about food diversity and lifestyle habits from the past and can complement other analytical methods such as microparticle analysis and stable isotope analysis. 10.1371/journal.pone.0226654
    Multi-omic detection of in archaeological human dental calculus. Fotakis Anna K,Denham Sean D,Mackie Meaghan,Orbegozo Miren Iraeta,Mylopotamitaki Dorothea,Gopalakrishnan Shyam,Sicheritz-Pontén Thomas,Olsen Jesper V,Cappellini Enrico,Zhang Guojie,Christophersen Axel,Gilbert M Thomas P,Vågene Åshild J Philosophical transactions of the Royal Society of London. Series B, Biological sciences Mineralized dental plaque (calculus) has proven to be an excellent source of ancient biomolecules. Here we present a genome (6.6-fold), the causative agent of leprosy, recovered via shotgun sequencing of sixteenth-century human dental calculus from an individual from Trondheim, Norway. When phylogenetically placed, this genome falls in branch 3I among the diversity of other contemporary ancient strains from Northern Europe. Moreover, ancient mycobacterial peptides were retrieved via mass spectrometry-based proteomics, further validating the presence of the pathogen. can readily be detected in the oral cavity and associated mucosal membranes, which likely contributed to it being incorporated into this individual's dental calculus. This individual showed some possible, but not definitive, evidence of skeletal lesions associated with early-stage leprosy. This study is the first known example of successful multi-omics retrieval of from archaeological dental calculus. Furthermore, we offer new insights into dental calculus as an alternative sample source to bones or teeth for detecting and molecularly characterizing in individuals from the archaeological record. This article is part of the theme issue 'Insights into health and disease from ancient biomolecules'. 10.1098/rstb.2019.0584