Hunter-Gatherers, Archaeogenomics, and the Evolutionary History of the Foxes of California's Channel Islands

Understanding human-animal relationships is a fundamental area of archaeological research. Throughout human history, animals have been sources of food, raw materials, labor, and companionship. Humans have also had an important influence on animal populations, including extinction, domestication, and translocation. Recently, archaeological research on the interactions between humans and animals has also helped us understand the contemporary status of animal populations, providing important insights for conservation biology and establishing a new research agenda, conservation archaeogenomics. In this dissertation, I define the field of conservation of archaeogenomics and develop a case study of how archaeological, genomic, and isotope data can be integrated to inform the conservation of an endangered carnivore. The endemic island fox (Urocyon littoralis) of California's Channel Islands is a federally listed endangered species and has been the subject of considerable conservation research, including a captive breeding program. Despite decades of research, significant questions remain about when foxes colonized the Channel Islands and the role that Native Americans may have played in their introduction and dispersal to six islands. Using mitochondrial genomes of 185 extant island and gray fox samples, I demonstrate that island and mainland lineages diverge ~9200-7100 cal BP and were quickly dispersed to the other Channel Islands, likely by humans. I also explore the possibility of a deliberate introduction by Native Americans using isotope data. I did not detect evidence of human resource provisioning of island foxes from early archaeological contexts as might be expected if they were introduced by ancient peoples. However, I did detect evidence of human resource provisioning on San Nicolas Island in the late Holocene and developed a long-term dataset documenting ~7300 years of foraging ecology in the endangered island fox. Archaeological investigations of human-animal relationships through time can help document the influence of Native Americans on species distribution, abundance, and ecology. Understanding how species and humans adapted to and influenced changing environments in the past will inform decisions about protecting, preserving, and restoring biodiversity in the future. This dissertation demonstrates the importance of integrating archaeology and genomics for understanding ancient and modern human environmental relationships and modern conservation biology

Integrating archaeology and ancient DNA analysis to address invasive species colonization in the Gulf of Alaska

The intentional and unintentional movement of plants and animals by humans has transformed ecosystems and landscapes globally. Assessing when and how a species was introduced are central to managing these transformed landscapes, particularly in island environments. In the Gulf of Alaska, there is considerable interest in the history of mammal introductions and rehabilitating Gulf of Alaska island environments by eradicating mammals classified as invasive species. The Arctic ground squirrel (Urocitellus parryii) is of concern because it affects vegetation and seabirds on Gulf of Alaska islands. This animal is assumed to have been introduced by historic settlers; however, ground squirrel remains in the prehistoric archaeological record of Chirikof Island, Alaska, challenge this timeline and suggest they colonized the islands long ago. We used 3 lines of evidence to address this problem: direct radiocarbon dating of archaeological squirrel remains; evidence of prehistoric human use of squirrels; and ancient DNA analysis of dated squirrel remains. Chirikof squirrels dated to at least 2000 years ago, and cut marks on squirrel bones suggested prehistoric use by people. Ancient squirrels also shared a mitochondrial haplotype with modern Chirikof squirrels. These results suggest that squirrels have been on Chirikof longer than previously assumed and that the current population of squirrels is closely related to the ancient population. Thus, it appears ground squirrels are not a recent, human‐mediated introduction and may have colonized the island via a natural dispersal event or an ancient human translocation.We thank T. Rick, D. Grayson, R. Fleischer, M. Hawkins, A. West, and C. Mikeska for their contributions to this research. We also thank 3 reviewers and the editors of Conservation Biology who greatly improved this paper. This work was funded by the National Geographic Society, the University of Maine, the Smithsonian Institution, and Boston University. The findings and conclusions in this article are those of the authors and do not necessarily represent the views of the U.S. Fish and Wildlife Service. (National Geographic Society; University of Maine; Smithsonian Institution; Boston University)Published versio

A unified protocol for simultaneous extraction of DNA and proteins from archaeological dental calculus

Archaeological materials are a finite resource, and efforts should be made to minimize destructive analyses. This can be achieved by using protocols combining extraction of several types of biomolecules or microparticles, which decreases the material needed for analyses while maximizing the information yield. Archaeological dental calculus is a source of several different types of biomolecules, as well as microfossils, and can tell us about the human host, microbiome, diet, and even occupational activities. Here, we present a unified protocol allowing for simultaneous extraction of DNA and proteins from a single sample of archaeological dental calculus. We evaluate the protocol on dental calculus from six individuals from a range of time periods and estimated preservation states, and compare it against previously published DNA-only and protein-only protocols. We find that most aspects of downstream analyses are unaltered by the unified protocol, although minor shifts in the recovered proteome can be detected, such as a slight loss of hydrophilic proteins. Total protein recovery depends on both the amount of starting material and choice of extraction protocol, whereas total DNA recovery is significantly reduced using the unified protocol (mean 43%). Nevertheless, total DNA recovery from dental calculus is generally very high, and we found no differences in DNA fragment characteristics or taxonomic profile between the protocols. In conclusion, the unified protocol allows for simultaneous extraction of two complementary lines of biomolecular evidence from archaeological dental calculus without compromising downstream results, thereby minimizing the need for destructive analysis of this finite resource

Millennial-scale sustainability of the Chesapeake Bay Native American oyster fishery

Estuaries around the world are in a state of decline following decades or more of overfishing, pollution, and climate change. Oysters (Ostreidae), ecosystem engineers in many estuaries, influence water quality, construct habitat, and provide food for humans and wildlife. In North America\u27s Chesapeake Bay, once-thriving eastern oyster (Crassostrea virginica) populations have declined dramatically, making their restoration and conservation extremely challenging. Here we present data on oyster size and human harvest from Chesapeake Bay archaeological sites spanning similar to 3,500 y of Native American, colonial, and historical occupation. We compare oysters from archaeological sites with Pleistocene oyster reefs that existed before human harvest, modern oyster reefs, and other records of human oyster harvest from around the world. Native American fisheries were focused on nearshore oysters and were likely harvested at a rate that was sustainable over centuries to millennia, despite changing Holocene climatic conditions and sea-level rise. These data document resilience in oyster populations under long-term Native American harvest, sea-level rise, and climate change; provide context for managing modern oyster fisheries in the Chesapeake Bay and elsewhere around the world; and demonstrate an interdisciplinary approach that can be applied broadly to other fisheries

The efficacy of whole human genome capture on ancient dental calculus and dentin

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 fourfold 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

The genetic prehistory of the Andean highlands 7000 years BP though European contact

The peopling of the Andean highlands above 2500 m in elevation was a complex process that included cultural, biological, and genetic adaptations. Here, we present a time series of ancient whole genomes from the Andes of Peru, dating back to 7000 calendar years before the present (BP), and compare them to 42 new genome-wide genetic variation datasets from both highland and lowland populations. We infer three significant features: A split between low- And high-elevation populations that occurred between 9200 and 8200 BP; a population collapse after European contact that is significantly more severe in South American lowlanders than in highland populations; and evidence for positive selection at genetic loci related to starch digestion and plausibly pathogen resistance after European contact. We do not find selective sweep signals related to known components of the human hypoxia response, which may suggest more complex modes of genetic adaptation to high altitude

Do I have something in my teeth? The trouble with genetic analyses of diet from archaeological dental calculus

Dental calculus and other preserved microbiome substrates are an attractive target for dietary reconstruction in past populations through a variety of physical, chemical, and molecular means. Recently, studies have attempted to reconstruct diet from archaeological dental calculus using archaeogenetic techniques. While dental calculus may provide a relatively stable environment for DNA preservation, the detection of plants and animals possibly consumed by an individual through DNA analysis is primarily hindered by microbial richness and incomplete reference databases. Moreover, high genomic similarity within eukaryotic groups - such as mammals - can obfuscate precise taxonomic identification. In the current study we demonstrate the challenges associated with accurate taxonomic identification and authentication of dietary taxa in ancient DNA data using both synthetic and ancient dental calculus datasets. We highlight common errors and sources of contamination across ancient DNA datasets, provide recommendations for dietary DNA validation, and call for caution in the interpretation of diet from dental calculus and other archaeological microbiome substrates.J.A.F.Y. was partially funded by the European Research Council (ERC) under the European Union's Horizon 2020 research innovation programme (ERC-2015-StG 678901-FoodTransforms to Philipp W. Stockhammer, Ludwig Maximilian University Munich, Germany). Z.F. was supported by the Werner Siemens Stiftung through Dr. Christina Warinner. This research was supported in part through computational resources provided by the Department of Archaeogenetics at the Max Planck Institute for the Science of Human History (J.A.F.Y).Ye

Proteomic evidence of dietary sources in ancient dental calculus.

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

Adaptive divergence despite strong genetic drift: genomic analysis of the evolutionary mechanisms causing genetic differentiation in the island fox (\u3ci\u3eUrocyon littoralis\u3c/i\u3e)

The evolutionary mechanisms generating the tremendous biodiversity of islands have long fascinated evolutionary biologists. Genetic drift and divergent selection are pre- dicted to be strong on islands and both could drive population divergence and specia- tion. Alternatively, strong genetic drift may preclude adaptation. We conducted a genomic analysis to test the roles of genetic drift and divergent selection in causing genetic differentiation among populations of the island fox (Urocyon littoralis). This species consists of six subspecies, each of which occupies a different California Chan- nel Island. Analysis of 5293 SNP loci generated using Restriction-site Associated DNA (RAD) sequencing found support for genetic drift as the dominant evolutionary mech- anism driving population divergence among island fox populations. In particular, pop- ulations had exceptionally low genetic variation, small Ne (range = 2.1–89.7; median = 19.4), and significant genetic signatures of bottlenecks. Moreover, islands with the lowest genetic variation (and, by inference, the strongest historical genetic drift) were most genetically differentiated from mainland grey foxes, and vice versa, indicating genetic drift drives genome-wide divergence. Nonetheless, outlier tests identified 3.6–6.6% of loci as high FST outliers, suggesting that despite strong genetic drift, divergent selection contributes to population divergence. Patterns of similarity among populations based on high FST outliers mirrored patterns based on morphology, providing additional evidence that outliers reflect adaptive divergence. Extremely low genetic variation and small Ne in some island fox populations, particularly on San Nicolas Island, suggest that they may be vulnerable to fixation of deleterious alleles, decreased fitness and reduced adaptive potential

Trans-ancestry genome-wide association study identifies 12 genetic loci influencing blood pressure and implicates a role for DNA methylation

We carried out a trans-ancestry genome-wide association and replication study of blood pressure phenotypes among up to 320,251 individuals of East Asian, European and South Asian ancestry. We find genetic variants at 12 new loci to be associated with blood pressure (P = 3.9 × 10-11 to 5.0 × 10-21). The sentinel blood pressure SNPs are enriched for association with DNA methylation at multiple nearby CpG sites, suggesting that, at some of the loci identified, DNA methylation may lie on the regulatory pathway linking sequence variation to blood pressure. The sentinel SNPs at the 12 new loci point to genes involved in vascular smooth muscle (IGFBP3, KCNK3, PDE3A and PRDM6) and renal (ARHGAP24, OSR1, SLC22A7 and TBX2) function. The new and known genetic variants predict increased left ventricular mass, circulating levels of NT-proBNP, and cardiovascular and all-cause mortality (P = 0.04 to 8.6 × 10-6). Our results provide new evidence for the role of DNA methylation in blood pressure regulation