Method of micro-sampling human dentine collagen for stable isotope analysis.

RATIONALE: Sampling of dentine for stable carbon (δ13 C) and nitrogen (δ15 N) isotope ratios in the direction of tooth growth allows the study of temporal changes to the diet and physiological stress of an individual during tooth formation. Current methods of sampling permanent teeth using 1 mm increments provide temporal resolution of 6-9 months at best depending on the tooth chosen. Although this gives sufficient sample sizes for reliable analysis by mass spectrometry, sectioning the dentine across the incremental structures results in a rolling average of the isotope ratios. A novel method of incremental dentine collagen sampling has been developed to decrease the collagen increment size to 0.35 mm along the incremental structures, thus reducing averaging and improving the temporal resolution of short-term changes within the δ13 C and δ15 N values. METHODS: This study presents data for a MicroMill-assisted sampling method that allows for sampling at 0.35 mm width × 1 mm depth increments following the incremental growth pattern of dentine. A NewWave MicroMill was used to sample the demineralised dentine section of modern donated human third molars from Sudan and compared to data from the same teeth using the 1 mm incremental sectioning method 2 established by Beaumont et al. RESULTS: The δ13 C and δ15 N isotopic data showed an increased temporal resolution, with each increment providing data for 2-4 months of dentine formation. CONCLUSIONS: The data show the potential of this method for studying dietary reconstruction, nutritional stress, and physiological change with greater temporal resolution potentially to seasonal level and with less attenuation of the δ13 C and δ15 N values than was previously possible from human dentine.

Scottish soldiers from the Battle of Dunbar 1650: A prosopographical approach to a skeletal assemblage.

After the Battle Dunbar between English and Scottish forces in 1650, captured Scottish soldiers were imprisoned in Durham and many hundreds died there within a few weeks. The partial skeletal remains of 28 of these men were discovered in 2013. Building on previous osteological work, here we report wide-ranging scientific studies of the remains to address the following questions: Did they have comparable diet, health and disease throughout their lives? Did they have common histories of movement (or lack of movement) during their childhoods? Can we create a collective biography of these men? Strontium and oxygen isotope analysis of tooth enamel investigated childhood movement. Carbon and nitrogen isotope analysis of incrementally sampled dentine addressed childhood diet and nutrition. Metaproteomic analysis of dental calculus investigated oral microbiomes and food residues; this was complemented by microscopic analysis of debris in calculus from ingested materials. Selected individuals were examined for dental microwear. The extent of hydroxylation of proline in collagen was examined as a potential biomarker for scurvy. An osteobiography for each man was created using the full range of data generated about him, and these were synthesised using an approach based on the historical method for a collective biography or prosopography. The childhood residences of the men were primarily within the Midland Valley of Scotland, though some spent parts of their childhood outside the British Isles. This is concordant with the known recruitment areas of the Scottish army in 1650. Their diets included oats, brassicas and milk but little seafood, as expected for lowland rather than highland diets of the period. Childhood periods of starvation or illness were almost ubiquitous, but not simultaneous, suggesting regionally variable food shortages in the 1620s and 1630s. It is likely there was widespread low-level scurvy, ameliorating in later years of life, which suggests historically unrecorded shortages of fruit and vegetables in the early 1640s. Almost all men were exposed to burnt plant matter, probably as inhaled soot, and this may relate to the high proportion of them with of sinusitis. Interpersonal violence causing skeletal trauma was rare. Based on commonalities in their osteobiographies, we argue that these men were drawn from the same stratum of society. This study is perhaps the most extensive to date of individuals from 17th century Scotland. Combined with a precise historical context it allows the lives of these men to be investigated and compared to the historical record with unprecedented precision. It illustrates the power of archaeological science methods to confirm, challenge and complement historical evidence.

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

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

Basilar portion porosity: A pathological lesion possibly associated with infantile scurvy.

Recent analysis of the juvenile (≤12 years) human remains from a 19th century site in Wolverhampton, England revealed a relatively high level of nutritional deficiency diseases within the population. Indeed, 41.7% of the 48 juvenile skeletons analysed exhibited a combination of porous and proliferative bone lesions consistent with the pathological alterations associated with nutritional stress. This paper describes a pathological lesion on the inferior surface of the basilar portion of the occipital bone, not previously reported in association with infantile scurvy, but which was exhibited by 90% (N=9) of the 10 scorbutic individuals identified during this study.

Limits and possibilities in the geolocation of humans using multiple isotope ratios (H, O, N, C) of hair from east coast cities of the USA.

We examined multiple natural abundance isotope ratios of human hair to assess biological variability within and between geographic locations and, further, to determine how well these isotope values predict location of origin. Sampling locations feature differing seasonality and mobile populations as a robust test of the method. Serially-sampled hair from Cambridge, MA, USA, shows lower δ(2)H and δ(18)O variability over a one-year time course than model-predicted precipitation isotope ratios, but exhibits considerable differences between individuals. Along a ∼13° north-south transect in the eastern USA (Brookline, MA, 42.3 ° N, College Park, MD, 39.0 ° N, and Gainesville, FL, 29.7 ° N) δ(18)O in human hair shows relatively greater differences and tracks changes in drinking water isotope ratios more sensitively than δ(2)H. Determining the domicile of humans using isotope ratios of hair can be confounded by differing variability in hair δ(18)O and δ(2)H between locations, differential incorporation of H and O into this protein and, in some cases, by tap water δ(18)O and δ(2)H that differ significantly from predicted precipitation values. With these caveats, randomly chosen people in Florida are separated from those in the two more northerly sites on the basis of the natural abundance isotopes of carbon, nitrogen, hydrogen, and oxygen.

Identification of cooked bone using TEM imaging of bone collagen.

Mild heating (≤100° C, 1 h)-typical of cooking-does not lead to detectable changes in any biochemical parameter yet measured; consequently bones that have been cooked, but which have not reached a temperature that will induce charring go undetected. We have used a microscopy based approach to investigate changes in the organization of the bone protein, collagen, as it is heated, using bone from heating experiments, short term burials, and archaeological sites. The work has revealed that the presence of a mineral matrix stabilizes the collagen enabling the damage to accumulate, but preventing it from causing immediate gelatinization. Once the mineral is removed, the damage can be observed using appropriate visualization methods. This chapter describes the transmission electron microscopy (TEM) technique that has been used to detect cooked bone by visualizing minor heat-induced damage at the level of the collagen fibril.

Mineralization of the metre-long biosilica structures of glass sponges is templated on hydroxylated collagen.

The minerals involved in the formation of metazoan skeletons principally comprise glassy silica, calcium phosphate or carbonate. Because of their ancient heritage, glass sponges (Hexactinellida) may shed light on fundamental questions such as molecular evolution, the unique chemistry and formation of the first skeletal silica-based structures, and the origin of multicellular animals. We have studied anchoring spicules from the metre-long stalk of the glass rope sponge (Hyalonema sieboldi; Porifera, Class Hexactinellida), which are remarkable for their size, durability, flexibility and optical properties. Using slow-alkali etching of biosilica, we isolated the organic fraction, which was revealed to be dominated by a hydroxylated fibrillar collagen that contains an unusual [Gly-3Hyp-4Hyp] motif. We speculate that this motif is predisposed for silica precipitation, and provides a novel template for biosilicification in nature.

Preservation of ancient DNA in thermally damaged archaeological bone.

Evolutionary biologists are increasingly relying on ancient DNA from archaeological animal bones to study processes such as domestication and population dispersals. As many animal bones found on archaeological sites are likely to have been cooked, the potential for DNA preservation must be carefully considered to maximise the chance of amplification success. Here, we assess the preservation of mitochondrial DNA in a medieval cattle bone assemblage from Coppergate, York, UK. These bones have variable degrees of thermal alterations to bone collagen fibrils, indicative of cooking. Our results show that DNA preservation is not reliant on the presence of intact collagen fibrils. In fact, a greater number of template molecules could be extracted from bones with damaged collagen. We conclude that moderate heating of bone may enhance the retention of DNA fragments. Our results also indicate that ancient DNA preservation is highly variable, even within a relatively recent assemblage from contexts conducive to organic preservation, and that diagenetic parameters based on protein diagenesis are not always useful for predicting ancient DNA survival.

Comment on "Protein sequences from mastodon and Tyrannosaurus rex revealed by mass spectrometry".

We used authentication tests developed for ancient DNA to evaluate claims by Asara et al. (Reports, 13 April 2007, p. 280) of collagen peptide sequences recovered from mastodon and Tyrannosaurus rex fossils. Although the mastodon samples pass these tests, absence of amino acid composition data, lack of evidence for peptide deamidation, and association of alpha1(I) collagen sequences with amphibians rather than birds suggest that T. rex does not.