The lower jaw of one of the individuals used in the experiment to assess whether the sex of human remains can be determined using distinctive peptides found in tooth enamel. (Image: Janet Montgomery)

In the first ‘Science Notes’ (CA 333), we discussed the identification of a possible female Viking warrior using ancient DNA analysis. This is a guaranteed way to confirm sex in human remains, but can be costly, time-consuming, and destructive to the bone, meaning that it is not feasible when a project needs to determine the sex of a large number of skeletons.

Also mentioned in that article was the fact that the skeleton in question had previously been identified as potentially female using osteological methods that look at the morphology of the skull and pelvis. This technique is based on developmental changes the skeleton undergoes during puberty due to the influx in sex hormones. While this can be a very effective method – its results are usually estimated at between 80% and 95% accuracy depending on the preservation of the bone, among other factors – individuals that have not yet undergone puberty (infant and juvenile skeletons) do not display these crucial differences. While previously some techniques have attempted to identify juvenile sex through various morphological and metric characteristics, none have proven to be consistently reliable.

This perennial problem in bioarchaeology has severely limited osteologists’ ability to analyse non-adult remains, particularly in terms of accurately reconstructing varying demographic and health profiles between boys and girls. A new technique, however – developed collaboratively by protein chemists, dentists, and archaeologists from the Universities of Brighton, São Paulo, and Durham, respectively – has potentially found a way of overcoming this limitation by analysing peptides. These short chains of amino acids are found in tooth enamel and one has been discovered to be distinct between the two sexes. Tooth enamel is one of the hardest, and hardiest, parts of the body: it often survives well in the burial environment and is less prone to contamination. This makes it ideal for use in sex analysis, as it means it can be both reliable and easily replicable in many instances.

There are peptide remnants from three major proteins that originate from tooth enamel: amelogenin, ameloblastin, and enamelin. Amelogenin has two sexually dimorphic variants: AMELX, which is found in both males and females, and AMELY, found only in males. For AMELY, a peptide contains an additional methionine amino acid. The research team discovered that in archaeological remains the oxidised version of the AMELY peptide was well preserved in the tooth enamel of older samples, while the unoxidised version may not be. They therefore decided to use the presence of oxidised AMELY as an indicator for male sex, and its absence as an indicator for female sex.

The team then did a blind study where they analysed the tooth enamel of seven skeletons of known sex – as identified through coffin plates and grave monuments – from a 19th-century cemetery in Fewston, near Harrogate in North Yorkshire. They also analysed male and female pairs (as confirmed using the morphological technique mentioned above) from three archaeological sites covering different periods – Neolithic, Anglo-Saxon, and late medieval – to determine the technique’s efficacy with older remains. In every case, they discovered that their peptide results agreed with the sex previously assigned to the skeleton, even in the Neolithic remains. Overall, this methodology proved to be not only accurate, but also less destructive and less expensive than ancient DNA analysis – winning the trifecta in terms of replicability in different archaeological and modern contexts.

While the technique is still in its early stages, it has the potential to broaden the types of research questions osteologists are able to address and could lead to an even greater understanding of child health and development in the past. As the authors summarise in the paper outlining their findings, ‘The ability to determine the sex of infant and juvenile remains completely revolutionises studies of growth, childcare, epidemiology, and demography in the past. For the first time, it will allow osteologists to examine sex-specific cultural treatment and differentiate between the health of boys and girls, as well as sex-specific growth trajectories and past developmental milestones, such as age of puberty and subsequent repercussions for fertility.’

The full paper, published in PNAS, can be read for free here:

This article appeared in CA 337.

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