Assessing the validity of a calcifying oral biofilm model as a suitable proxy for dental calculus

<ns3:p>Background Dental calculus is increasingly used by researchers to study dietary patterns in past populations. The benefits of using dental calculus for this purpose have been clearly demonstrated, with dental calculus harbouring a wealth of microremains and biomarkers for health and diet. Previous studies have demonstrated some of the limitations and biases of how methods of processing may overlook, or even remove, some of the important information contained within the mineralised matrix. However, there are many factors that are impossible to account for *in vivo* and in archaeological material, such as exact dietary intake, and individual factors such as pH and enzyme activity, leaving some limitations that require a different approach. Methods We present a protocol for creating a calcifying oral biofilm model that can be used to explore the biases and limitations of dental calculus as a medium for paleodietary reconstructions. We report the microbial and mineral composition, using metagenomic classification and Fourier transform infrared (FTIR) spectroscopy, respectively, in an effort to validate the model calculus as an appropriate proxy to natural dental calculus. Results Metagenomic classification showed a microbial profile predominantly made up of (facultative) anaerobes, with a community structure that was somewhat distinct from other oral reference samples. The core genera of the model consisted of oral species, but clustered separately from oral reference samples due to a higher abundance of anaerobes. Mineral and organic components of our model mimic that of the modern and archaeological reference calculus, with carbonated hydroxyapatite as the principal compound, consistent with natural human-derived calculus. Conclusions Overall our model is indicative of an oral environment, and has potential to validate current methods used in the analysis of archaeological dental calculus, and should be used to complement, rather than replace current *in vivo* studies.</ns3:p>