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Multidisciplinary analyses on ancient dental calculus revealed the possibility to reconstruct habits and diet of ancient human populations, investigate individual health status, as well as provide information on past environments. In the present study we applied both metagenomic and microscopic analysis on ancient human dental calculus in order to obtain a cross-section of the life conditions in a population of central Italy belonging to the Copper Age culture of Rinaldone (IV millennium BCE). The metagenomic profile suggested an agricultural subsistence and a dietary regimen particularly enriched in complex carbohydrates with low soluble fiber. Even bacterial functional profile seems to indicate an almost exclusive carbohydrates intake that could have favoured the occurrence of nutritional stress in the individuals. Exploring the diversity of the plant food consumed, we detected direct evidence of cereals such as wheat and/or barley, and found signals of the use of leaf vegetables, thus providing additional information on human/environment relationship. The presence of oral pathogens, even if at low abundance (<0.1%), can be related to the high consumption of carbohydrates and finds correspondence with the palaeopathological evidence. In conclusion, starting from very minute amounts of ancient dental calculus, our molecular and microscopic analysis jointly provided complementary data in support of past life condition reconstruction in ancient human populations. [ABSTRACT FROM AUTHOR]

In this study, we used metatranscriptomics for the first time to investigate microbial composition, functional signatures, and antimicrobial resistance gene expression in endodontic infections. Root canal samples were collected from ten teeth, including five primary and five persistent/secondary endodontic infections. RNA from endodontic samples was extracted, and RNA sequencing was performed on a NovaSeq6000 system (Illumina). Taxonomic analysis was performed using the Kraken2 bacterial database. Then, sequences with a taxonomic classification were annotated against the Universal Protein Knowledgebase for functional annotation and the Comprehensive Antibiotic Resistance Database for AR-like gene identification. Proteobacteria, Bacteroidetes, Firmicutes, and Actinobacteria represented the dominant phyla, whereas Fusobacteria, Spirochetes, and Synergistetes were among the nondominant phyla. The top ten species were mainly represented by obligate (or quasiobligate) anaerobes, including Gram-negative (eg, Capnocytophaga sp. oral taxon 323, Fusobacterium nucleatum , Prevotella intermedia , Prevotella oris , Tannerella forsythia, and Tannerella sp. oral taxon HOT−286) and Gram-positive species (eg, Olsenella uli and Parvimonas micra). Transcripts encoding moonlighting proteins (eg, glycolytic proteins, translational elongation factors, chaperonin, and heat shock proteins) were highly expressed, potentially affecting bacterial adhesion, biofilm formation, host defense evasion, and inflammation induction. Endodontic bacteria expressed genes conferring resistance to antibiotic classes commonly used in dentistry, with a high prevalence and expression of tetracycline and lincosamide resistance genes. Antibiotic efflux and antibiotic target alteration/protection were the main resistance mechanisms. Metatranscriptomics revealed the activity of potential endodontic pathogens, which expressed putative virulence factors and a wide diversity of genes potentially involved in AR. [ABSTRACT FROM AUTHOR]