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Brucellosis is a disease caused by the bacterium Brucella and typically transmitted through contact with infected ruminants. It is one of the most common chronic zoonotic diseases and of particular interest to public health agencies. Despite its well-known transmission history and characteristic symptoms, we lack a more complete understanding of the evolutionary history of its best-known species-Brucella melitensis. To address this knowledge gap we fortuitously found, sequenced and assembled a high-quality ancient B. melitensis draft genome from the kidney stone of a 14th-century Italian friar. The ancient strain contained fewer core genes than modern B. melitensis isolates, carried a complete complement of virulence genes, and did not contain any indication of significant antimicrobial resistances. The ancient B. melitensis genome fell as a basal sister lineage to a subgroup of B. melitensis strains within the Western Mediterranean phylogenetic group, with a short branch length indicative of its earlier sampling time, along with a similar gene content. By calibrating the molecular clock we suggest that the speciation event between B. melitensis and B. abortus is contemporaneous with the estimated time frame for the domestication of both sheep and goats. These results confirm the existence of the Western Mediterranean clade as a separate group in the 14th CE and suggest that its divergence was due to human and ruminant co-migration., Competing Interests: The authors have declared that no competing interests exist., (Copyright: © 2023 Long et al. This is an open access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited.)

[This corrects the article DOI: 10.1371/journal.ppat.1006750.].

Hepatitis B virus (HBV) is a ubiquitous viral pathogen associated with large-scale morbidity and mortality in humans. However, there is considerable uncertainty over the time-scale of its origin and evolution. Initial shotgun data from a mid-16th century Italian child mummy, that was previously paleopathologically identified as having been infected with Variola virus (VARV, the agent of smallpox), showed no DNA reads for VARV yet did for hepatitis B virus (HBV). Previously, electron microscopy provided evidence for the presence of VARV in this sample, although similar analyses conducted here did not reveal any VARV particles. We attempted to enrich and sequence for both VARV and HBV DNA. Although we did not recover any reads identified as VARV, we were successful in reconstructing an HBV genome at 163.8X coverage. Strikingly, both the HBV sequence and that of the associated host mitochondrial DNA displayed a nearly identical cytosine deamination pattern near the termini of DNA fragments, characteristic of an ancient origin. In contrast, phylogenetic analyses revealed a close relationship between the putative ancient virus and contemporary HBV strains (of genotype D), at first suggesting contamination. In addressing this paradox we demonstrate that HBV evolution is characterized by a marked lack of temporal structure. This confounds attempts to use molecular clock-based methods to date the origin of this virus over the time-frame sampled so far, and means that phylogenetic measures alone cannot yet be used to determine HBV sequence authenticity. If genuine, this phylogenetic pattern indicates that the genotypes of HBV diversified long before the 16th century, and enables comparison of potential pathogenic similarities between modern and ancient HBV. These results have important implications for our understanding of the emergence and evolution of this common viral pathogen.