A 5500-year-old Treponema pallidum genome from Sabana de Bogotá, Colombia.

Treponematosis, a bacterial infection caused by Treponema pallidum subspecies and T. carateum (yaws, bejel, syphilis, pinta), has afflicted humans for millennia. Despite paleopathological evidence and emerging genomic data, little is known about the evolutionary history of these pathogens. We report a 5500-year-old Treponema genome (TE1-3) from Middle Holocene hunter-gatherer contexts of the rock shelter Tequendama I in Colombia. Our analyses place TE1-3 as a sister lineage to all known T. pallidum subspecies, positioning this pathogen in the Americas millennia before European contact and before diversification of the subspecies causing syphilis, yaws, and bejel. This discovery broadens the known diversity of T. pallidum while extending the genomic record of treponemal pathogens by millennia, providing molecular support for a deep history of T. pallidum in the Americas. Editor's summary: Treponemes are infectious bacteria that cause the human diseases yaws, bejel, syphilis, and pinta. Despite skeletal pathology consistent with treponematosis being present in human remains for millennia, the origin and evolution of these bacteria remain opaque. Bozzi et al. identified a 5500-year-old Treponema genome during sequencing of human remains from Colombia without skeletal signs of treponematosis (see the Perspective by Zuckerman and Bailey). This genome falls outside of known T. pallidum lineages today, but it has many genetic hallmarks associated with virulence in modern pathogens of these subspecies. This work provides genetic evidence of treponematosis in the Americas thousands of years before the arrival of Europeans and shows the value of screening metagenomic sequences for ancient pathogens. —Corinne Simonti INTRODUCTION: The origins and early diversification of Treponema pallidum subspecies, which cause syphilis, yaws, and bejel, remain poorly resolved despite paleopathological evidence for treponemal disease in pre-Columbian populations. Sparse genomic data from ancient, precolonial contexts create a multimillennial disconnect between osteological evidence of treponematosis and its molecular record. RATIONALE: To better resolve the evolutionary history of T. pallidum, we performed genomic reconstruction and analysis of pathogen DNA detected in shotgun-sequenced DNA data for a human population genomics project. Although the skeleton showed no macroscopic signs of treponematosis, our metagenomic screening identified T. pallidum in an individual who died ~5500 years ago [5464 to 5309 calendar years before the present (cal yr B.P.), 2σ] and was excavated from Middle Holocene contexts of the Tequendama I rock shelter in Sabana de Bogotá, Colombia. We applied multiple methods (competitive mapping, phylogenetics reconstructions, genome-wide average nucleotide identity statistics, and patristic distance) to confidently identify this pathogen as T. pallidum, reconstruct the genome (TE1-3), and evaluate its relationship to other genomically characterized Treponema. With this genome, we inferred divergence estimates of TE1-3 from other T. pallidum to reconstruct the early diversification of T. pallidum subspecies. RESULTS: Our analyses place TE1-3 as an early-diverging sister lineage to all modern T. pallidum subspecies, unveiling a previously unknown subspecies. Bayesian molecular clock analyses estimate the divergence between TE1-3 and other T. pallidum lineages to ~13,700 years ago [95% highest posterior density (HPD): 6768 to 20,592 cal yr B.P.]. This suggests that the divergence between TE1-3 and the modern, genomically characterized T. pallidum subspecies occurred during the Late Pleistocene to Early Holocene, closely following the peopling of the Americas, whereas the diversification of the known subspecies themselves took place more recently, within the Holocene ~6000 cal yr B.P. (3622 to 9452 cal yr B.P. 95% HPD). After assessing the breadth of coverage, we determined that TE1-3 likely possessed virulence-associated genes found in modern T. pallidum strains, suggesting similar genetic potential for virulence. CONCLUSION: Our results show the presence and previously unknown diversity of T. pallidum from a Middle Holocene hunter-gatherer context in Sabana de Bogotá, Colombia, extending the genomic record of treponematoses by more than three millennia. TE1-3's early divergence supports models in which treponemal pathogens had already diversified in the Americas during the Holocene. The availability of this ancient South American genome provides a rare calibration point for reconstructing the early diversification of T. pallidum subspecies. This study also underscores the value of metagenomic screening of ancient remains to uncover previously undetectable infectious disease histories in deep time. These findings open new questions about the timing, routes, and drivers of treponemal spread, and on the longstanding interplay among Treponema pallidum, human hosts, and the broader socioecological landscapes in which these diseases have evolved, persisted, and spread to affect human populations. Deeply divergent Treponema pallidum lineage in the Americas and implications for pathogen evolution.: We identified and reconstructed a ~5500-year-old 1.7× T. pallidum genome (TE1-3) from a Middle Holocene individual at Tequendama I rock shelter in Colombia. Phylogenomic analysis reveals that TE1-3 is an early-branching sister lineage to all extant T. pallidum subspecies, with divergence dating to ~13,700 years ago (95% HPD: 6768 to 20,592 cal yr B.P.) and coinciding with early human migration and ecological shifts in the Americas. Despite its antiquity, TE1-3 possesses the T. pallidum virulence-associated genes, suggesting conserved pathogenic capacity. TE1-3 provides the earliest molecular evidence of T. pallidum in the Americas to date, bridging a multimillennial gap between skeletal signs and genomic data and offering a rare glimpse into the long-term evolution of treponemal pathogens. [Figure created in BioRender] [ABSTRACT FROM AUTHOR]