Treponema pallidum subsp. pallidum is the causative agent of syphilis, a human-specific sexually transmitted infection that causes a multistage disease with diverse clinical manifestations. Treponema pallidum undergoes rapid vascular dissemination to penetrate tissue, placental, and blood-brain barriers and gain access to distant tissue sites. The rapidity and extent of T. pallidum dissemination are well documented, but the molecular mechanisms have yet to be fully elucidated. One protein that has been shown to play a role in treponemal dissemination is Tp0751, a T. pallidum adhesin that interacts with host components found within the vasculature and mediates bacterial adherence to endothelial cells under shear flow conditions. In this study, we further explore the molecular interactions of Tp0751-mediated adhesion to the vascular endothelium. We demonstrate that recombinant Tp0751 adheres to human endothelial cells of macrovascular and microvascular origin, including a cerebral brain microvascular endothelial cell line. Adhesion assays using recombinant Tp0751 N-terminal truncations reveal that endothelial binding is localized to the lipocalin fold-containing domain of the protein. We also confirm this interaction using live T. pallidum and show that spirochete attachment to endothelial monolayers is disrupted by Tp0751-specific antiserum. Further, we identify the 67-kDa laminin receptor (LamR) as an endothelial receptor for Tp0751 using affinity chromatography, coimmunoprecipitation, and plate-based binding methodologies. Notably, LamR has been identified as a receptor for adhesion of other neurotropic invasive bacterial pathogens to brain endothelial cells, including Neisseria meningitidis, Haemophilus influenzae, and Streptococcus pneumoniae, suggesting the existence of a common mechanism for extravasation of invasive extracellular bacterial pathogens. IMPORTANCE Syphilis is a sexually transmitted infection caused by the spirochete bacterium Treponema pallidum subsp. pallidum. The continued incidence of syphilis demonstrates that screening and treatment strategies are not sufficient to curb this infectious disease, and there is currently no vaccine available. Herein we demonstrate that the T. pallidum adhesin Tp0751 interacts with endothelial cells that line the lumen of human blood vessels through the 67-kDa laminin receptor (LamR). Importantly, LamR is also a receptor for meningitis-causing neuroinvasive bacterial pathogens such as Neisseria meningitidis, Haemophilus influenzae, and Streptococcus pneumoniae. Our findings enhance understanding of the Tp0751 adhesin and present the intriguing possibility that the molecular events of Tp0751-mediated treponemal dissemination may mimic the endothelial interaction strategies of other invasive pathogens.

Extracellular bacteria that spread via the vasculature employ invasive mechanisms that mirror those of metastatic tumor cells, including intravasation into the bloodstream and survival during hematogenous dissemination, arrestation despite blood flow, and extravasation into distant tissue sites. Several invasive bacteria have been shown to exploit normal platelet function during infection. Due to their inherent ability to interact with and influence other cell types, platelets play a critical role in alteration of endothelial barrier permeability, and their role in cancer metastasis has been well established. The highly invasive bacterium and causative agent of syphilis, Treponema pallidum subspecies pallidum, readily crosses the endothelial, blood-brain and placental barriers. However, the mechanisms underlying this unusual and important aspect of T. pallidum pathogenesis are incompletely understood. In this study we use darkfield microscopy in combination with flow cytometry to establish that T. pallidum interacts with platelets. We also investigate the dynamics of this interaction and show T. pallidum is able to activate platelets and preferentially interacts with activated platelets. Platelet-interacting treponemes consistently exhibit altered kinematic (movement) parameters compared to free treponemes, and T. pallidum-platelet interactions are reversible. This study provides insight into host cell interactions at play during T. pallidum infection and suggests that T. pallidum may exploit platelet function to aid in establishment of disseminated infection.

that had an established Ng infection. In this regard, a single dose of carbamazepine was effective in curing (!99% Ng killing by 24h post-treatment) Pex cells infected with multidrugresistant Ng strains, including the ceftriaxone-resistant strains WHO-X (H041) and WHO-Y (F89). Conclusion Our data identify safe, repurposed, drugs that may have efficacy in preventing and treating Ng cervical infection in women. Disclosure No significant relationships.

Introduction Treponema pallidum ssp. pallidum, the causative agent of syphilis, is a highly invasive pathogen that interacts with a diverse repertoire of host cells during infection. The pathogen invades immunologically privileged sites and crosses the placental, blood-brain, endothelial and blood-retina barriers to establish widespread infection. Treponema pallidum disseminates via the circulatory and lymphatic systems, avoiding the prevalent inflammatory reactions raised against other blood-borne pathogens. In this study we investigate if T. pallidum uses an interaction with human platelets, key mediators of homeostasis and immune surveillance, to facilitate host persistence. We demonstrate that T. pallidum adheres to human platelets enabling survival for an extended period, and we discuss how this interaction may aid T. pallidum pathogenesis. MethodsPlatelet rich plasma prepared from donor blood was incubated under host-mimicking microaerophilic conditions with viable T. pallidum, followed by examination for T. pallidum-platelet interactions via darkfield microscopy and flow cytometry analyses. Viability was confirmed using microscopic and fluorescent staining methodologies.Results Treponema pallidum binds both the rounded and spread morphologies of activated platelets via a polar tip structure, maintaining a firm tether under fluidic conditions. A lack of interaction between heat-killed T. pallidum and platelets confirmed specificity and identified heat-labile T. pallidum surface components as mediators of this interaction. Viability assays illustrated T. pallidum retained viability in platelet rich plasma for >3 days under these conditions.ConclusionThe demonstration in this study of (1) prolonged T. pallidum survival within human platelet rich plasma and (2) T. pallidum-platelet interactions indicates that platelets do not exhibit a direct antimicrobial effect on T. pallidum and that T. pallidum mediates a strong and specific interaction with human platelets. These findings may reveal a novel mechanism of host survival employed by this elusive pathogen.

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