Human placental trophoblasts support sustained Treponema pallidum replication and reveal new candidate host pathways implicated in congenital syphilis

Abstract Congenital syphilis is a leading cause of preventable stillbirth, yet the mechanisms that enable Treponema pallidum subsp. pallidum, the etiological agent of syphilis, to traverse and persist within the human placenta are virtually unknown. This knowledge gap reflects, in part, the fastidious nature of T. pallidum , which has historically only been propagated in rabbit epithelial cells. Here, we demonstrate that T. pallidum replicates and can be propagated long-term in human placental trophoblast models. We identified three human trophoblast cell lines, JEG-3, BeWo, and HTR-8, that sustained robust T. pallidum replication for over 55 days. Confocal imaging with GFP-expressing T. pallidum demonstrated bacterial adherence to trophoblast cultures. To identify human placental pathways that may support T. pallidum replication and persistence, we performed bulk transcriptomic profiling of the three trophoblast lines co-cultured with T. pallidum for 3 or 7 days. Our findings revealed conserved host responses involving increased cholesterol synthesis, suppressed type I interferon signaling, and disrupted extracellular matrix organization. These data implicate host metabolic rewiring, innate immune attenuation, and extracellular matrix remodeling as candidate pathways that may promote T. pallidum invasion, replication and persistence within the placenta. Together, these models represent the first human placental systems capable of supporting efficient, long-term T. pallidum growth and provide a foundation for mechanistic studies of congenital syphilis pathogenesis.