Long-Term <i>In Vitro</i> Culture of the Syphilis Spirochete <i>Treponema pallidum</i> subsp. <i>pallidum</i>

Abstract

Investigation of <i>Treponema pallidum</i> subsp. <i>pallidum</i>, the spirochete that causes syphilis, has been hindered by an inability to culture the organism continuously <i>in vitro</i> despite more than a century of effort. In this study, long-term logarithmic multiplication of <i>T. pallidum</i> was attained through subculture every 6 to 7 days and periodic feeding using a modified medium (<i>T. pallidum</i> culture medium 2 [TpCM-2]) with a previously described microaerobic, rabbit epithelial cell coincubation system. Currently, cultures have maintained continuous growth for over 6 months with full retention of viability as measured by motility and rabbit infectivity. This system has been applied successfully to the well-studied Nichols strain of <i>T. pallidum</i>, as well as to two recent syphilis isolates, UW231B and UW249B. Light microscopy and cryo-electron microscopy showed that <i>in vitro</i>-cultured <i>T. pallidum</i> retains wild-type morphology. Further refinement of this long-term subculture system is expected to facilitate study of the physiological, genetic, pathological, immunologic, and antimicrobial susceptibility properties of <i>T. pallidum</i> subsp. <i>pallidum</i> and closely related pathogenic <i>Treponema</i> species and subspecies.<b>IMPORTANCE</b> Syphilis, a sexually transmitted disease with a global distribution, is caused by a spiral-shaped bacterium called <i>Treponema pallidum</i> subspecies <i>pallidum</i> Previously, <i>T. pallidum</i> was one of the few major bacterial pathogens that had not been cultured long-term <i>in vitro</i> (in a test tube), greatly hindering efforts to better understand this organism and the disease that it causes. In this article, we report the successful long-term cultivation of <i>T. pallidum</i> in a tissue culture system, a finding that is likely to enhance our ability to obtain new information applicable to the diagnosis, treatment, and prevention of syphilis.