Compositions and Methods for Long-Term in Vitro Culture Of The Syphilis Spirochete.

Background

Syphilis is a multistage sexually transmitted infection of world-wide importance, with an estimated 5.6 million new cases per year. Treponema pallidum, the causative agent of syphilis, was first identified by Schaudinn and Hoffman in 1905, but in the past researchers have been unable to maintain T. pallidum viability or multiplication beyond 12 to 18 days. The inability to culture these organisms continuously in vitro has necessitated their propagation in rabbits for use in research and the production of antigens for serodiagnostic tests.

Discovery

UTHealth researchers have developed a culture system comprising of a co-culture of a population of T. pallidum along with mammalian epithelial cells.  This system utilizes a customized medium and incubation in low concentrations of oxygen,  with subculture at 6-7 day intervals.  Multiple strains of T. pallidum were tested with successful outcomes of incubation times of greater than 47 days for UW249B strain, 70+ days for UW231B strain, and currently over one year for the Nichols strain. Structural integrity and viability of strains were tested and confirmed using multiple forms of microscopy, quantitative PCR, and infectivity studies.

Benefits/Advantages of New Cell Culture Method

•       Elimination of costly maintenance of T. pallidum by rabbit infection

•       Yields of up to 9 x 10⁸ T. pallidum per 75 cm²  tissue culture flask

•       Full retention of infectivity and structural integrity of T. pallidum

Potential Cell Culture Applications

•       production of T. pallidum for use in syphilis serodiagnostic tests

•       genome sequencing of in vitro-cultured T. pallidum

•       antimicrobial susceptibility testing

•       potential use for isolation and characterization of new strains from syphilis, yaws, and bejel (endemic syphilis) patients.

Inventors