The species Treponema pallidum includes three subspecies (pallidum, pertenue, and endemicum) that cause syphilis, yaws, and bejel, respectively. A closely related species, Treponema paraluiscuniculi, is the etiologic agent of venereal syphilis in rabbits but does not infect humans. Although these treponemes cause distinct diseases, no laboratory method for differentiation has been reported. Genetic signatures were defined in the 5' and 3' flanking regions of the 15-kDa lipoprotein gene (tpp15) that distinguish the human pathogens and T. paraluiscuniculi, as well as distinguishing T. pallidum subsp. pallidum from the causes of human nonvenereal treponematoses. A single Eco47III restriction site in the 5' flanking region differentiates T. pallidum subsp. pallidum from the other subspecies and species, and an XcmI site in the 3' flanking region differentiates T. paraluiscuniculi from the human pathogens. Polymerase chain reaction methods and restriction polymorphism were used to analyze 27 strains of pathogenic Treponema species.

The 15-kDa lipoprotein of Treponema pallidum is a major immunogen during natural syphilis infection in humans and experimental infection in other hosts. The humoral and cellular immune responses to this molecule appear late in infection as resistance to reinfection is developing. One therefore might hypothesize that this antigen is important for protective immunity. This possibility is explored by using both genetic and antigenic approaches. Limited or no cross-protection has been demonstrated between the T. pallidum subspecies and strains or between Treponema species. We therefore hypothesized that if the 15-kDa antigen was of major importance in protective immunity, it might be a likely site of antigenic diversity. To explore this possibility, the sequences of the open reading frames of the 15-kDa gene have been determined for Treponema pallidum subsp. pallidum (Nichols and Bal-3 strains), T. pallidum subsp. pertenue (Gauthier strain), T. pallidum subsp. endemicum (Bosnia strain), Treponema paraluiscuniculi (Cuniculi A, H, and K strains), and a littlecharacterized simian isolate of Treponema sp. (Fribourg-Blanc strain). No significant differences in DNA sequences of the genes for the coding region of the 15-kDa antigen were found among the different species and subspecies studied. In addition, all organisms showed expression of the 15-kDa antigen as determined by monoclonal antibody staining. The role of the 15-kDa antigen in protection against homologous infection with T. pallidum subsp. pallidum Nichols was examined in rabbits immunized with a purified recombinant 15-kDa fusion protein. No alteration in chancre development was observed in immunized, compared to unimmunized, rabbits, and the antisera induced by the immunization failed to enhance phagocytosis of T. pallidum subsp. pallidum by macrophages in vitro. These results do not support a major role for this antigen in protection against syphilis infection.

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