Molecular Subtyping of Treponema pallidum Subspecies pallidum

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Abstract
Background and Objectives: Epidemiologic studies on syphilis have been hampered by the fact that strains ofTreponema pallidumsubspeciespallidum(T. pallidum), the causative agent of this disease, cannot be differentiated by either protein-based or deoxyribonucleic acid-based methods. Syphilis is endemic in many developing countries and is common in some industrialized nations. In addition, the disease has been shown to increase the risk of infection with the human immunodeficiency virus. Goal: To develop a molecular subtyping method forT. pallidum. Study Design: Two genes exhibiting intrastrain variability were identified as potential targets for strain differentiation: the acidic repeat protein (arp) gene, which contains a variable number of 60 base pair repeats, and a member of the treponema pallidum repeat (tpr) gene family. Polymerase chain reaction amplification and restriction endonuclease digestion of polymerase chain reaction products from laboratory strains and clinical specimens were used to develop a molecular subtyping scheme forT. pallidum. Results: Determining the number of repeats in thearpgene by polymerase chain reaction resulted in 12 different subtypes among the 63 isolates that were studied. Among those, most(54.2%) hadarpgenes with 14 repeats. The other 11 subtypes hadarpgenes with 7 to 21 repeats, each accounting for 2% to 14% of the isolates. Polymerase chain reaction amplification of a member of thetprgene family from a subset of 46 isolates followed by digestion of the polymerase chain reaction product withMseI resulted in seven restriction fragment length polymorphism patterns designatedatog.Strains with 14 repeats could be grouped into five restriction fragment length polymorphism subtypes. By combining the two systems we observed 16 subtypes among 46 isolates examined. This typing system is stable, reproducible, and easy to perform. In addition, the use of the ABI Genetic Analyzer for the determination of fragment size and banding patterns makes the results unbiased. Conclusion: This is the first molecular subtyping system that distinguishes among clinical isolates ofT. pallidum.