MOLECULAR SUBTYPING OF TREPONEMA-PALLIDUM SUBSPECIES PALLIDUM

Background and Objectives: Epidemiologic studies on syphilis have been hampered by the fact that strains of Treponema pallidum subspecies pa llidum (T, pallidum), the causative agent of this disease, cannot be d ifferentiated by either protein-based or deoxyribonucleic acid-based m ethods. Syphilis is endemic in many developing countries and is common in some industrialized nations, In addition, the disease has been sho wn to increase the risk of infection with the human immunodeficiency v irus. Goal: To develop a molecular subtyping method for T, pallidum. S tudy Design: Two genes exhibiting intrastrain variability were identif ied 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 fami ly. Polymerase chain reaction amplification and restriction endonuclea se digestion of polymerase chain reaction products from laboratory str ains and clinical specimens were used to develop a molecular subtyping scheme for T, pallidum. Results: Determining the number of repeats in the alp gene by polymerase chain reaction resulted in 12 different su btypes among the 63 isolates that were studied. Among those, most (54. 2%) had alp genes with 14 repeats. The other II subtypes had mp genes with 7 to 21 repeats, each accounting for 2% to 14% of the isolates. P olymerase chain reaction amplification of a member of the tpr gene fam ily from a subset of 46 isolates followed by digestion of the polymera se chain reaction product with MseI resulted in seven restriction frag ment length polymorphism patterns designated a to g, Strains with 14 r epeats could be grouped into five restriction fragment length polymorp hism subtypes, By combining the two systems we observed 16 subtypes am ong 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 sy stem that distinguishes among clinical isolates of T, pallidum.