Aim: To describe the molecular types of Treponema pallidum and the proportion of macrolide and tetracycline resistance mutations in Barcelona. Materials & methods: Molecular type was determined using the Enhanced-CDC Typing system and antibiotic resistance was determined by sequencing the 23S and 16S rRNA genes. Results: A total of 183 patients were enrolled and 213 specimens (99 ulcers, 114 bloods) were collected. Sixty-two (70.5%) of 88 ulcers and 0 (0%) of bloods T. pallidum-DNA containing samples were fully typed. Up to 21 different strain types were identified (14d/g in 27.4%; 14f/g in 14.5%). Macrolide resistance mutations were present in 95% and tetracycline in 0%. Conclusion: Several different strains co-exist in Barcelona with a high proportion of macrolide resistance and absence of tetracycline resistance.