Background: Since its discovery, penicillin has been the primary treatment option for syphilis, caused by the spirochete Treponema pallidum. Its efficacy against the pathogen is significantly greater compared to doxycycline, the only other viable treatment option thus far. Due to previous inability to culture T. pallidum in vitro, efforts to identify compounds that rival the efficacy of penicillin have been hindered. With up to 10% of the global population reporting penicillin allergies, global penicillin shortages, and growing concern about antibiotic resistance, alternative treatment options are needed. Methods: In this study a novel drug screen of almost 100 B-lactams was performed to determine their efficacy against T. pallidumin vitro. A multiphase, iterative approach of manual enumeration and qRT-PCR was used to identify a small subset of high preforming compounds. The top 10% of compounds were further evaluated to determine their in vitro minimum inhibitory concentration (MIC). Finally, we used a fluorescent D-amino acid to visualize peptidoglycan synthesis in drug treated and untreated controls to validate the efficacy of the antibiotics on a cellular level. Results: We identified multiple B-lactams with similar or lower in vitro MICs compared to benzathine penicillin G, the current standard of care. Of note, was nafcillin which had an in vitro MIC of 0.553 ng/mL and had several additional promising characteristics such as a long half-life, low cost:efficacy ratio, and its current use in clinical settings treating penicillin resistant bacteria. Additionally, we determined that T. pallidum incorporates peptidoglycan ubiquitously across the sacculus indicating a lateral mechanism of growth. All of the top preforming antibiotics caused a decrease in peptidoglycan remodeling and growth. Conclusion: This is the first major drug screen conducted for the syphilis agent and was successful in identifying several potential therapeutics for future clinical investigation. In addition, this work provides new insights into T. pallidum peptidoglycan biology.
