Influence of dTMP on the phenotypic appearance and intracellular persistence of Staphylococcus aureus

Area of Science:

• Microbiology
• Molecular Biology
• Infectious Diseases

Background:

• Thymidine-dependent small-colony variants (SCVs) of Staphylococcus aureus are implicated in persistent and recurrent infections, particularly in cystic fibrosis patients.
• The intracellular concentration of thymidylate (dTMP) is hypothesized to influence the phenotypic expression and persistence of S. aureus SCVs.

Purpose of the Study:

• To investigate the role of intracellular dTMP concentration in the development and persistence of S. aureus SCVs.
• To explore therapeutic strategies targeting dTMP metabolism for managing S. aureus infections.

Main Methods:

• Metabolic pathway assays were used to measure intracellular dTMP concentrations.
• Inhibition of thymidylate synthase and nucleoside transporters was employed to induce or affect SCV phenotypes.
• High-performance liquid chromatography (HPLC) was used to quantify dTMP levels.
• Cell culture experiments with cystic fibrosis cells assessed bacterial intracellular persistence.

Main Results:

• Inhibition of thymidylate synthase with 5-fluorouracil and co-trimoxazole induced an SCV phenotype.
• Inhibition of nucleoside transporters impaired SCV growth, while extracellular dTMP supplementation reverted SCVs to normal-colony variants (NCVs).
• HPLC confirmed lower intracellular dTMP in SCVs compared to NCVs.
• SCVs with diminished dTMP exhibited significantly enhanced intracellular persistence in cystic fibrosis cell cultures.

Conclusions:

• Intracellular dTMP concentration is a critical factor determining both the phenotypic appearance (SCV vs. NCV) of Staphylococcus aureus.
• dTMP levels also play a key role in the intracellular persistence of S. aureus, suggesting potential therapeutic targets.