Zander Johannes, Besier Silke, Saum Stephan H, Dehghani Faramarz, Loitsch Stefan, Brade Volker, Wichelhaus Thomas A
Institute of Medical Microbiology and Infection Control, Hospital of Johann Wolfgang Goethe University, Paul-Ehrlich-Str. 40, 60596 Frankfurt/Main, Germany.
Infect Immun. 2008 Apr;76(4):1333-9. doi: 10.1128/IAI.01075-07. Epub 2007 Dec 26.
Thymidine-dependent small-colony variants (SCVs) of Staphylococcus aureus are frequently associated with persistent and recurrent infections in cystic fibrosis patients. The phenotypic appearance of S. aureus SCVs or normal-colony variants (NCVs) is postulated to be affected by the intracellular amount of dTMP. This hypothesis was proven by metabolic pathway assays revealing altered intracellular dTMP concentrations, followed by investigation of the associated phenotype. Inhibition of the staphylococcal thymidylate synthase, which generated intracellular dTMP from dUMP, using 5-fluorouracil and co-trimoxazole resulted in an SCV phenotype. Inhibition of a nucleoside transporter, which provided the bacterial cell with extracellular thymidine, caused growth inhibition of SCVs. In turn, reversion of SCVs to NCVs was achieved by supplying extracellular dTMP. High-performance liquid chromatography additionally confirmed the intracellular lack of dTMP in SCVs, in contrast to NCVs. Moreover, the dTMP concentration is postulated to influence the intracellular persistence of S. aureus. Cell culture experiments with cystic fibrosis cells revealed that clinical and co-trimoxazole-induced SCVs with a diminished amount of dTMP showed significantly better intracellular persistence than NCVs. In conclusion, these results show that the dTMP concentration plays a key role in both the phenotypic appearance and the intracellular persistence of S. aureus.
金黄色葡萄球菌的胸腺嘧啶核苷依赖性小菌落变体（SCV）常与囊性纤维化患者的持续性和复发性感染相关。据推测，金黄色葡萄球菌SCV或正常菌落变体（NCV）的表型外观受细胞内dTMP含量的影响。代谢途径分析揭示细胞内dTMP浓度改变，随后对相关表型进行研究，从而证实了这一假设。使用5-氟尿嘧啶和复方新诺明抑制从dUMP生成细胞内dTMP的葡萄球菌胸苷酸合成酶，会导致SCV表型。抑制为细菌细胞提供细胞外胸腺嘧啶核苷的核苷转运体，会导致SCV生长受抑制。反过来，通过提供细胞外dTMP可使SCV逆转为NCV。高效液相色谱进一步证实，与NCV相比，SCV细胞内缺乏dTMP。此外，据推测dTMP浓度会影响金黄色葡萄球菌在细胞内的存活。对囊性纤维化细胞进行的细胞培养实验表明，临床分离株和复方新诺明诱导产生的dTMP含量减少的SCV在细胞内的存活能力明显优于NCV。总之，这些结果表明，dTMP浓度在金黄色葡萄球菌的表型外观和细胞内存活方面均起关键作用。
