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Abstract：
BACKGROUND: Long-term treatment with trimethoprim-sulfamethoxazole (SXT) can lead to the formation of small-colony variants (SCVs) of Staphylococcus aureus. However, the mechanism behind SCVs formation remains poorly understood. In this study, we explored the phenotype and omics-based characterization of S. aureus SCVs induced by SXT and shed light on the potential causes of SCV formation.
METHODS: Stable SCVs were obtained by continuously treating S. aureus isolates using 12/238 µg/ml of SXT, characterized by growth kinetics, antibiotic susceptibility testing, and auxotrophism test. Subsequently, a pair of representative strains (SCV and its parental strain) were selected for genomic, transcriptomic and metabolomic analysis.
RESULTS: Three stable S. aureus SCVs were successfully screened and proven to be homologous to their corresponding parental strains. Phenotypic tests showed that all SCVs were non-classical mechanisms associated with impaired utilization of menadione, heme and thymine, and exhibited slower growth and higher antibiotic minimum inhibitory concentrations (MICs), compared to their corresponding parental strains. Genomic data revealed 15 missense mutations in 13 genes in the representative SCV, which were involved in adhesion, intramolecular phosphate transfer on ribose, transport pathways, and phage-encoded proteins. The combination analysis of transcriptome and metabolome identified 35 overlapping pathways possible associated with the phenotype switching of S. aureus. These pathways mainly included changes in metabolism, such as purine metabolism, pyruvate metabolism, amino acid metabolism, and ABC transporters, which could play a crucial role in promoting SCVs development by affecting nucleic acid synthesis and energy metabolism in bacteria.
CONCLUSIONS: This study provides profound insights into the causes of S. aureus SCV formation induced by SXT. The findings may offer valuable clues for developing new strategies to combat S. aureus SCV infections.
METHODS: Stable SCVs were obtained by continuously treating S. aureus isolates using 12/238 µg/ml of SXT, characterized by growth kinetics, antibiotic susceptibility testing, and auxotrophism test. Subsequently, a pair of representative strains (SCV and its parental strain) were selected for genomic, transcriptomic and metabolomic analysis.
RESULTS: Three stable S. aureus SCVs were successfully screened and proven to be homologous to their corresponding parental strains. Phenotypic tests showed that all SCVs were non-classical mechanisms associated with impaired utilization of menadione, heme and thymine, and exhibited slower growth and higher antibiotic minimum inhibitory concentrations (MICs), compared to their corresponding parental strains. Genomic data revealed 15 missense mutations in 13 genes in the representative SCV, which were involved in adhesion, intramolecular phosphate transfer on ribose, transport pathways, and phage-encoded proteins. The combination analysis of transcriptome and metabolome identified 35 overlapping pathways possible associated with the phenotype switching of S. aureus. These pathways mainly included changes in metabolism, such as purine metabolism, pyruvate metabolism, amino acid metabolism, and ABC transporters, which could play a crucial role in promoting SCVs development by affecting nucleic acid synthesis and energy metabolism in bacteria.
CONCLUSIONS: This study provides profound insights into the causes of S. aureus SCV formation induced by SXT. The findings may offer valuable clues for developing new strategies to combat S. aureus SCV infections.
摘要：
背景：使用甲氧苄啶-磺胺甲恶唑（SXT）的长期治疗可导致金黄色葡萄球菌的小菌落变体（SCV）的形成。然而,SCV形成背后的机制仍然知之甚少。在这项研究中,我们探索了SXT诱导的金黄色葡萄球菌SCV的表型和基于组学的表征,并阐明了SCV形成的潜在原因.
方法：通过使用12/238µg/ml的SXT连续处理金黄色葡萄球菌分离株，获得稳定的SCV，以生长动力学为特征，抗生素药敏试验,和营养缺陷测试。随后，选择一对代表性菌株（SCV及其亲本菌株）进行基因组，转录组学和代谢组学分析。
结果:成功筛选了三个稳定的金黄色葡萄球菌SCV,并证明其与其相应的亲本菌株同源。表型测试表明，所有SCV都是与甲萘醌利用受损相关的非经典机制，血红素和胸腺嘧啶,并表现出较慢的生长和较高的抗生素最低抑制浓度（MIC），与它们相应的亲本菌株相比。基因组数据显示有代表性的SCV中13个基因中有15个错义突变，参与粘附，分子内磷酸在核糖上的转移，运输途径，和噬菌体编码的蛋白质。转录组和代谢组的组合分析鉴定了可能与金黄色葡萄球菌的表型转换相关的35个重叠途径。这些途径主要包括代谢的变化，比如嘌呤代谢，丙酮酸代谢，氨基酸代谢，和ABC运输机，通过影响细菌的核酸合成和能量代谢,对促进SCV的发育起到至关重要的作用。
结论：这项研究为SXT诱导的金黄色葡萄球菌SCV形成的原因提供了深刻的见解。这些发现可能为开发对抗金黄色葡萄球菌SCV感染的新策略提供有价值的线索。
方法：通过使用12/238µg/ml的SXT连续处理金黄色葡萄球菌分离株，获得稳定的SCV，以生长动力学为特征，抗生素药敏试验,和营养缺陷测试。随后，选择一对代表性菌株（SCV及其亲本菌株）进行基因组，转录组学和代谢组学分析。
结果:成功筛选了三个稳定的金黄色葡萄球菌SCV,并证明其与其相应的亲本菌株同源。表型测试表明，所有SCV都是与甲萘醌利用受损相关的非经典机制，血红素和胸腺嘧啶,并表现出较慢的生长和较高的抗生素最低抑制浓度（MIC），与它们相应的亲本菌株相比。基因组数据显示有代表性的SCV中13个基因中有15个错义突变，参与粘附，分子内磷酸在核糖上的转移，运输途径，和噬菌体编码的蛋白质。转录组和代谢组的组合分析鉴定了可能与金黄色葡萄球菌的表型转换相关的35个重叠途径。这些途径主要包括代谢的变化，比如嘌呤代谢，丙酮酸代谢，氨基酸代谢，和ABC运输机，通过影响细菌的核酸合成和能量代谢,对促进SCV的发育起到至关重要的作用。
结论：这项研究为SXT诱导的金黄色葡萄球菌SCV形成的原因提供了深刻的见解。这些发现可能为开发对抗金黄色葡萄球菌SCV感染的新策略提供有价值的线索。
