Interfaculty Institute for Genetics and Functional Genomics, University Medicine Greifswald Greifswald, Germany.
ZIK-FunGene Junior Research Group Applied Proteomics, Interfaculty Institute for Genetics and Functional Genomics, University Medicine Greifswald Greifswald, Germany.
Front Microbiol. 2014 Aug 1;5:392. doi: 10.3389/fmicb.2014.00392. eCollection 2014.
Staphylococcus aureus is a human pathogen that can cause a wide range of diseases. Although formerly regarded as extracellular pathogen, it has been shown that S. aureus can also be internalized by host cells and persist within these cells. In the present study, we comparatively analyzed survival and physiological adaptation of S. aureus HG001 after internalization by two human lung epithelial cell lines (S9 and A549), and human embryonic kidney cells (HEK 293). Combining enrichment of bacteria from host-pathogen assays by cell sorting and quantitation of the pathogen's proteome by mass spectrometry we characterized S. aureus adaptation during the initial phase between 2.5 h and 6.5 h post-infection. Starting with about 2 × 10(6) bacteria, roughly 1450 S. aureus proteins, including virulence factors and metabolic enzymes were identified by spectral comparison and classical database searches. Most of the bacterial adaptation reactions, such as decreased levels of ribosomal proteins and metabolic enzymes or increased amounts of proteins involved in arginine and lysine biosynthesis, enzymes coding for terminal oxidases and stress responsive proteins or activation of the sigma factor SigB were observed after internalization into any of the three cell lines studied. However, differences were noted in central carbon metabolism including regulation of fermentation and threonine degradation. Since these differences coincided with different intracellular growth behavior, complementary profiling of the metabolome of the different non-infected host cell types was performed. This revealed similar levels of intracellular glucose but host cell specific differences in the amounts of amino acids such as glycine, threonine or glutamate. With this comparative study we provide an impression of the common and specific features of the adaptation of S. aureus HG001 to specific host cell environments as a starting point for follow-up studies with different strain isolates and regulatory mutants.
金黄色葡萄球菌是一种人类病原体，可以引起广泛的疾病。尽管以前被认为是细胞外病原体，但已表明金黄色葡萄球菌也可以被宿主细胞内化并在这些细胞内持续存在。在本研究中，我们比较分析了金黄色葡萄球菌 HG001 被两种人肺上皮细胞系（S9 和 A549）和人胚肾细胞（HEK 293）内化后的存活和生理适应。通过细胞分选从宿主-病原体测定中富集细菌并通过质谱法定量病原体的蛋白质组，我们在感染后 2.5 小时至 6.5 小时的初始阶段表征了金黄色葡萄球菌的适应。从大约 2×10^6 个细菌开始，通过光谱比较和经典数据库搜索鉴定了约 1450 种金黄色葡萄球菌蛋白，包括毒力因子和代谢酶。大多数细菌适应反应，如核糖体蛋白和代谢酶水平降低，或参与精氨酸和赖氨酸生物合成、编码末端氧化酶和应激响应蛋白的酶或 sigma 因子 SigB 的激活，在被内化到研究的三种细胞系中的任何一种后都观察到。然而，在中心碳代谢中观察到差异，包括发酵和苏氨酸降解的调节。由于这些差异与不同的细胞内生长行为相吻合，因此对不同未感染宿主细胞类型的代谢组进行了补充分析。这揭示了细胞内葡萄糖的相似水平，但氨基酸（如甘氨酸、苏氨酸或谷氨酸）的量存在宿主细胞特异性差异。通过这项比较研究，我们对金黄色葡萄球菌 HG001 适应特定宿主细胞环境的常见和特定特征有了初步了解，为后续使用不同分离株和调节突变体进行研究提供了起点。
