Exploring differentially expressed genes of staphylococcus aureus exposed to human tonsillar cells using RNA sequencing

Nasal colonization is well described; however, we have limited knowledge about S. aureus throat colonization. The main objective of this project was to explore differentially expressed genes (DEGs) in S. aureus throat isolate TR145 exposed for 1 or 3 hours (h) to human tonsil epithelial cells (HTEpiC) by using RNA sequencing (RNA-seq) and pathway analysis. We have shown the suitability of using HTEpiC as an in vitro model for investigating key determinants in S. aureus during co-incubation with the HTEpiC cells. Among the DEGs were genes encoding proteins involved in adhesion and immune evasion, as well as iron acquisition and transport. As their expression is induced upon meeting with the HTEpiC, they might be explored further for future targets for intervention to prevent either colonization or infection in the throat region. In this study, a Staphylococcus aureus throat isolate was cultured with or without a tonsillar cell line prior to RNA sequencing to find differentially expressed genes (DEGs). Test samples were defined as S. aureus strains exposed to HTEpiC and control samples were defined as S. aureus without HTEpiC cells. Three independent experiments were run in triplicates. Total RNA extracted from three replicates of S. aureus TR145 grown in absence of HTEpiC cells collected at time point of 0h, 1h and 3h (bacteriaonly, control samples (C)) and three replicas of S. aureus TR145 after 1h and 3h exposure to HTEpiC cells (bacteriaVSHTEpiC, test samples (T)) were selected for RNA-seq library preparation

鼻腔定植现象已有充分研究阐释，但目前我们对金黄色葡萄球菌（Staphylococcus aureus，简称S. aureus）咽部定植的认知仍较为有限。本研究的核心目标为，通过RNA测序（RNA-seq）及通路分析，探究暴露于人扁桃体上皮细胞（human tonsil epithelial cells，HTEpiC）1小时或3小时的金黄色葡萄球菌咽部分离株TR145的差异表达基因（differentially expressed genes，DEGs）。本研究证实，HTEpiC可作为体外模型，用于探究金黄色葡萄球菌与HTEpiC共孵育过程中的关键致病决定因子。本次筛选得到的差异表达基因中，包含编码黏附、免疫逃逸相关蛋白以及铁摄取与转运相关蛋白的基因。由于这些基因在与HTEpiC接触后表达被诱导上调，因此可作为潜在干预靶点，用于未来预防咽部定植或感染的相关研究。本研究中，我们将金黄色葡萄球菌咽部分离株与扁桃体细胞系共培养或单独培养，随后开展RNA测序以筛选差异表达基因（DEGs）。实验组样本定义为暴露于HTEpiC的金黄色葡萄球菌菌株，对照组样本则为未接触HTEpiC的金黄色葡萄球菌菌株。本研究共开展3次独立实验，每组均设置3次重复。我们选取以下样本进行RNA-seq文库构建：未接触HTEpiC的金黄色葡萄球菌TR145在0h、1h、3h三个时间点的3次重复样本（纯细菌培养组，对照组C），以及暴露于HTEpiC 1h和3h的金黄色葡萄球菌TR145的3次重复样本（细菌-HTEpiC共培养组，实验组T）。