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. Overall design: 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

鼻腔定植已有充分研究报道，但目前我们对金黄色葡萄球菌（S. aureus）咽部定植的认知仍较为匮乏。本研究的核心目标为：通过RNA测序（RNA-seq）与通路分析，探究暴露于人扁桃体上皮细胞（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（DEGs）。本研究证实，采用HTEpiC作为体外模型，可有效用于探究金黄色葡萄球菌与HTEpiC共孵育过程中的关键调控因子。本次筛选得到的差异表达基因，涵盖了编码黏附、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接触后表达被显著诱导，后续可进一步探索其作为干预靶点的潜力，以预防咽部区域的细菌定植或感染。总体实验设计：本研究中，将一株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，随后通过RNA测序筛选差异表达基因。实验组定义为暴露于HTEpiC的金黄色葡萄球菌菌株，对照组定义为未与HTEpiC共培养的金黄色葡萄球菌。本研究开展了三次独立实验，每个实验均设置三次技术重复。本次选取用于构建RNA测序文库的样本包括：未添加HTEpiC、分别于0h、1h、3h收集的金黄色葡萄球菌TR145的三次重复样本（仅细菌培养，对照组C组），以及暴露于HTEpiC 1h、3h的金黄色葡萄球菌TR145的三次重复样本（细菌与HTEpiC共培养，实验组T组）。