目的 分析致角膜溃疡的金黄色葡萄球菌和正常人结膜囊分离的金黄色葡萄球菌的差异表达基因,探讨金黄色葡萄球菌性角膜溃疡可能的发病机制.方法 收集2018年1—8月在青岛眼科医院检验科分离培养的10株金黄色葡萄球菌,其中5株来自经培养证实的细菌性角膜溃疡患者;5株来自正常人结膜囊细菌培养.应用转录组测序技术(RNA-Seq)对10株金黄色葡萄球菌进行转录组测序,对获得的数据进行生物信息学分析,以P≤0.05且差异倍数≥2为条件筛选显著差异表达基因(DEGs),并利用基因本体(GO)以及京都基因和基因组百科全书(KEGG)数据库对DEGs进行功能注释和富集分析.结果 2个组间共发现270个DEGs,其中138个表达上调,132个表达下调.分析差异基因的功能发现,编码α溶血素、δ溶血素、毒力因子EsxA和LysR家族转录调节器的基因在角膜溃疡分离株中均显著上调.GO富集分析显示,DEGs的功能主要涉及与代谢相关的生物学功能,其中次黄嘌呤核苷酸的合成与代谢最显著.KEGG通路分析表明,多条代谢通路发生显著变化,其中金黄色葡萄球菌感染、双组分信号系统、丙酮酸代谢、嘌呤代谢等途径对探讨金黄色葡萄球菌性角膜溃疡的致病机制至关重要.结论 角膜溃疡患者分离得到的金黄色葡萄球菌和正常人结膜囊分离的金黄色葡萄球菌之间存在差异基因表达,为进一步研究金黄色葡萄球菌性角膜溃疡的致病机制和治疗靶点相关的基因或通路提供了实验基础.%Objective To investigate the possible pathogenesis of Staphylococcus aureus ( S. aureus) corneal ulcer by analyzing the differentially expressed genes ( DEGs) of S. aureus isolated from the patients with corneal ulcer and healthy conjunctival sac. Methods Ten strains of S. aureus isolates were obtained from January to August 2018 in the clinical laboratory of Qingdao Eye Hospital. Five strains of S. aureus isolated from patients with corneal ulcer and five strains of S. aureus isolated from healthy conjunctival sac were included. The gene expression profiles of 10 strains of S. aureus were sequenced by Illumina high-throughput RNA-sequencing ( RNA-Seq) . P≤0. 05 and fold change≥2 were used as the threshold to determine the statistically DEGs. Gene Ontology ( GO ) and Kyoto Encyclopedia of Genes and Genomes ( KEGG) pathway enrichment analyses were used to determine the biological functions of DEGs. Results The genome-wide transcriptional analysis demonstrated that 270 genes were differentially expressed with 138 upregulated genes and 132 downregulated genes in the strains from corneal ulcer. Function analysis of DEGs revealed that genes encoding alpha hemolysin,delta hemolysin,virulence factor EsxA and LysR family transcriptional regulators were significantly upregulated in strains isolated from cornea ulcer. GO enrichment analysis showed that most DEGs was involved in the metabolic process with the biosynthesis, the most significantly related process was the metabolism of inosine monophosphate. The KEGG pathways suggested that a number of metabolic pathways had significant changes,such as S. aureus infection,two-component system and pyruvate metabolism,purine metabolism, which were critical to the pathogenesis of S. aureus corneal ulcer. Conclusions Identification of the DEGs between corneal ulcer isolates and healthy conjunctival isolates of S. aureus is helpful for further investigations on genes or pathways associated with the pathogenesis and therapeutic targets of S. aureus corneal ulcer.
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