基于比较转录组学分析NaCl胁迫影响德尔卑沙门氏菌的耐渗机制

NaCl stress affects the permeability of Salmonella enterica subsp. enterica Derby: a study based on comparative transcriptomics

摘要:

【目的】研究高渗胁迫条件下德尔卑沙门氏菌(Salmonella enterica subsp. enterica Derby, S. Derby)的转录组调控机制，分析差异表达基因(differentially expressed genes, DEGs)表达水平，探究在高渗胁迫影响下德尔卑沙门氏菌耐渗反应的相关代谢通路。【方法】通过高渗胁迫诱导德尔卑沙门氏菌的耐渗性，提取菌株的总RNA，去除rRNA，构建cDNA文库。利用转录组测序技术及生物学信息技术分析相关DEGs，并通过实时荧光定量PCR (real-time fluorescence quantitative PCR, qRT-PCR)进行验证。【结果】胁迫组德尔卑沙门氏菌通过转录组测序结果发现有3 950个DEGs，其中具有显著上调的基因21个，显著下调基因38个。涉及到细胞膜蛋白、氨基酸的代谢等相关基因上调，协助德尔卑沙门氏菌在高渗环境中存活。与此同时，胁迫组德尔卑沙门氏菌的糖转运系统(sugar transport system, PTS)、糖酵解过程以及抗氧化性相关基因表达显著下调，这是由于高渗环境菌体需要在体内储存大量糖类等物质，从而降低了糖原的消耗，进而导致细胞外膜的脂多糖合成受到抑制，降低了高渗胁迫下德尔卑沙门氏菌细胞膜表面的O抗原的合成。【结论】高渗环境诱导后显著提高了德尔卑沙门氏菌的耐渗性，其中Na⁺/H⁺逆向转运蛋以及谷氨酸的代谢通路发挥着重要的作用，为进一步了解以及更好地控制其在食品中的污染提供了理论依据。

关键词:高渗胁迫;德尔卑沙门氏菌;差异表达基因(DEGs);转录组分析;实时荧光定量PCR

Abstract:

[Objective] This study aims to decipher the mechanism of Salmonella enterica subsp. enterica Derby (S. Derby) adapting to hypertonic stress at the transcriptional level. We mined the differentially expressed genes (DEGs) to explore the metabolic pathways associated with the response of S. Derby to the stress. [Methods] After the hyperosmotic tolerance of S. Derby was induced, we extracted the total RNA, removed the rRNA, and constructed a cDNA library. The relevant DEGs were identified by transcriptome sequencing and bioinformatics tools and verified by real-time fluorescence quantitative PCR. [Results] After hyperosmotic stimulation, 3 950 DEGs were identified by transcriptome sequencing, which included 21 significantly up-regulated genes and 38 significantly down-regulated genes. The genes involved in the efflux of Na⁺ from the cell membrane and the metabolism of amino acids were up-regulated, which can provide energy and help S. Derby survive in a hyperosmotic environment. The genes associated with the sugar transport system (PTS), glycolysis, and anti-oxidation of S. Derby were significantly down-regulated in the stress group. Under hyperosmotic stress, the bacteria cannot take up carbohydrates from the external environment and thus the synthesis of lipopolysaccharide in the outer membrane of the cell is inhibited, which reduces the invasiveness of S. Derby, thereby increasing the toxicity of S. Derby. [Conclusion] Under saturated NaCl stress, the osmotic tolerance of S. Derby is significantly improved, during which the Na⁺/H⁺ antiporter and the glutamate metabolic pathway play a key role. The findings provide a theoretical basis for the further understanding and control of S. Derby contamination in food.

Key words:hyperosmotic stress;Salmonella enterica subsp. enterica Derby;differentially expressed genes;transcriptome analysis;real-time fluorescence quantitative PCR