Lyu Chongyang, Hu Haijing, Cai Linlin, He Shuwen, Xu Xinglian, Zhou Guanghong, Wang Huhu
State Key Laboratory of Meat Quality Control and Cultured Meat Development, College of Food Science and Technology, Nanjing Agricultural University, Nanjing, People's Republic of China.
State Key Laboratory of Meat Quality Control and Cultured Meat Development, College of Food Science and Technology, Nanjing Agricultural University, Nanjing, People's Republic of China; College of Food Science and Pharmacy, Xinjiang Agricultural University, Urumqi, Xinjiang, People's Republic of China.
J Adv Res. 2025 May;71:127-139. doi: 10.1016/j.jare.2024.06.008. Epub 2024 Jun 7.
Salmonella Enteritidis has brought great harm to public health, animal production and food safety worldwide. The biofilm formed by Salmonella Enteritidis plays a critical role in microbial cross-contamination. Small non-coding RNAs (sRNAs) have been demonstrated to be responsible for regulating the formation of biofilm. The sRNA SaaS has been identified previously, that promotes pathogenicity by regulating invasion and virulence factors. However, whether the SaaS is implicated in regulating biofilm formation in abiotic surfaces remains unclear.
This study aimed to clarify the effect of SaaS in Salmonella Enteritidis and explore the modulatory mechanism on the biofilm formation.
Motility characteristics and total biomass of biofilm of test strains were investigated by the phenotypes in three soft agar plates and crystal violet staining in polystyrene microplates. Studies of microscopic structure and extracellular polymeric substances (EPS) of biofilm on solid surfaces were carried out using confocal laser scanning microscope (CLSM) and Raman spectra. Transcriptomics and proteomics were applied to analyze the changes of gene expression and EPS component. The RNA-protein pull-down and promoter-reporter β-galactosidase activity assays were employed to analyze RNA binding proteins and identify target mRNAs, respectively.
SaaS inhibits biofilm formation by repressing the adhesion potential and the secretion of EPS components. Integration of transcriptomics and proteomics analysis revealed that SaaS strengthened the expression of the flagellar synthesis system and downregulated the expression of curli amyloid fibers. Furthermore, RNA-protein pull-down interactome datasets indicated that SaaS binds to Hfq (an RNA molecular chaperone protein, known as a host factor for phage Qbeta RNA replication) uniquely among 193 candidate proteins, and promoter-reporter β-galactosidase activity assay confirmed target mRNAs including hilD, cheA, and csgA.
SaaS inhibits the properties of bacterial mobility, perturbs the secretion of EPS, and contributes to the inhibition of biofilm formation by interacting with target mRNA (hilD, cheA, and csgA) through the Hfq-mediated pathway.
肠炎沙门氏菌已对全球公共卫生、动物生产和食品安全造成了巨大危害。肠炎沙门氏菌形成的生物膜在微生物交叉污染中起着关键作用。小非编码RNA（sRNA）已被证明负责调节生物膜的形成。先前已鉴定出sRNA SaaS，它通过调节侵袭和毒力因子来促进致病性。然而，SaaS是否参与调节非生物表面生物膜的形成仍不清楚。
本研究旨在阐明SaaS在肠炎沙门氏菌中的作用，并探索其对生物膜形成的调节机制。
通过在三个软琼脂平板上的表型以及在聚苯乙烯微孔板中的结晶紫染色来研究测试菌株的运动特性和生物膜的总生物量。使用共聚焦激光扫描显微镜（CLSM）和拉曼光谱对固体表面生物膜的微观结构和胞外聚合物（EPS）进行研究。应用转录组学和蛋白质组学来分析基因表达和EPS成分的变化。分别采用RNA-蛋白质下拉和启动子-报告基因β-半乳糖苷酶活性测定来分析RNA结合蛋白并鉴定靶标mRNA。
SaaS通过抑制黏附潜能和EPS成分的分泌来抑制生物膜形成。转录组学和蛋白质组学分析的整合表明，SaaS增强了鞭毛合成系统的表达并下调了卷曲菌毛淀粉样纤维的表达。此外，RNA-蛋白质下拉相互作用组数据集表明，在193种候选蛋白中，SaaS仅与Hfq（一种RNA分子伴侣蛋白，是噬菌体Qβ RNA复制的宿主因子）结合，启动子-报告基因β-半乳糖苷酶活性测定证实了包括hilD、cheA和csgA在内的靶标mRNA。
SaaS抑制细菌的运动特性，扰乱EPS的分泌，并通过Hfq介导的途径与靶标mRNA（hilD、cheA和csgA）相互作用，从而有助于抑制生物膜的形成。
