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Abstract：
OBJECTIVE: To evaluate the expression dynamics of biofilm genes in methicillin-resistant Staphylococcus aureus (MRSA) retrieved from endotracheal tubes (ETT) and to determine how gene regulation is attenuated in vitro where host-environmental factors are no longer present.
METHODS: Biofilm was grown (24 h) in tryptic broth soy plus 0.25% glucose for a clinical MRSA isolate in planktonic state and after sessile growth named ETT-MRSA (S2, S3, S4, S5, S6, S7). Gene expression of five biofilm-related genes (icaC, clfB, ebps, fnbB, and RNA III) was assessed consecutively from day 1 to day 4 after ETT growth through real-time PCR. 16S rRNA was used as a control.
RESULTS: The MRSA isolates retrieved from ETT were capable of producing biofilms dependent on ica. The gene expression dynamics of ETT-MRSA changed progressively compared to planktonic MRSA gene expression under both ambient air (p < 0.001) and ambient air with 5% CO2 (p < 0.001). Dynamic assessment of icaC expression in both atmospheric conditions showed progressive downregulation in vitro compared to in vivo ETT biofilms. The expression patterns of clfB and ebps genes were similar to icaC. In contrast, the expression of the RNA III gene showed progressive upregulation from day 1 to day 4 (p < 0.001).
CONCLUSIONS: MRSA loses its biofilm gene expression in vitro, by adaptive features across multiple generations, as evidenced by the progressive downregulation of icaC and upregulation of RNA III. These findings underscore the significance of host-environment dependence in regulating bacterial biofilm genes, highlighting its importance in diagnostics. Bacterial strains lose their host-specific characteristics as they are cultured in vitro.
METHODS: Biofilm was grown (24 h) in tryptic broth soy plus 0.25% glucose for a clinical MRSA isolate in planktonic state and after sessile growth named ETT-MRSA (S2, S3, S4, S5, S6, S7). Gene expression of five biofilm-related genes (icaC, clfB, ebps, fnbB, and RNA III) was assessed consecutively from day 1 to day 4 after ETT growth through real-time PCR. 16S rRNA was used as a control.
RESULTS: The MRSA isolates retrieved from ETT were capable of producing biofilms dependent on ica. The gene expression dynamics of ETT-MRSA changed progressively compared to planktonic MRSA gene expression under both ambient air (p < 0.001) and ambient air with 5% CO2 (p < 0.001). Dynamic assessment of icaC expression in both atmospheric conditions showed progressive downregulation in vitro compared to in vivo ETT biofilms. The expression patterns of clfB and ebps genes were similar to icaC. In contrast, the expression of the RNA III gene showed progressive upregulation from day 1 to day 4 (p < 0.001).
CONCLUSIONS: MRSA loses its biofilm gene expression in vitro, by adaptive features across multiple generations, as evidenced by the progressive downregulation of icaC and upregulation of RNA III. These findings underscore the significance of host-environment dependence in regulating bacterial biofilm genes, highlighting its importance in diagnostics. Bacterial strains lose their host-specific characteristics as they are cultured in vitro.
摘要：
目的：评估从气管导管（ETT）中提取的耐甲氧西林金黄色葡萄球菌（MRSA）中生物膜基因的表达动态，并确定在宿主环境因素不再存在的情况下，基因调控如何在体外减弱。
方法：生物膜在胰蛋白酶肉汤大豆加0.25％葡萄糖中生长（24小时），用于浮游状态的临床MRSA分离物，并在固着生长后命名为ETT-MRSA（S2，S3，S4，S5，S6，S7）。五个生物膜相关基因的基因表达(icaC，clfB,Ebps,fnbB,和RNAIII）在ETT生长后第1天至第4天通过实时PCR连续评估。使用16SrRNA作为对照。
结果：从ETT回收的MRSA分离株能够产生依赖于ica的生物膜。与在环境空气(p<0.001)和具有5%CO2的环境空气(p<0.001)下的浮游MRSA基因表达相比,ETT-MRSA的基因表达动力学逐渐改变。在两种大气条件下icaC表达的动态评估显示，与体内ETT生物膜相比，体外逐渐下调。clfB和ebps基因的表达模式与icaC相似。相比之下,从第1天到第4天，RNAIII基因的表达显示出进行性上调（p<0.001）。
结论：MRSA在体外失去其生物膜基因表达，通过跨多代的自适应特征，icaC的逐渐下调和RNAIII的上调证明了这一点。这些发现强调了宿主环境依赖性在调节细菌生物膜基因中的重要性。强调其在诊断中的重要性。细菌菌株在体外培养时失去其宿主特异性特征。
方法：生物膜在胰蛋白酶肉汤大豆加0.25％葡萄糖中生长（24小时），用于浮游状态的临床MRSA分离物，并在固着生长后命名为ETT-MRSA（S2，S3，S4，S5，S6，S7）。五个生物膜相关基因的基因表达(icaC，clfB,Ebps,fnbB,和RNAIII）在ETT生长后第1天至第4天通过实时PCR连续评估。使用16SrRNA作为对照。
结果：从ETT回收的MRSA分离株能够产生依赖于ica的生物膜。与在环境空气(p<0.001)和具有5%CO2的环境空气(p<0.001)下的浮游MRSA基因表达相比,ETT-MRSA的基因表达动力学逐渐改变。在两种大气条件下icaC表达的动态评估显示，与体内ETT生物膜相比，体外逐渐下调。clfB和ebps基因的表达模式与icaC相似。相比之下,从第1天到第4天，RNAIII基因的表达显示出进行性上调（p<0.001）。
结论：MRSA在体外失去其生物膜基因表达，通过跨多代的自适应特征，icaC的逐渐下调和RNAIII的上调证明了这一点。这些发现强调了宿主环境依赖性在调节细菌生物膜基因中的重要性。强调其在诊断中的重要性。细菌菌株在体外培养时失去其宿主特异性特征。
