Guangdong Provincial Key Laboratory of Veterinary Pharmaceutics Development and Safety Evaluation, Guangzhou, China; College of Life Science and Technology, Southwest University for Nationalities, Chengdu, China.
Environ Toxicol Pharmacol. 2010 Jul;30(1):5-10. doi: 10.1016/j.etap.2010.03.004. Epub 2010 Mar 9.
In our studies, qualitative (scanning electron microscope) and semi-quantitative (modified crystal violet staining method) methods had been used to evaluate Escherichia coli biofilm-forming ability. Broth microdilution method and enterobacterial repetitive intergenic consensus-based PCR (ERIC-PCR) were performed to study E. coli drug resistance pattern and genetic typing. Based on the results above, we studied the correlation between biofilm-forming ability phenotype, drug resistance pattern and genetic typing in E. coli. Real-time qPCR (qRT-PCR) was used to reveal mRNA expression level of E. coli biofilm related multiple antibiotics resistance genes (acrA, agn43, csgA, csgD, ompF and pgaA) under different concentrations of four aminoglycoside pressures. Our results showed that: (i) forty-nine out of 64 strains of E. coli (76.56%) showed significant production of biofilm and most of them performed weak biofilm-forming ability; (ii) ERIC-PCR showed that there was significant correlation between biofilm-forming ability and genotype; while there was weak correlation between biofilm-forming ability and drug resistance patterns based upon the results of semi-quantitative method and antibiotics susceptibility test; (iii) qRT-PCR revealed mRNA expression of acrA, agn43, csgA, csgD, ompF and pgaA genes changed accordingly by stimulation of different concentrations of four aminoglycosides.
在我们的研究中,使用了定性(扫描电子显微镜)和半定量(改良结晶紫染色法)方法来评估大肠杆菌生物膜形成能力。采用肉汤微量稀释法和肠杆菌重复基因间一致性聚合酶链反应(ERIC-PCR)来研究大肠杆菌的耐药模式和遗传分型。基于以上结果,我们研究了大肠杆菌生物膜形成能力表型、耐药模式和遗传分型之间的相关性。采用实时 qPCR(qRT-PCR)揭示了在不同浓度的四种氨基糖苷类药物压力下,大肠杆菌生物膜相关的多种抗生素耐药基因(acrA、agn43、csgA、csgD、ompF 和 pgaA)的 mRNA 表达水平。我们的结果表明:(i)64 株大肠杆菌中有 49 株(76.56%)表现出明显的生物膜生成能力,其中大多数表现出较弱的生物膜形成能力;(ii)ERIC-PCR 显示生物膜形成能力与基因型之间存在显著相关性;而基于半定量方法和抗生素药敏试验的结果,生物膜形成能力与耐药模式之间的相关性较弱;(iii)qRT-PCR 显示 acrA、agn43、csgA、csgD、ompF 和 pgaA 基因的 mRNA 表达在受到四种氨基糖苷类药物不同浓度的刺激时相应发生变化。