Objective To investigate the mechanism of Carbapenem resistance in C. Freudii. Methods Antibiotic susceptibilities were determined by agar dilution method. Conjugation experiments were carried out in mixed broth cultures. Plasmid DNA preparations were obtained by using an alkalinelysis technique and were digested by various endonucleases; The crude β-lactamase extracts of C. Freudii and E. Coli transconjugant were subjected to analytical isoelectric focusing( IEF )Specific PCR amplification and DNA sequence analysis were performed to confirm the β-lactamse type. Results The C. Freundii isolate showed resistance against Carbapenemes. The MICs of imipen-em and meropenem were both 64 mg · L-1 . The isolate was also resistant against penicillins, cephalosorins, cefoxitins, aztreonam, quinolo-nes,and aminoglycosides. The conjugant results showed the antibiotics can transfer by plasmid. Isoelectric focusing demonstrated two β-lactamases with the isoelectric points of 5. 0 and 7. 5 in conjugant. Specific PCR amplification and DNA sequence analysis show ed that the C. Freudii produce the gene of KPC-2. Coclusion the product of KPC-2 Carbapenem was the first and foremost dues of Carbapenem-risitance and it can transfer by plasmid.%目的 研究弗劳地枸橼酸杆菌对碳青霉烯类抗生素的耐药机制.方法 采用琼脂对倍稀释法检测弗劳地枸橼酸杆菌对亚胺培南和美罗培南以及其他常见药物的最低抑菌浓度(MIC).等电聚集电泳分析其β-内酰胺酶类型,聚合酶链反应(PCR)和DNA序列分析检测β-内酰胺酶基因型,接合试验分析其耐药质粒传递情况.结果 弗劳地枸橼酸杆菌对亚胺培南和美罗培南的MIC均为64 mg·L-1,对青霉素类、头孢菌素类、头孢西丁、氨曲南和氨基糖苷类均耐药.转移接合结果显示对亚胺培南和美罗培南的耐药性可以通过质粒转移.等电聚焦电泳结果显示转移接合子具有等电点(PI)约为5.0、7.5的2种β-内酰胺酶.PCR检测及序列分析结果显示该菌产生KPC-2基因.结论 KPC-2型碳青酶烯酶的产生是引起弗劳地枸橼酸杆菌对碳青酶烯酶耐药的主要原因,并且该耐药性能通过质粒进行转移.
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