CTX-M-14型超广谱β-内酰胺酶原核表达载体的构建及其抗药活性分析

Construction and detection of antibiotic susceptibilities of CTX-M-14 type extended-spectrum-lactamase

目的分析头孢噻肟—水解酶-14(CTX-M-14)型超广谱β-内酰胺酶(ESBL)型菌抗药活性,为临床治疗提供参考。方法收集分离产ESBL大肠埃希菌46株,采用PCR克隆目的基因,采用基因重组技术构建pET28a-CTX-M-14质粒,采用BL21大肠埃希菌作为宿主菌进行表达,采用液体稀释法检测其抗药活性。结果PCR扩增条带在约900bp位置处,序列分析结果显示其与目的基因符合;转化子对青霉素类、头孢一、二代及三代中头孢噻肟、头孢曲松耐药,对庆大霉素、四环素、喹诺酮类药物、碳青酶烯类敏感;对头孢他啶体外试验敏感,对加有酶抑制剂的抗生素稳定敏感。结论本研究成功构建了CTX-M-14型ESBL原核表达载体,且具有广泛抗药活性;此为临床合理选择治疗药物提供了理论依据。

Objective To analyze the antibiotic susceptibilities of CTX-M-14 type extended-spectrum-lactamase（ ESBL）, so as to provide reference for the chnical anti-infection treatment. Methods 46 strains, producing ESBL E. coil were collected from clinical samples, and the target genes were cloned by PCR, pET28a-CTX-M-14 gene were constructed by recombination techinical and expressed in BL21 E. coli , then the antibiotic susceptibilities were detected by agar dilution test. Results The amplified product was at 900 bp in agarose electrophoresis and the DNA sequence was right. The transformant was resistant to penicillins, the first and second generation cephalosporins, cefotaxime and ceftriaxone, sensitive to tetracycline, gentamicin, aminoglycosides,imipenem and ceftazidime in vitr. And it was sensrtive to antibiotics including beta-lactamase inhibitors. Conclusion CTX-M-14 ESBL can be successfully constructed, and transformant of which possess wide resistance to antibiotics. This study provides basis for reasonable application of antibiotics .