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目的探讨HMGN2对肺炎克雷伯氏杆菌耐药性的影响。方法分离、纯化并鉴定兔胸腺组织HMGN2。用质粒提取试剂盒提取耐药肺炎克雷伯杆菌中的质粒pMG252,PCR扩增其耐药基因qnrA;凝胶阻滞实验检测HMGN2与耐药质粒pMG252及qnrA基因结合;用不同浓度的HMGN2刺激培养肺炎克雷伯氏杆菌,然后用试管二倍稀释法测定环丙沙星对其MIC值的改变,微量法测定细菌的生长曲线,检测HMGN2对耐药菌株耐药性的逆转效果;将重组HMGN2及耐药质粒pMG252转化到大肠杆菌DH5α中,检测重组HMGN2对细菌耐药性的影响。结果HMGN2可明显阻滞pMG252质粒及qnrA基因的泳动速率,且随HMGN2浓度增加,qnrA基因的泳动速率逐渐减慢;15μg/ml HMGN2与细菌共孵育后,环丙沙星对耐药肺炎克雷伯氏杆菌的MIC值明显下降,且该耐药株的生长与对照组和低剂量组相比明显的受到抑制;pET-32a-c(+)-HMGN2重组表达质粒可时环丙沙星对大肠杆菌DH5α的MIC值明显小于其对照组,下降到其1/16。结论HMGN2能有效逆转含耐药质粒pMG252的肺炎克雷伯氏杆菌等对喹诺酮类抗生素的耐药性。
Objective To investigate the effect of HMGN2 on the drug resistance of Klebsiella pneumoniae. Methods The rabbit thymus HMGN2 was isolated, purified and identified. Plasmid pMG252 was extracted from Klebsiella pneumoniae resistant to Klebsiella pneumoniae by plasmid extraction kit, and its resistance gene qnrA was amplified by PCR. Gel-block assay was used to detect the binding of HMGN2 to drug-resistant plasmids pMG252 and qnrA. Different concentrations of HMGN2 Corynebacterium pneumoniae was cultured, and then the change of MIC value of ciprofloxacin was determined by two-fold dilution in vitro. The growth curve of bacteria was determined by micro-method and the reversal effect of drug resistance of HMGN2 on drug- HMGN2 and drug-resistant plasmid pMG252 were transformed into E. coli DH5α to detect the effect of recombinant HMGN2 on bacterial resistance. Results The migration rate of pMG252 plasmid and qnrA gene was significantly blocked by HMGN2. With the increase of HMGN2 concentration, the migration rate of qnrA gene gradually slowed down. When incubated with 15 μg / ml HMGN2 and bacteria, the effect of ciprofloxacin on the resistance to pneumonia The MIC value of Klebsiella pneumoniae was significantly decreased, and the growth of the drug-resistant strain was significantly inhibited compared with the control group and the low-dose group; pET-32a-c (+) - HMGN2 recombinant expression plasmid Star E. coli DH5α MIC value was significantly less than its control, down to 1/16. Conclusion HMGN2 can effectively reverse the resistance of quinolone antibiotics such as Klebsiella pneumoniae containing drug-resistant plasmid pMG252.