Analysis Of Drug Resistance Mechanism And Molecular Epidemiology Of NDM Producing Escherichia Coli Strains

Objective:To investigate the drug resistance determinants,genetic relatedness and gene horizontal transfer of the clinically isolated NDM-5 producing Escherichia coli strains,so can provide useful suggestions for hospital-acquired infection and prevention.Methods:Carbapenem-resistant Escherichia coli strains confirmed by Vitek 2 Compact system were collected during the period from July 2015 to March 2016.Modified Hodge test and IPM-EDTA synergy test were used to screen the production of metallo-β-lactamase.Microdilution method(for ceftazidime,sulfamethoxazole,piperacillin/tazobactam,aztreonam,amikacin,gentamicin,levofloxacin,imipenem,meropenem,etapenem and polymyxin)and E-test strips(for tigecyclin)were conducted to confirm MICs.PCR and nucleotide sequencing methods were performed to detect the resistant genes of carbapenems,ESBLs,aminoglycosides,quinolones and polymyxin.Pulse-field gel electrophoresis(PFGE)and multilocus sequence typing(MLST)methods were applied to analyze the clonal relatedness.Conjugation experiment and PCR-based replicon typing methods were used to investigate the characteristics of blaDM-5 gene-carrying plasmids.Results:The results of Vitek 2 Compact system identified a total of 920 Escherichia coli strains during the period from July 2015 to March 2016.Among them,28 exhibited resistance to carbapenems(imipenem and meropenem).The modified Hodge test and IPM-EDTA synergy test indicated 18 strains can produce metallo-β-lactamase.Drug susceptibility testing results showed all strains exhibited high resistance to ceftazidime,piperacillin/tazobactam,imipenem,meropenem,etapenem and levofloxacin.In contrast,the resistance rate of amikacin,gentamicin and polymycin were 33.3%,0%and 0%,respectively.Followed sequencing of positive amplicons of carbapenem-resistant genes showed the type of carbapenemase-encoding gene were all blaNDm-5 Moreover,the ESBLs,aminoglycosides and quinolones resistant genes were detected in 16,4 and 7 strains,respectively.Fortunately,no mcr-1 gene was detected.MLST devided the 18 blaNDM-5 positive E.coli strains into 8 sequence types,among which the ST167,ST410 and ST101 were three main types.Furthermore,similarities between the PFGE patterns revealed strains in ST167(except CREC-133),ST410 and ST101 had ever caused clonal spread in our hospital.Fifteen blaNDM-5 positive E.coli strains transferred blaNDM-5-toE.coli J53 successfully and the blaNDM-5 gene were all located one IncX3 plasmids,causing the MIC of carmbapenems in conjugations increased at least 32 folds.Conclusion:Producing NDM-5 was the main mechanism for E.coli strains being resistant to carbapenem in our hospital.Here,we firstly reported a high incidence and spread of the blaNDM-5 positive E.coli strains.Clonal spread combined with horizontal transfer of blaNDM-5 gene contributed to the high prevalence of blaNDM-5 positive E.coli strains in our hospital.Owing to the common situation of multi-drug resistance in blaNDM-5 positive E.coli strains had brought great challenge to the clinical anti-infection therapy,strict infection control measures and reasonable application of antibiotics were urgently needed.