Drug Resistance Mechanisms Of Carbapenems-Resistance Escherichia Coli

ObjectiveTo investigate the prevalence of resistance genes,resistance mechanisms and molecular characteristics of carbapenems-resistance Escherichia coli(E.coli)isolated from Tai'an central hospital,July 2014 to December 2015,and to provide evidences for the prevalence and outbreak of nosocomial infection,and for timely preventing dissemination of carbapenems-resistance E.coli.MethodsA total of 12 nonduplicated carbapenems-resistance E.coli were isolated from Tai'an central hospital.Drug sensitivity of piperacillin tazobactam,cefotaxime,ceftazidime,cefepime,cefoxitin,imipenem,meropenem,ertapenem,aztreonam,amikacin,levofloxacin,gentamicin,sulfamethoxazole were determined by Kirby-Bauer test,E-test,and fully automated microbial identification system which include SIEMENS microscan96 plus,BD PHOENIXTM System and VITEK? 2 COMPACT.Phenotype of carbapenemasesproducing E.coli were determined by modified Hodge test and EDTA-disk synergy test.Resistance genes encoding ?-lactamases,aminoglycosidase,and anti-quinolone related genes were detected by PCR,the amplification products were then sequenced and verified by whole genome sequencing(shotgun method).Chromosomal polymorphism of drug-resistant strains were determined by pulsed-field gel electrophoresis(PFGE)analysis,sequence type and phylogenetic relationships were also detected by multilocus sequence typing(MLST).Carbonyl cyanidem-chlorophenylhydrazone(CCCP)tests were performed to observe whether the active efflux mechanism contributed to the carbapenems resistance.Expression of outer membrane protein was analyzed by SDS-PAGE electrophoresis.Then,to determine the cotransferable characteristics between strains,conjugation tests were performed.ResultsA total of 12 carbapenems-resistance E.coli were collected,7 were isolated from ICU,1 from endocrine ward I,1 from pediatric surgical ward,1 from general surgery eastern ward,1 from neurosurgery ICU,and 1 from emergency ward.Drug susceptibility testing results showed that the 12 strains were all carbapenems-resistance,and were all positive for modified Hodge test and EDTA-disk synergy test.According to the resistance genes,2 strains harbored blaNDM-1,1 harbored blaNDM-4,and 9 harbored blaNDM-5.To avoid redundancy,other resistance genes were not presented here.PFGE results evidenced that 12 strains were divergent and not clone-related.Strain 7 and 9,while strain 9 and 10 showed relatively closer phylogenetic relationship,when isolates clustering together with 80% were considered to be the same genotype.MLST confirmed that 12 strains were classified to ST10,ST167,ST405,ST5703.CCCP could not improve the drug sensitivity of the strains to carbapenems(imipenem,meropenem and ertapenem).Deletion of outer membrane protein was not detected in the strains.Successful conjugation was determined in 9 out of 12 strains,suggesting that part of the resistance genes could disseminate between different species.ConclusionCarbapenems-resistance E.coli were verified in Tai'an,the New Delhi metallo-betalactamase genes were major contributors of the resistance mechanism,and resistance genes could be disseminated between species,which may lead to the spread of drug resistance.