| ObjectiveTo investigate the drug resistance phenotype,genotype and homology of carbapenem-resistant Enterobacteriaceae isolated from childern or neonates at two general hospital in China so as to prevent nosocomial infection and provide practice guidelines for the reasonable use of antibiotics.MethodsTotal 21 non-duplicate carbapenem-resistant Enterobacteriaceae isolates,including 8 Enterobacter cloacae?YG18?,10 Klebsiella pneumoniae?FK110?and 3Klebsiella oxytoca?CS13?,were collected,and the identification and antibiotic susceptibility tests of these isolates were done by using VITEK-2 Compact system?BioMérieux?.The putative carbapenemase producers screening were detected by modified Hodge test?MHT?and metallo-beta-lactamases E-test?MBL,IPM+EDTA?.The drug resistance genes,encoding the carbapenemases,extended-spectrum-?-lactamases,AmpC enzymes,PMQR genes and others,were confirmed by using PCR and product sequencing.The clonal relatedness of these strains were determined by pulsed-field gel electrophoresis?PFGE?and multilocus sequence typing?MLST?.The mobility of plasmids was detected by conjugation tests for investigating the transfer characteristics of plasmid.The incompatibility groups of mobile plasmids were determined by the PCR-based replicon typing?PBRT?method.And the plasmids carrying blaNDM-1DM-1 were located by Southern blot hybridization.ResultsThese isolates were highly resistant to most of available antibiotics such as cephalosporins,aminoglycosides and carbapenems.Both Hodge tests and metallo-beta-la-ctamases tests were positive in all collected isolates,and all of them harbored blaNDM-1.The antibiotics resistant genes,including blaNDM-1,blaSHV-12,blaVIM-1,blaCTX-M,blaACC-1,blaTEM-1EM-1 and blaOXA-1XA-1 genes,were detected in Enterobacter cloacae.Both ten Klebsiella pneumoniae and three Klebsiella oxytoca harbored blaNDM-1,blaOXA-1,blaDHA-1,qnrB4,intI and aac?6'?-Ib-cr except blaSHV-12.Pulsed-field gel electrophoresis?PFGE?show that three NDM-1 producing Enterobacter cloacae isolates?YG57?were clonally related,and belong to the clonal group ST177.The ten isolates of NDM-1 producing K.pneumoniae and the three NDM-1 producing K.oxytoca were categorized into the same PFGE types,and belong to ST30 and ST2,respectively.Conjugation experiment confirmed that the plasmids harboring the blaNDM-1DM-1 in 21 strains were transferable.The S1PFGE and Southern hybridization with a NDM-specific probe showed that the blaNDM-1gene was located on a nearly 50kb plasmid in Enterobacter cloacae,280kb IncHIB plasmid in K.pneumoniae,and 275kb IncHIB plasmid in K.oxytoca,respectively.While the PCR-based replicon typing showed that the incompatibility groups of mobile plasmids in Enterobacter cloacae were untyped.ConclusionThe results indicated that there were the NDM-1-producing Enterobacteriaceae spread at the pediatric wards in two general hospital in Zhejiang province and Tianjin city.To our knowledge,it is the first report in worldwide that there was an outbreak of ST30 K.pneumoniae harboring the bla NDM-1,blaOXA-1,bla DHA-1,qnr B4,intI,aac?6'?-Ib-cr,blaSHV-12HV-12 and ST2 K.oxytoca harboring the blaNDM-1,blaOXA-1,blaDHA-1,qnrB4,intI and aac?6'?-Ib-cr in the same neonatal unit in Tianjin city.And that there was another outbreak of ST177 Enterobacter cloacae harboring the blaNDM-1DM-1 genes at the pediatric ward in Zhejiang province.Hence,there is an urgent need on strengthening the surveillance of carbapenem-resistant Enterobacteriaceae and reasonable use of antibiotics. |
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