Study On Genotype And Resistance Characters Of Plasmid-mediated AmpC β-lactamases And ESBLs In Escherichia Coli

Objectives:Escherichia coli(E.coli)is one of the most common gram-negative bacterias causing clinical infection,especially nosocomial infection. Its drug resistant mechanisms are very complex,one of which toβ-lactam antibiotics is mainly producing AmpCβ-lactamases and extended-spectrumβ-lactamases (ESBLs).When one bacteria can produce both of the enzymes,it will be more resistant towards antibiotics.In order to provide a reference for understanding the trend of drug resistance,the reasonably use of antibiotics and the control of the resistant bacteria's dissemination,we collected clinical E.coli isolates from two hosipial in Tianjin,analysed their resistance to antibiotics,and investigated the prevalence and genotypes of plasmid-mediated AmpCβ-lactamases and ESBLs.Methods:Clinical E.coli isolates at two hospitals in Tianjin were collected during March to December in 2006,which were categorized into nosocomial infection isolates and community infection isolates.Kirby-Bauer(K-B)test and agar dilution test were used to test antimicrobial susceptibility.Phenotypic confirmatory test and three-dimensional extract test were used for identification of ESBLs and AmpCβ-lactamases separately.Polymerase chain reaction(PCR)was performed to identify the genotypes of plasmid-mediated AmpCβ-lactamases.Location of theβ-lactamase genes was determined by plasmid conjugation test,and K-B test was performed to test antimicrobial susceptibility of both donor isolates and conjugons. ESBLs genotypes of isolates were identified by PCR.Some PCR products were sequenced and analyzed for homology on Genebank.Results:1 203 strains at two hospitals in Tianjin were collected during March to December in 2006,and the proportion of nosocomial infection isolates was 19.7%.Imipenem was the most sensitive antibiotics,which resistant rate was only 0.5%.The resistant rates to ceftazidime,cefepime,cefoperazone/sulbactam,cefoxitin and amikacin were between 10%～20%,while the resistant rates to cefazolin,cefuroxime,piperacillin, gentamicin,ofloxacin,levofloxacin and ciprofloxacin were higher than 60%. nosocornial infection isolates were resistant to antibiotics more severely.2 55 strains were positive for ESBLs phenotypic confirmatory test,the positive rate was 27.1%.62.5%nosocomial infection isolates were ESBLs positive,this rate was obviously higher than community infection isolates'(P＜0.05).The antibiotics resistant rates of isolates producing ESBLs were higher than non-producing isolates.The resistant rate of isolates producing ESBLs to cefotaxime was significantly higher than ceftazidime.12 strains were positive for three-dimensional extract test,and the positive rate was 5.9%.10.0%nosocomial infection isolates were AmpCβ-lactamases positive.The rate was higher than community infection isolates', but the difference was not statistically significant(P＞0.05).The detection rate of double positive isolates was 3.4%.3 8 strains of E.coli were positive for plasmid-mediated AmpCβ-lactamases gene amplification:6 strains for CIT genotype and 2 strains for DHA genotype.Those 8 isolates were all successfully conjugated,and the resistant patterns were transmitted. Antimicrobial susceptibility test showed that,donor strains were resistant to almost all third-generation cephoiosporins,gentamicin and quinolones,only sensitive to imipenem.Compared with those above,the resistant of conjugons to cefepime and cefoperazone/sulbactam were lower,while two appear resistant to aminoglycosides.4 Among those 8 strains mentioned above,three were ESBLs positive,their genotypic combinations were TEM and CTX-M-3(E59);TEM and CTX-M-14(E112); TEM,CTX-M-3 and CTX-M-14(E220),respectively.Compared in Genebank,the sequences of CIT type from E59 strain;DHA type,TEM type,CTX-M-3 group type and CTX-M-14 group type from E220 strain were found to be 100%homogenous to CMY-2,DHA-1,TEM-1,CTX-M-55 and CTX-M-19 genotypes.Couclusions:The resistant rates to antibiotics of E.coli at two hospitals in Tianjin was slightly higher than the country's average level in 2005,while the positive rates of strains producing plasmid-mediated AmpCβ-lactamases and ESBLs were lower than some other regions in China.Our study was the first to report the genotype distribution of plasmid-mediated AmpCβ-lactamases in E.coli in Tianjin, and the results were similar with reports from abroad,but inconsistent with those reports of majority regions in China,which means that different areas may have different enzyme genotypes.We also reported 3 strains that could produce both plasmid-mediated AmpCβ-lactamases and ESBLs,one of which had a genotypic combination(AmpCβ-lactamases DHA-1 gene,TEM-1β-lactamases gene, CTX-M-19 and CTX-M-55 ESBLs genes)that has not been reported before.