The Detection Of Plasmid-Mediated ESBLs And AmpC β-lactamase Genes In Escherichia Coli Isolated From Animals In Sichuan Province

Escherichia coli is one of the most common zoonotic pathogens, and plays an important role in clinical medicine, veterinary medicine and public health. With the worldwide use of antimicrobial agents in veterinary medicine, the incidence of resistance and multidrug-resistance among pathogenic bacterium has been increasing, which has become a major concern for both medicine and public health problems in the world. Informations from the resistant surveillance and epidemio logical study of plasmid-mediated β-lactam antibiotics resistance genes in Escherichia coli can contribuie to learn emergence and dissemination of resistant genes, and guide the rational use of antimicrobial agents in veterinary medicine.A total of467E. coli strains were isolated from animas in different farms of Sichuan province during2006to2011year. The MIC values of13β-lactam antibiotics and14non-β-lactam antimicrobial agents were determined by broth microdilution method according to CLSI guidelines. The results showed that except the susceptible rate of PM was100%, the resistant rates of other β-lactam antibiotics were above16%, resistant rate of AMC, AMP, AMX. LEX, CFZ, CEF, AZM, CRO, CTX, CAZ, FOX, FEP was91.65%,91.01%,89.08%,80.09%,68.95%,65.52%,61.88%,49.89%,32.76%,24.63%,19.91%,16.70%, respectively. The isolates showed different resistant levels to14non-β-lactam antimicrobial agents, whose resistant rates ranged15±100%. All isolates had multiple resistance, most isolates exhibited multidrug resistance to17-24antimicrobial agents. Different degree of cross-resistance existed in E.coli to different cephalosporins.The crossresistant rate of LEX against other8agents tested was88～99%, and The cross-resistant rate of FEP against other agents tested was the minimum (ranged19.35～50.98%). The cross-resistant rate between two cephalosporins were not always the same, and some were very different. The results suggested that the production of cross-resistance maybe bidirection or unidirection. Cross-resistance also existed in E.coli between cephalosporins and non-(3-lactam antimicrobial agents, these isolates resistant to cephalosporins, were also were resistant to aminoglycosides, chloramphenicol, fluoroquinolones and sulfonamides.ESBLs and AmpC β-lactamase in E.coli isolates were detected using double disk confirmatory test according to CLSI guidelines and three-dimensional test, respectively. The results showed that36.83%(172/467) and13.70%(64/467) of isolates were ESBLs and AmpC P-lactamase-producing positive strains, respectively.60of172ESBLs postive strains were also AmpC β-lactamase-producing positive strains. The resistant level of P-lactamase-producing postive strains was severer than that of negative strains. PCR and PCR-sequencing were used to detect7genes encoding ESBLs,6genes encoding AmpC P-lactamase and1gene encoding NDM-1β-lactamase. The PCR results were as followed:168and64isolates harboured ESBLs and AmpC genes, respectively. TEM, CTX-M, OXA, CMY-2and DHA genes postive strains were148,107,26,63and3strains, respectively.151of all β-lactamase postive strains simultaneously harboured more than one resistant genes, and TEM gene could stimultaneously be deteded in most of CTX-M or CMY-2postive strains. CTX-M and CMY-2were the main p las mid-mediated. ESBLs and AmpC P-lactamase genes in E.coli isolates from animals in Sichuan province.Serological test and conjugation experiment were carried out among176β-lactamase postive strains. Results were as folllowed:31serovars which mainly including O9、O20、 O78、O107、O127、O131and O157, were identified; There were no significant correlation between serovars and resistance; Strains haboured identical resistant gene had different serovars, strains belonging to identical serovar harboured different resistant genes; Rates of AMX-resistant transconjugants in168ESBLs postive strains, and FOX-resistant transconjugants in64AmpC postive, were44.64%and39.06%, respectively. MIC values and PCR results of replicons showed that resistance plasmids carrying genes encoding ESBLs or AmpC P-lactamase could spreaded horizontally among bacterium, which contributed to resistance to β-lactam antibiotics and non-β-lactam antimicrobials, then mukidrug resistance was prevalence.