Genotype Detection Of Extended-spectrum β-lactamases Produced By E.coli Isolates From Duck

Plasmid-encoded Extended-spectrumβ-lactamases is a large species ofβ-lactamases capable of hydrolyzing oxyimino-cephalosporins like third-generation cephalosporins(cefotaxime,cefetaxime,ceftriaxone) and monobactams (like aztreonam).They are inhibited by clavulanic acid. ESBLS are produced by Eschrichia coli.,Klebsiella pneumoniae ,Serratia marcescens,Protens mirabilis,Enterobacter cloacae ,Acinetobacter sp. And Psendomonas aernginosa and so on. So far, more than 400 kinds of ESBLS have been detected,and the types of new genotypes is increasing. In accordance with classification of the Bradford method,ESBLS can be divided into five types: TEM,CTX-M,SHV,OXA and other types..At present,ESBLS has become a major reasons thatβ-lactam antibiotics resistance. With the use of a large number of antibiotics,animals also appeared in varying degrees of resistance and multi-drug resistance phenomenon.Production of Extended-spectrumβ-lactamases is a major cause of resistance.In this paper ,we did works as follows: 40 isolates of E.coli were detected ESBLS by CLSI commended methods. MIC and drug susceptibility were detected by susceptibility test and K-B disk diffusion menthod. ESBLS genotypes and subtypes were detected by PCR .The experiment results showed that 18 E.coli isolates were ESBLs producers.A total of 45% strains were positive. The results of susceptibility testing showed that resistance rates of ESBLs-producing strains to 21 antibiotics were 17%～100%,which were higher than non-ESBLs -producing strains. The sensitive rates of Carbapenems to ESBLs-producing and non-ESBLs -producing strains were 100%. The MIC was significantly reduced through different drug combination against ESBLS-producing strains.The ESBLs-producing strains represented multiple resistance and cross resistance.(like Aminoglycosides, Quinolones, Chloramphenicol).Antibiotics/β-lactam inhibitor and/or different antibiotics can improve distinctly against antibiotic activity of ESBLs-producing strains.18 ESBLs-producing isolates were subjected to PCR experiments to detect ESBL genes,including blaTEM,blaSHV,blaCTX-M,blaOXA.The results showed that 18 starains were TEM(861bp),12 starains were CTX-M and 9 starains were OXA.None of blaSHV was present in all isolates. Then the genotype of ESBLs were characterized by PCR and sequencing. The results showed that 12 starains carrying TEM and CTX-M, 9 starains carrying TEM and OXA,and 3 starains carrying TEM ,CTX-M and OXA. The sequence homology was 99.8% ~ 100% among TEM-type clinical isolates. The sequence homology of submitted sequence FJ405215 and AY293072(TEM-1) was 99.9%. There were two mutations, but not caused by mutations of amino acid sites, it was still broad-spectrum TEM-1 type enzyme.The sequence homology was 99.9% ~ 100% among CTX-M-type clinical isolates .The submitted sequence FJ405218,FJ405219 were the same as that of in Genebank AF252622,AY156923,so they are also CTX-M-14–type and CTX-M-27-type ESBLS.The sequence homology was 100% among OXA-type clinical isolates.The submitted sequence FJ841897 was the same as that of in Genebank EF415651(OXA-1),so it is also OXA-1-type.