Study On The Common Genotype Of ESBLs Producing Escherichia Coli And Its Mechanism Of Quinolone-resistance In Primary Hospital

Objective:To understand the distribution of common genotypes of ESBLs-producing E.coli isolated from primary hospital in this area,and to provide experimental basis for clinical treatment,we investigate the mechanism of chromosome and plasmid mediated combined with the fluoroquinolone-resistance situation of these bacteria.Methods:1.114 E.coli strains were isolated from clinical samples from March 2014 to March 2015 in the local hospital.In these strains ESBLs were detested by ESBLs confirmatory test by disk diffusion method,and the antimicrobial susceptibility results were tested by using automatic bacteria identification and antimicrobial sensitivity instrument.2.PCR method were used to detect whether blaCTX-M-1,blaCTX-M-2,blaCTX-M-9 and blaSHV genes being carried by these strains.10 strains would be randomly selected from each group of genotype to determine the gene subtype by direct bidirectional DNA sequencing of PCR products.3.20 Strains fluoroquinolones resistanc of the above ESBLs-producing E.coli were randomly selected to amplified gyrA/gyrB/parC/parE genes by PCR method,the PCR products were sequenced by bidirectional DNAsequencing method.The mutation sites were determined by blast comparison of the sequencing results.We used PCR method to detect whether qnrA,qnrB,qnrS or aac(6’)-Ib-cr genes were carried by the above 50 strains ESBLs-producing E.coli,and the PCR positive products were direct bidirectional DNA sequenced.Results:1.50 strains of ESBLs-producing E.coli(43.9%)were isolated from 114 E.coli strains of 50 ESBLs-producing strains.Antimicrobial resistence rates of amoxicillin,piperacillin,ceftriaxone and cefuroxime are 100%,resistance rates of levofloxacin and ciprofloxacin were 70%and 80%;and there were more high drug resistance to the cefotaxime,sulfamethoxazole,gentamycin,aztreonam,levofloxacin and ciprofloxacin,but had a high sensitivity to compound preparations of β-lactamase inhibit,carbapenems,amikacin and cefoxitin,in addition to the resistant rate of amoxicillin/clavulanic acid was 24%,the rest were below 10%.2.There were 46(92%)of 50 strains carrying blaCTX-M gene;Among them,33 strains(66%)carryed the blaCTX-M-9,19 strains(38%)carryed the blaCTX-M-1,6 strains carryed above two genes,blaCTX-M-2 and blaSHV were not detected.In CTX-M-1 group,the CTX-M-15 subtype was dominant;in CTX-M-9 group,the CTX-M-14 was dominant.3.In the randomly selected 20 strains of ESBLs-producing E.coli which resistant to fluoroquinolones,20 strains of bacteria had site mutations in GyrA 83 and 87;19 strains had site mutation in ParC 80,and 3 strains in ParC 84;9 strains had site mutation in ParE 458,and 1strain in ParE 445;site mutation in gyrB was not found.All of the strains carrying the qnr-gene produced CTX-M genotype ESBL.Meanwhile,7 strains in 8 AAC(6 ’)-Ib-cr positive strains produced CTX-M genotype ESBL.Conclusion:1.The rate of ESBLs-producing E.coli was high in the local primary hospital,these strains are usually multiple antimicrobial resistant bacteria,but they were generally sensitive with a high degree to Compound preparations of β-lactamase inhibitors,carbapenems,amikacin and cefoxitin.2.There was a certain amount of CTX-M genotype ESBLs in local primary hospital.The genotype CTX-M-9 and CTX-M-1 are the dominant type in ESBLs-producing E.coli in this area.3.The fluoroquinolone resistance in ESBLs-producing E.coli in the local primary hospital was related to the bacterial target gene mutation mechanism.The mutation points not only occurred in gyrA and parC,but also occurred in parE,which had been rarely reported in other experiments.At the same time,we also found that the mechanism of plasmid-mediated fluoroquinolone resistance in this area,and these strains were producing CTX-M genotype of ESBLs.