The Role Of Integrons And Gene Cassettes In Mediation And Dissemination Of Antibiotic Resistance

ObjectiveTo investigate the antibiotic resistance, the presence and class of integrons and the content and order of gene cassettes in clinical isolates of Escherichia coli. To observe the relationship between gene cassettes and resistance phenotype , To analyze the distribution of integrons among different genotype isolates. To illuminate the molecular mechanism of integrons and gene cassettes in mediation and dissemination of antibiotic resistance.MthodsIdentification, ESBLs detection and susceptibility to 20 antibiotics of 94 Escherichia coli isolates were performed by Microscan WalkAway40, and susceptibility to 4 more antibiotics determined by the disk diffusion assay. Class 1 integrons,intI2 and intI3 were detected by PCR with total DNA and plasmid DNA respectively. The RFLP analysis of amplicons of class 1 integrons with identical size was carried out. The purified amplicons of class 1 integrons were sequenced. We tried to find out the content and order ofgene cassettes in integrons by searching in GeneBank. Escherichia coli isolates were typed by the pulsed-field gel electrophoresis(PFGE).ResultsOver 50% of the isolates were resistant to 15 antibiotics. Antibiotics with high resistance rate included Ampicillin(94%), Piperacillin(87%),and so on. 45 isolates(47.9%) produced ESBLs. 90(95.7%) isolates were multidrug resistant(resistance to over 3 antibiotics). 52(55.3%) isolates had class 1 integrons on total DNA , 41 (43.6%) on plasmid DNA and 14(14.9%) on Chromosomal DNA. 3 isolates had class 1 integrons on both plasmid DNA and Chromosomal DNA. The size of class 1 integrons ranged from 600bp to 3000bp. Each isolate had 1 or 2 class 1 integrons. 5 isolates(5.3%) had 2 class 1 integrons. intI2 and intI3 were not detected. Class 1 integrons indistinguishable with respect to size had identical restriction patterns. It was supposed that they might have the same sequences. The predominant gene cassettes included dfrl7(encoding resistance to trimethoprim) and aadA5 (encoding resistance to streptomycin and spectinomycin). The main order of inserted gene cassettes was dfrl7-aadA5. The major isolates carrying dfr were resistant to sulfamethoxazole/trimethoprim, Most isolates carrying aadA were not susceptible to streptomycin. 82 Escherichia coli isolates which could be typed by PFGE had various genotypes. Isolates with different genotype harbored identical integronsConclusionThe situation of antibiotic resistance in clinical isolates of Escherichia coli is serious. Class 1 integrons are widespread in clinical Escherichia coli. Gene cassettes in integrons are greatly related with old antibiotics such as trimethoprim and old aminoglycoside compounds. As a whole, isolates carrying gene cassette have corresponding resistance phenotype. Integrons mediated antibiotic resistance. Genotyping is suggestive of horizontal intraspecies transfer of integrons. Through this study we obtained molecular epidemiological data about integrons and gene cassettes in clinical isolates of Escherichia coli, and contributes to understand deeply molecular mechanism of integrons and gene cassettes in dissemination and mediation of antibiotic resistance. It supplys theoretic basis for making plan for antibiotics, rational use of antibiotics and preventing development of antibiotic resistance.