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Abstract
Antibacterial drugs are one of the most important therapeutic agents of bacterial infections but multidrug resistant Escherichia coli (MDREC) is an increasing problem worldwide. Major resistance mechanism of MDREC is horizontal gene transfer of R plasmids harboring integrons, which the integron integrase (IntI) catalyzes gene cassette insertion and excision through site specific recombination. In this study, resistance profiles of integron harboring E. coli isolated in Korea and the genetic environments of integron gene cassettes were analyzed by PCR and direct sequencing to clarify the mechanisms of spread of integron harboring E. coli. Resistance rates of integron harboring E. coli, including β-lactams, aminoglycosides, and fluoroquinolones and MDR frequencies were significantly higher than that of E. coli without integron (p<0.01). Majority (80%) of integron harboring E. coli showed resistance transfer by conjugation. Most (80%) of E. coli had dfrA17-aadA5 cassette array and PcH1 hybrid promoter; 16.7% of E. coli had dfrA12-orfF-aadA2 cassette array and PcW promoter. The higher prevalence of weak Pc variants among most (96.7%) of integron harboring MDREC suggests that a flexible cassette array is more important than enhanced expression. All the integrons had LexA binding motif suggests that SOS responses control the expression of these integrons. In conclusion, the genetic bases of integrons were diverse, and the spread and the expression of prevalent gene cassette arrays may be deeply related with strengths of Pc promoters in integrons. These informations will provide important knowledge to control the increase of integron harboring MDREC.
Figures and Tables
Fig. 1
Incidences of multiple resistance to increasing numbers of antimicrobial agents in integron-positive (▒) and integron-negative (■) Escherichia coli isolated in Korea. Total 117 isolates (30 integron-positive and 87 integron-negative isolates) we re tested.
Fig. 2
Schematic representation of integrons from multidrug resistant Escherichia coli isolated in Korea. A, Integron (n=1 isolate) with aadA2 gene cassette. B, Integrons (n=24 isolates) with dfrA17 and aadA5 gene cassettes. C, Integrons (n=5 isolates) with dfrA12, orfF and aadA2 gene cassettes. 5'-CS, 5'-conserved segment. 3'-CS, 3'-c onserved segment. intI1, integron integrase gene. LexA box, LexA repressor binding motif. attI, site specific recombination region of integron. attC, site specific recombin ation region of gene cassette. qacEdelta, quarternary ammonium compound efflux gene. sulI, sulfonamide resistance gene.
Fig. 3
The DNA sequences of class 1 integrons and their gene cassettes from multidrug resistant Escherichia coli isolated in Korea. A, Integron with aadA2 gene cassette containing the aadA2 aminoglycoside adenyltransferase gene and its attC. B, Integron with dfrA17 gene cassette containing the dfrA17 dihydrofolate reductase gene and its attC, and aadA5 gene cassette containing the aadA5 aminoglycoside adenyltransferase gene and its attC. C, Integron with dfrA12 gene cassette containing the dfrA12 dihydrofolate reductase gene and its attC, orfF gene cassette and its attC, and aadA2 gene cassette containing the aadA2 aminoglycoside adenyltransferase gene and its attC. The putative initiation codons and stop codons are in italics. R", L", L' and R' indicate imperfect inverted repeat sequences (underlined). RBS, ribosomal binding site.
Table 2
Antibiotic susceptibility of integron-positive and integron-negative Escherichia coli isolated in Korea
Table 4
DNA sequencesa associated with site specific recombination of integron gene cassettes from Escherichia coli isolated in Korea
a, All the DNA sequences are bottom strand sequences of 3'→5' direction; b, Inverted repeat sequences R" pairs with R', and L" pairs with L' to form R and L boxes, respecti vely, and DNA folding occurs within the sequences between L" and L' to form stem loop: c, EHB, Extra helical bases generated in the folded attC site; d, VTS, Variable term inal structure of stem loop.
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