Research Distribution Of Transposons In Multidrug-Resistant ESBLs-producing Escherichia Coli

Objective:To explore the transposons distribution and drug resistant in multi-drug resistant ESBLs-producing E. Coil isolates.Method:1.Screening test:the presence of 71 ESBLs-producing strains including CTX、CAZ、 CS、AZT were determined by kerby-Bauer test according to the National Committee for clinical Laboratory Standards(NCCLS). If the diameter of inhibition of ceftazidime was smaller than 22mm, that of ceftriaxone was smaller than 25mm, and that of cefotaxime or aztreonam was smaller than 27mm, probably these strains produced ESBLs.2.Confirmatory test:The strains of ESBLs-producing E. coli matched preliminary results were detected by confirmatory test。 Suspect ESBLs-producing strains were determined by Disk diffusion test. Adding ceftazidime and ceftazidime/clavulanic acid, cefotaxime and cefotaxime/clavulanic acid in each group, observed diameter of inhibition zone of adding clavulanic acid was larger 5mm than not adding clavulanic acid.3. Disk diffusion test:The susceptibility of 71 strains of E. coil to 6 antibiotics were detected by disk diffusion which was recommended by the National Committee for Clinical Laboratory Standards (NCCLS). Using standard strains of Escherichia coli ATCC 25922 to control quality. The levofloxacin, streptomycin, cefoxitin, amikacin, cefotaxime and ciprofloxacin were detected respectively in 71 strains of Escherichia coli resistance.4. PCR Technology:Using Biospin bacterial genome DNA were extracted Escherichia coli DNA by the kit. According to the known transposon Tn21, Tn501, Tnl, Tn2, Tn3, Tn1000 Tn1548 were designed seven pairs of primers, E. coli DNA template were amplified by PCR from positive transposon genetic markers. The positive amplification length:Tn21 for 317 bp; Tn501 for 373 bp; Tnl for 497bp; Tn2 for 479bp; Tn3 for 323bp; Tn1000 for 372bp; Tn1548 for 497bp; The PCR products were subjected to agarose condensate gel electrophoresis, UV detector observed the results.5. Gene sequencing:to extract transposon DNA amplification-positive strains, PCR amplification products were measured by nucleic acid quantitative instrument to obtained the DNA concentration> 300ug/ml and sent to Shanghai Sangon Biotechnology Company to sequence. Results:1.ESBLs-producing strain:In 71 strains of Escherichiacoli,34(49.30%) ESBLs-producing strains were’confirmed by disk diffusion method confirmatory test.2. The resistance to 6 antibiotics in 71 strains of Escherichia coli:multi-resistant 45 (63.38%), the dual-resistance 8 (11.27%), single-resistant 11 (15.49%), the all-sensitive 7 (9.86%).3. In 71 strains of Escherichia coli, transposon Tn21, Tn501 are 15 strains.Transposon Tn1, Tn2, Tn3, Tn1000, Tn1548 was not detected.3.1 Distribution of Transposon Tn21, Tn501 in the various resistance patterns:13 strains isolates were multi-drug resistant transposon strains.8 strains isolates were double-resistant strains.1 strains isolates were single-resistant strains.7 strains sensitive resistant isolates were not detected in the presence of transposon. There was a statistically significant (X2= 4.56,p=0.032) in the whole group CMH; There was a statistically significant association (X2= 4.44 p=0.0350) between Multi-resistant and dual resistance+single resistance+ sensitive.3.2 Distribution of Transposon Tn21 and Tn501 in ESBLs-producing strains and non-ESBLs-producing strains:35 ESBLs-producing strains isolates were 11 strains.36 non-ESBLs-producing strains isolates were 4 strains; they were statistically significant (X2=4.33;?=0.0373).3.3.Distribution of resistance patterns of transposon Tn21, Tn501:Transposon Tn21 isolates was detected 7 strains, in which were 6 strains multi-resistant,0 strains dual-resistant,1 strains single-resistance; Transposon Tn501 isolates was detected 5 strains, in which were 4 strains muti-resistant; Tn21 and Tn501 overlapping are multi-resistant strains.3.4.Distribution of transposon Tn21 and Tn501 in ESBLs-producing strains and non-ESBLs-producing strains:Transposon Tn21 was detected 7 strains, including ESBLs-producing strains 7 strain (100%), non-ESBLs-producing enzyme 0; Transposon Tn501 was isolated 5 strains containing,1 of ESBLs-producing strains strains,4 of non-ESBLs-producing strains; Tn21 and Tn501 overlaping were 3 strains including ESBLs-producing strains (100%).3.5. Distribution of Transposon Tn21,Tn501 in resistance patterns:Contaning 9 strains multi-resistant,1 single-resistant, dual resistance and full sensitivity mode are not detected; Transposon Tn501 was detected 7 multi-resistant,1 strains double-resistant. 3.6. Distribution of transposon Tn501 Tn21 ESBLs-producing strains and non-ESBLs-producing strains:Transposon Tn21, the former was detected 10, the later was 0; Transposon Tn501, the former detected 8, the later was 4; (p= 0.0229). Conclusion:1.Transposon Tn21, Tn501 form are common, especially in the multi-drug resistant Escherichia pattern from clinical isolates of Escherichia coli in this region.2.Transposon strains was distributed into ESBLs-producing strains and non-ESBLs-producing strains, especially in the rate of detection ESBLs-producing strains is more than non ESBLs-producing strains. The transposon was probably related to ESBLs.3, Transposon TN21 Tn501 can be seen in the overlapping presence of multidrug-resistant Escherichia coli.