| Objective:To investigate the distribution and drug-resistant features of Escherichia coli carrying TEM beta lactamases,which were isolated from the patients in Affiliated Hospital of Lu Zhou Medical College,and to discuss the role of TEM beta lactamases in E.coli.Methods : 1.Polymerase Chain Reaction(PCR)technique :Firstly,71 strains E.coli were cultured,then the plasmid DNA was extracted according to specification,and designing the public primer of bla TEM.The refined DNA template were used to PCR.The PCR Amplified products were visualized after electrophoresis on agarose gel,and photographed with gelatin camera.Secondly,the primer of TEM-1,TEM-2,TEM-116,TEM-151,TEM-135 were designed.The detection of TEM?-lactamase-encoding genes was carried out using PCR with primers that correspond to conserved regions of several bla TEM-type genes.The amplicons were visualized after electrophoresis on agarose gel.Electrophoresis results were observed by Ultraviolet detector,and photographed with gelatin camera.Thirdly,the positive gene amplification products were sequenced.2.Disk diffusion test:Antibiotic susceptibility of the E.coli was performed by the disk diffusion technique.Piperacillin(PRL),piperacillin-tazobactam(TZP),cefoperazone(CPZ),cefoperazone-Sulbactam(SCF),cefotaxime(CTX),cefotaxime-clavulanic acid(CDO3),cefradine(CE),cefepime(FEP),aztreonam(ATM),latamoxef(MOX),cefoxitin(FOX),imipenem(IPM),Gentamicin(GEN),kanamycin(K),ciproflox-acin(CIP),norfloxacin(NOR)were choosen?Quality control was performed as recommended using the E.coli strain ATCC 25922.Results :1.The stains carrying TEM beta lactamases: The total detection rate of E.coli carrying TEM beta lactamases was63.38%(45/71),including that TEM-1 beta lactamases was 37.78%(17/45),TEM-2 was 28.89%(13/45),TEM-151 was 11.11%(5/45),TEM-116 was15.56%(7/45),TEM-135 was 6.67%(3/45).2.The drug-resistances of the 16 antibiotics to the E.coli carrying TEM beta lactamases:1)All of 45 strains were sensitive to imipenem(IPM)? The drug-resistance rates of piperacillin(PRL),cefoperazone(CPZ),Cefotaxime(CTX)were highest,accounted for 80%.2)the analysis resistance patterns: There were 22(48.89%)multi-drug resistant strains,13(28.89%)double-resistant strains and 10(22.22%)one-resistant strains among 45 E.coli strains carrying TEM beta lactamases.The resistance rate of multi-drug resistant strains was significantly higher than those of double-resistant and one-resistant strains for most antibiotics.To beta lactamase inhibitors,the drug resistance were piperacillin(PRL)33 strains(86.84%),piperacillintazobactam(TZP)2 strains(5.26%),cefoperazone(CPZ)32 strains(84.21%),cefoperazone-sulbactam(SCF)3 strains(7.89%),cefotaxime(CTX)33 strains(86.84%),cefotaxime-Clavulanic acid(CDO3)5 strains(13.15%).The resistance differences of 6 antibiotics were compared by Chi-square test,P<0.05,the difference was satistically significant;3.The distribution and drug resistances of TEM beta lactamases : 1)The distribution of TEM beta lactamases:TEM-1 and TEM-2 beta lactamases was uniformly distributed in one-resistant,double-resistant and multi-drug resistant E.coli,while TEM-151,TEM-116 and TEM-135 beta lactamases were mainly distributed in double-resistant and multi-drug resistant E.coli.2)Drug-resistance of TEM beta lactamases: 2.1)The resistance differences were compared between TEM-2 and TEM-1 beta lactamases by Chi-square test,the differences was not satistically significant(P>0.05).2.2)For the beta lactamase inhibitors,The resistance differences were compared between TEM-1 and TEM-151 beta lactamases by Chi-square test,the differences was satistically significant(P<0.05);2.3)The drug resistances of TEM-116 and TEM-1 beta lactamases:This experiment was checking out 7 strains(15.56%)E.coli coutains TEM-116 by PCR and sequences.The resistance differences were compared between TEM-116 and TEM-1 beta lactamases by Chi-square test.The resistance differences of Gentamicin(GEN),ciprofloxacin(CIP),norfloxacin(NOR),Cefepime(FEP)were satistically significant(P < 0.05).The resistance differences of other antibiotic were not satistically significant,just that the resistance rates of TEM-116 were higher than TEM-1 for all antibiotic;2.4)The drug-resistances of TEM-135:3 strains(6.67%)had been checking out in this experiment,which was absolutely resistant to cephalosporins and fluoroquinolones.2.5)The drug resistances of TEM-1/TEM-2 and TEM-116/TEM-151/ TEM-135 in multi-drug E.coli:11 strains had been checking out in TEM-1 and TEM-2,and 13 strains had been checking out in TEM-116,TEM-151 and TEM-135.The resistance difference of Gentamicin(GEN),ciprofloxacin(CIP),cefoxitin(FOX)were satistically significant(P<0.05).Conclusion:TEM beta lactamases genotypes occurrs in E.coil in LuZhou.In this experiment,TEM-1 and TEM-2 can be found mostly,at the same time,the TEM-116?TEM-151 and TEM-135 also be found.Using Disk diffusion test and Statistical analysis,the drug resistance rates of TEM-116 ? TEM-151 and TEM-135 were higher than TEM-1 and TEM-2 obviously.The drug resistance rate of multi-drug resistant E.coli was higher than double-resistant and one-resistant E.coli. |
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