| Objective: 1. To investigate the drug-resistant pattern of Escherichia coliand Klebsiella isolated from our hospital and the genotype disttribution ofextended-spectrum β-lactamases (ESBLs) produced by Escherichia coli andKlebsiella. 2. to compare the 4 detection methods for ESBLs produced byEscherichia coli and Klebsiella and to construct an appropriate detection method.Methods: 1. 296 strains of nonrepetitive Escherichia coli and Klebsiella isolatedfrom our hospital from May 2004 to October 2004 were collected .2. two kinds ofdouble-disk synergy test , NCCLS phenotypic confirmatory test andthree-dimensional test were used to detect ESBLs in Escherichia coli andKlebsiella for experimental comparison. 3. ESBLs and AmpC enzyme weredetected by plasmid conjugation, isoelectric focusing test and three-dimensionaltest. 4. 78 strains of clinical isolates and 56 strains of transconjugants werecollected for in vitro susceptibility test using international standard plate dilutionmethod to assay the MIC50 and MIC90 as indicators representing theantibacterial activity of antimicrobial agents. The resistant rate (R%) ,intermediate rate ( I%) and susceptibility rate (S%) of drugs were calculatedaccording to the criteria in guidelines of NCCLS(2004). 5)the ESBLs genes fromtransconjugants and clinical isolates were amplified by PCR.Results1. 4 methods showed no difference for the detected ESBLs inEscherichia coli and Klebsiella(P>0.05). 2. The rates of ESBLs in Escherichiacoli and Klebsiella were 55.16% (123/223) and 50.68%(37/73)respectively. 3. 78ESBLs-produing Escherichia coli and Klebsiella isolates and 56 transconjugantswere susceptible to imipenem and meropenem and MIC was ≤0.5μg/m.Susceptibility rate of cefoxitin was 12.5%~ 66.7%,78.9%~91.9 respectively.Susceptibility rate of piperacillin/tazobactam was50%~80%, 78.9%~100%respectively. Resistance rates of 78 clinical isolates and 56 transconjugants toampicillin, piperacillin, cefazolin, ampicillin/sulbactum, cefuroximem,trimethoprim/sulfamethoxazole,gentamycin,cefoperazone were higher than 60%.4. Resistance (MIC≧64μg/ml) rates of 78 ESBLs-produing Escherichia coli andKlebsiella to ceftazidime were observed in 5 out of 78 strains,while resistance(MIC≧64μg/ml) rates of 78 ESBLs-produing Escherichia coli and Klebsiellato cefotaxime and ceftriaxone were 58%, 46%, 66% and 56% respectively. 5.Resistant rates of ESBLs-produing Escherichia coli and Klebsiella were ≥1.5times higher than those of non-ESBLs-produing strains. All tested strains howeverwere susceptible to imipenem and meropenem and resistant to ampicillin. 6.Resistance to extended-spectrum -lactams was successfully transferred from 56 of78 ESBL producers to E.coli. Among 56 transconjugants, production of ESBLswas inferred in all strains by three-dimensional test for ESBLs. 7. On IEF gels,the -lactamases with pIs of 7.8(8 strains)and 8.8(1 strain)were inhibited bycloxacillin but not by clavulanic acid, but other enzymes with pIs of 5.2(76strains), 8.1(58 strains), 8.4(17 strains), 8.2(3 strains), 8.3(1 strains) and 7.0 (5strains) -lactamase were inhibited by clavulanic acid but not by cloxacillin in 78clinical isolates.On IEF gels, -lactamases enzymes with pIs of 8.1(40 strains),8.4(16 strains) were inhibited by clavulanic acid but not by cloxacillin in 56transconjugants.8.Among 56 transconjugants and 22 clinical isolates, The blaTEMgenes were amplified from 57 isolates, The blaSHV genes were amplified from 6isolates, the blaCTX-M genes were amplified from 70 isolates, the blaCTX-M-1GROOPgenes were amplified from 18 isolates, the blaCTX-M-9 genes were amplified from53 isolates,1 isolate carried 2 types of ESBLs,producing CTX-M-9 groop andCTX-M-1 groop. blaCTX-M-2 genes, blaCTX-M-8 genes, blaOXA-2 genes and blaOXA-10... |
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