| Summary: | 碩士 === 國立嘉義大學 === 生物科技研究所 === 94 === Because of the improper use of antibiotics, multiple antibiotic resistant Enterobacteriaceae are becoming more common and causing severe problems in treatment of infection diseases. The common clinically antibiotic resistant bacteria include Escherichia coli, Mycobacterium tuberculosis, Klebsiella pneumoniae and Salmonella spp. Bacteria have developed the resistance mechanisms to neutralize the antibiotic action. Although β-lactam drugs have been used to cure the bacterial infection, bacteria can produce β-lactamases which hydrolyze theβ-lactam drugs, rendering them inactive. Recent, the genotypes and kinds of β-lactamases increase dramatically. The most popular extended spectrumβ-lactamases (ESBLs) are class A and class C-type β-lactamases. The purposes of this study were to determine the dissemination mechanism and the distribution of conjugatable class A and class C-type bla plasmids in Enterobacteriaceae. Including 4 K. pneumoniae, 23 Salmonella and 53 E. coli strains isolated from Chang Gung Memorial Hospital （CGMH） between 1999 to 2004. Plasmid size and number of all isolates were determined by alkaline miniprep method. Antibiotic susceptibility of all isolates was determined. The strains carrying the genes forβ-lactamase were determined by PCR method. Plasmid-encoding bla was determined by Southern blot and PCR analysis. The results showed that blaCMY, blaCTX-M14, blaCTX-M3, blaSHV and blaTEM were found in 39%, 8.6%, 17.4%, 4% and 22% of Salmonella strains individually. To determine whether bla- encoded plasmid were capable of conjugation, the purified plasmid DNA were transformated into E. coli DH5α, and transformants were used as donor and mated with Salmonella Typhimurium LBNP4417. The plasmid profile and genes blas of transconjugants were analyzed. The Salmonella S12 strain encodes three bla genes and consists of two big plasmids, including blaCMY encoded by 90 kb conjugatable plasmid and the blaTEM and blaCTX-M3 encoded by 85 kb plasmid. All 4 K. pneumoniae strains carry blaCTX-M3, blaSHV and blaTEM. The blaSHV was located in 120 kb plasmid and chromosome. The blaTEM and blaCTX-M3 were located in 85 kb plasmid simultaneously. And the K. pneumonia KP14 strains carried 95 kb blaDHA-1- encoded plasmid. The percentage of blaCMY, blaCTX-M14, blaCTX-M3 and blaTEM were 18.9%, 5.7%, 7.5% and 69.8% individually. The E. coil E45, E49 strains carried 85 kb blaCMY - encoded conjugatable plasmid. The E. coil pE7T-1, pE23T-1, pE30T-1 strains carried 90 kb blaCMY - encoded conjugatable plasmid. The E. coil pE3T-1, pE8T-3 and pE20T-3 strains carried 100 kb blaTEM - encoded conjugatable plasmid. The E. coil pE20T-1 and E21 strains carried 85 kb blaCTX-M3-encoded conjugatable plasmid. The increased genotype of β-lactamases produced from Enterobacteriacea and spread of such resistance genes by plasmid conjugation play an important threat to cure the bacterial infection. Therefore, surveillance of multiple antibiotic resistance strains carrying the popular class A and class C β-lactamases will assist us to assess the spread of such genes and prevent their infection.
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