| Summary: | 碩士 === 國立陽明大學 === 醫學生物技術暨檢驗學系暨研究所 === 98 === Our laboratory previously generated the imipenem-resistant strain ab1254R from the imipenem-susceptible strain ab1254S to study the mechanisms involved in the imipenem resistance of Acinetobacter balumannii. Comparism of the outer membrane proteins(OMPs) profiles gave two protein spots between ab1254S strain and ab1254R strain using two-dimensional gel electrophoresis(2-DE), that were significantly increased in strain ab1254R. Mass spectrometry analyses of these protein spots showed CarO and Omp25. Since these two proteins are very similar in physicochemical parameters (Mw and pI). The separation of these two proteins was very difficult even though using 2-DE analysis. However, it was reported that the clinical imipenem resistant strains of A. baumannii were associated with the loss of an outer membrane protein CarO, which may allow the influx of carbapenem antibiotics. Therefore, we decided to generate the Omp25 recombinant protein for antibody production for western blotting analysis to determine the protein expression level of Omp25 between strains ab1254S and ab1254R or between imipenem-susceptible and -resistant clinical strains. In contrast, several protein spots in 2-DE showed down-regulation in ab1254R. Of these proteins, the mRNA level of the putative ferric siderophore receptor protein I and II, OstA, CsuD, the putative DcaP-like and the putative OprB-like in ab1254R were lower than that of ab1254S. The putative ferric siderophore receptor protein II is almost 50% reduction. Whether these proteins associated with imipenem resistance in A. baumannii or not need furthermore investigation.
In addition to OXA-58, our laboratory identified another β-lactamase, IMP-1 present in Acinetobacter genomic species 3. Analysis the bacterial periplasmic fraction using SDS-PAGE, a protein band about 27kDa was significantly increased in imipenem-resistant strains. This protein was identified using 2-DE and mass spectrometry. We also showed that IMP-1, rather than OXA-58, may contribute to imipenem resistance of genomic species 3.
To investigate the additional mechanisms involved in the imipenem resistance. Comparism of outer membrane protein profiles between imipenem-susceptible and imipenem-resistant of Acinetobacter genomic species 3, YaeT, Catalase hydroperoxidase II (HPII) and putative long-chain fatty acid transport protein(FadL) were up-regulated in imipenem-resistant strains. Furthermore, the mRNA level of fadL gene was significantly increased in imipenem-resistant strains. However, the mRNA levels of yaeT and katE showed no difference between susceptible strains and resistant strains. Thus, how FadL influences an imipenem resistance of genomic species 3 needs further investigation.
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