碩士 / 國立成功大學 / 生物科技研究所碩博士班 / 95 / Aeromonas hydrophila is a gram-negative bacterium which can cause disease in wide variety of waterborne animals, including reptiles, amphibians, and fish. It is also an emerging human pathogens that cause a wide array of diseases, such as gastroenteritis, wound infections, and septicemia. Since two-component systems are assumed to be one of the prime bacterial systems and likely play a major role in bacterial virulence, one of our purpose is to investigate whether one of two-component systems, CpxA and CpxR, in A. hydrophila is involved in the pathogenesis of this bacterium or not. Previous studies reported that CpxA is a histidine kinase sensor protein and CpxR is a response regulatory protein and CpxA/R system controls transcription of the dsbA locus and activation of the Cpx pathway in 5- to 10-fold the synthesis of DsbA(disulfide bond formation protein A) in Escherichia coli. Since DsbA is a oxidative catalyst in the periplasm and it affects disulfide bond formation of secretory and periplasmic proteins and since A. hydrophila produces several extracellular proteins whose disulfate-bond formation may be catalyzed by DsbA and these extracellular proteins may act as the virulence factors. In this study, we constructed cpxA isogenic mutant and cpxR isogenic mutant by homologous recombination, and determined the properties of these two isogenic mutant. In our data, no significantly difference in DTT(dithiothreitol) assay, extracellular proteaolytic and lipolytic activities between two isogenic mutant and wild type strain was observed. However, both the two isogenic mutant exhibited significantly difference lower survival than the wild-type strain when incubated for 15 and 30 min in LB broth containing 20 µg/ml polymyxin B. This findings suggest that the CpxA/R system may be required for resistant to polymyxin B. After incubating with 75% human serum for 30 min, the survival rates of both isogenic mutant strains were shown to be significantly lower than wild type strain. The appearance in human serum-resistant assay match with polymyxin B assay was observed.We suggest that CpxA/R system could be make the response in undergoing stress at early stage. However, the value LD50 of mice infected by the two isogenic mutant via intraperitioneal was not significantly difference with wild type. Finally, we suggest that cpx two-component system was played a less important role in pathogenesis of Aeromonas hydrophila. However, expression of DsbA did not influenced by interruption of cpx in Aeromonas hydrophila.
| Identifer | oai:union.ndltd.org:TW/095NCKU5111012 |
| Date | January 2007 |
| Creators | Chu-yang Hsieh, 謝主揚 |
| Contributors | Ming-chung Chang, 張敏政 |
| Source Sets | National Digital Library of Theses and Dissertations in Taiwan |
| Language | zh-TW |
| Detected Language | English |
| Type | 學位論文 ; thesis |
| Format | 88 |
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