Identification and characterization of type 3 fimbrial adhesins of Klebsiella pneumoniae

• Main Authors: Wen-Ling Lu, 呂文鈴
• Other Authors: Hwan-You Chang
• Format: Others
• Language: en_US
• Published: 2001
• Online Access: http://ndltd.ncl.edu.tw/handle/03713213332827002508

碩士 === 國立清華大學 === 生命科學系 === 89 === Klebsiella pneumoniae is an important opportunistic pathogen that often causes urinary infection, pneumonia and septicemia in immunocompromised patients. In Taiwan, K. pneumoniae infection is closely associated with liver abscess among patients with diabetes mellitus, which may cause serious complications. As generally known, attachment of pathogens to their host is a prerequisite step of infection. In order to understand the pathogenesis of K. pneumoniae, I investigate the fimbriae and fimbrial adhesins of the clinical isolates of K. pneumoniae in Taiwan. About 100 isolates were analyzed by Dot blot hybridization and RFLP analysis. The results showed that, almost all the isolates analyzed possessed a gene relating to type 1 fimbrial adhesin. In addition to the type 3 fimbrial genes that have been reported previously, I identified several new adhesin genes, herein, namely mrkD-L1, 2, 3. Clones with the entire mrkD-L1 gene cluster were then isolated from the K. pneumoniae CG43 genomic DNA phage library. Sequence analysis showed that, the arrangement of the mrk genes was similar to that reported previously in another strain, except that the regulator, mrkE, was absent in K. pneumoniae CG43. The result of Southern hybridization indicated that, the mrk genes were chromosomally located by strain CG43. Mutations of the structural gene, mrkA, were subsequently generated. However, the effects of these mutants have not yet determined. Finally, the mrkD-L1 gene was over-expressed in E. coli, and the recombinant protein was purified and used to immunize mice. The production of MrkD-L1 specific antibody in mice has been confirmed by western analysis and ELISA. No protection was observed when the MrkD-L1 immunized mice were challenged with a lethal dose of K. pneumoniae CG43. However, lack of protection can not rule out the importance of type 3 fimbrial adhesin in K. pneumoniae pathogenesis. Abstract -------------------------------------------------------------- 1 Introduction --------------------------------------------------------- 5 Material and Methods -------------------------------------------- 10 Results ------------------------------------------------------------- 19 Discussion --------------------------------------------------------- 25 Reference ---------------------------------------------------------- 28 Figures ------------------------------------------------------------- 35