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完整後設資料紀錄
DC 欄位 | 值 | 語言 |
---|---|---|
dc.contributor.advisor | 王錦堂(Jin-Town Wang) | |
dc.contributor.author | Ai-Lin Hsiao | en |
dc.contributor.author | 蕭艾琳 | zh_TW |
dc.date.accessioned | 2021-06-15T12:49:38Z | - |
dc.date.available | 2021-08-26 | |
dc.date.copyright | 2016-08-26 | |
dc.date.issued | 2016 | |
dc.date.submitted | 2016-07-21 | |
dc.identifier.citation | 1. Roberts, I.S., The biochemistry and genetics of capsular polysaccharide production in bacteria. Annual Review of Microbiology, 1996. 50: p. 285-315.
2. Edmondson, A.S. and E.M. Cooke, The development and assessment of a bacteriocin typing method for Klebsiella. J Hyg (Lond), 1979. 82(2) : p.207–223. 3. Hsu, C.R., et al., Isolation of a bacteriophage specific for a new capsular type of Klebsiella pneumoniae and characterization of its polysaccharide depolymerase. PLoS One, 2013. 8(8): p. e70092. 4. Pan, Y.J., et al., Capsular polysaccharide synthesis regions in Klebsiella pneumoniae serotype K57 and a new capsular serotype. J Clin Microbiol, 2008. 46(7): p. 2231-40. 5. Podschun R. and Ullmann U., Klebsiella spp. as Nosocomial Pathogens: Epidemiology, Taxonomy, Typing Methods, and Pathogenicity Factors. Clinical Microbiology Reviews, 1998. 11(4): p. 589-603. 6. Fang, C.T., et al., A novel virulence gene in Klebsiella pneumoniae strains causing primary liver abscess and septic metastatic complications. J Exp Med, 2004. 199(5): p. 697-705. 7. Tsai, Y.K., et al., Klebsiella pneumoniae outer membrane porins OmpK35 and OmpK36 play roles in both antimicrobial resistance and virulence. Antimicrob Agents Chemother, 2011. 55(4): p. 1485-93. 8. Tsai, F.C., et al., Pyogenic liver abscess as endemic disease, Taiwan. Emerg Infect Dis, 2008. 14(10): p. 1592-600. 9. Wang JH, L.Y., Lee SS, Yen MY, Chen YS, Wann SR, et al, Primary liver abscess due to Klebsiella pneumoniae in Taiwan. Clin Infect Dis, 1998. 26: p. 1434-8. 10. Ko W-C, P.D., Sagnimeni AJ, Hansen DS, Gottberg AV, Mohapatra S., Community-Acquired Klebsiella pneumoniae Bacteremia: Global Differences in Clinical Patterns. Emerg Infect Dis., 2002. 8(2): p.160-166. 11. Pen˜a, C., M. Pujol, C. Ardanuy, A. Ricart, R. Pallares, J. Lin˜ares, J. Ariza, and F. Gudiol. , Epidemiology and successful control of a large outbreak due to Klebsiella pneumoniae producing extended-spectrum b-lactamases. Antimicrob. Agents Chemother., 1998. 42(1): p.53-8. 12. Jacoby, G.A. and A.A. Medeiros, More extended-spectrum beta-lactamases. Antimicrob Agents Chemother. , 1991. 35(9): p.1697–1704. 13. Bush, K., G. A. Jacoby, and A. A. Medeiros., A functional classification scheme for b-lactamases and its correlation with molecular structure. Anti-microb. Agents Chemother, 1995. 39(6): p.1211-33. 14. Nordmann, P., L. Dortet, and L. Poirel, Carbapenem resistance in Enterobacteriaceae: here is the storm! Trends Mol Med, 2012. 18(5): p. 263-72. 15. Lee, N.Y., et al., Characterization of carbapenem-nonsusceptible Klebsiella pneumoniae bloodstream isolates at a Taiwanese hospital: clinical impacts of lowered breakpoints for carbapenems. Eur J Clin Microbiol Infect Dis, 2012. 31(8): p. 1941-50. 16. Lance R. Peterson, Bad bugs, no drugs: no ESCAPE revisited. Clin Infect Dis., 2009. 49(6): p.992-993. 17. Ambler R.P., The structure of beta-lactamases. Philos Trans R Soc Lond B Biol Sci. , 1980. 289(1036): p.321-31. 18. Bush K, J.G., Medeiros AA., A functional classification scheme for β-lactamases and its correlation with molecular structure. Antimicrob Agents Chemother., 1995. 39(6): p.1211-33. 19. Queenan, A.M. and K. Bush, Carbapenemases: the versatile beta-lactamases. Clin Microbiol Rev, 2007. 20(3): p. 440-58. 20. P., N., D. L., and P. L., Carbapenem resistance in Enterobacteriaceae: here is the storm! Trends Mol Med. , 2012. 18(5): p.263-72. 21. Shah, P.M. and R.D. Isaacs, Ertapenem, the first of a new group of carbapenems. J Antimicrob Chemother, 2003. 52(4): p. 538-42. 22. Cheng, N.-C., et al., In vitro activities of tigecycline, ertapenem, isepamicin, and other antimicrobial agents against clinically isolated organisms in Taiwan. Microbial Drug Resistance, 2005. 11(4): p.330-41. 23. Papp-Wallace, K.M., et al., Carbapenems: past, present, and future. Antimicrob Agents Chemother, 2011. 55(11): p. 4943-60. 24. Chow JW, S.D., Imipenem resistance associated with the loss of a 40-kDa outer membrane protein in Enterobacter aerogenes. J Antimicrob Chemother, 1991. 28 (4): p.499-504. 25. Munoz-Price, L.S. and J.P. Quinn, The spread of Klebsiella pneumoniae carbapenemases: a tale of strains, plasmids, and transposons. Clin Infect Dis, 2009. 49(11): p. 1739-41. 26. Doumith M, E.M., Livermore DM, Woodford N, Molecular mechanisms disrupting porin expression in ertapenem-resistant Klebsiella and Enterobacter spp. clinical isolates from the UK. J Antimicrob Chemother, 2009. 63(4): p.659-67. 27. Woodford N, D.J., Hill RL, Palepou MF, Pike R, et al. , Ertapenem resistance among Klebsiella and Enterobacter submitted in the UK to a reference laboratory. Int J Antimicrob Agents, 2007. 29(8): p. 3917–3921. 28. Patrice Nordmann, G.C., Thierry Naas, The real threat of Klebsiella pneumoniae carbapenemase-producing bacteria. Lancet Infect Dis, 2009. 9(4): p.228-36. 29. Scott K. Fridkin, C.D.S., et al., Surveillance of Antimicrobial Use and Antimicrobial Resistance in United States Hospitals: Project ICARE Phase 2. CID, 1999. 29(2): p.245-52. 30. Bradford, P., Urban C, Mariano N, Projan SJ, Rahal JJ, Bush K., Imipenem resistance in Klebsiella pneumoniae is associated with the combination of ACT-1, a plasmid-mediated AmpC b-lactamase, and the loss of an outer membrane protein. Antimicrob Agents Chemother 1997. 47(3): p.563-9. 31. MacKenzie FM, F.K., Dorai-John T, Amyes SG, Gould IM., Emergence of a carbapenem-resistant Klebsiella pneumoniae. Lancet 1997. 350(9080): p.783. 32. Bratu S, L.D., Haag R, et al., Rapid spread of carbapenem-resistant Klebsiella pneumoniae in New York City: a new threat to our antibiotic armamentarium. Arch Intern Med., 2005. 165(12): p.1430-5. 33. Schwaber MJ, K.-L.S., Navon-Venezia S, Schwartz D, Leavitt A, Carmeli Y, Predictors of carbapenem-resistant Klebsiella pneumoniae acquisition among hospitalized adults, and effect of acquisition on mortality. Antimicrob Agents Chemother, 2008. 52(3): p.1028-33. 34. Villegas MV, L.K., Correa A, et al., First detection of the plasmid-mediated class A carbapenemase KPC-2 in clinical isolates of Klebsiella pneumoniae from South America. Antimicrob Agents Chemother., 2006. 50(8): p.2880-2. 35. Wei ZQ, D.X., Yu YS, Shen P, Chen YG, Li LJ, Plasmid-mediated KPC-2 in a Klebsiella pneumoniae isolate from China. Antimicrob Agents Chemother, 2007. 51(3):711-4. 36. 產碳氫黴烯酶KPC腸桿菌之群聚事件之監測102年度研究報告. 衛生福利部疾病管制署, 2013. 37. Annual report of Taiwan Nosocomial Infection Surveillance Systems. Kaohsiung Medical University Hospital, Kaohsiung, Taiwan. Kaohsiung Medical University Hospital, 2014. 38. Wu, J.J., et al., Prevalence and characteristics of ertapenem-resistant Klebsiella pneumoniae isolates in a Taiwanese university hospital. Microb Drug Resist, 2011. 17(2): p. 259-66. 39. Patel, G., et al., Outcomes of carbapenem-resistant Klebsiella pneumoniae infection and the impact of antimicrobial and adjunctive therapies. Infect Control Hosp Epidemiol, 2008. 29(12): p. 1099-106. 40. Kanj, S.S. and Z.A. Kanafani, Current concepts in antimicrobial therapy against resistant gram-negative organisms: extended-spectrum beta-lactamase-producing Enterobacteriaceae, carbapenem-resistant Enterobacteriaceae, and multidrug-resistant Pseudomonas aeruginosa. Mayo Clin Proc, 2011. 86(3): p. 250-9. 41. Delcour, A.H., Outer membrane permeability and antibiotic resistance. Biochim Biophys Acta, 2009. 1794(5): p. 808-16. 42. Nikaido H., Molecular basis of bacterial outer membrane permeability revisited. Microbiol Mol Biol Rev, 2003. 67(4):593-656. 43. Ardanuy C, L.J., Domínguez MA, Hernández-Allés S, Benedí VJ, Martínez-Martínez L, Outer Membrane Profiles of Clonally Related Klebsiella pneumoniae Isolates from Clinical Samples and Activities of Cephalosporins and Carbapenems. Antimicrobial Agents And Chemotherapy, 1998. 42(7):1636-40. 44. A, D.-S., et al., Identification and characterization of a new porin gene of Klebsiella pneumoniae: its role in beta-lactam antibiotic resistance. JOURNAL OF BACTERIOLOGY, 1999. 181(9):2726-32. 45. Herna´ndez-Alle´s, S., S. Albertı´, X. Rubires, S. Merino, J. M. Toma´s, and V. J. Benedı´. Isolation of FC3-11, a bacteriophage specific for the Klebsiella pneumoniae porin OmpK36, and its use for the isolation of porin-deficient mutants. Can. J. Microbiol, 1995. 41(4-5): p. 399-406. 46. Albertı´, S., F. Rodrı´guez-Quin˜ones, T. Schirmer, G. Rummel, J. M. Toma´s, J. P. Rosenbusch, and V. J. Benedı´. A porin from Klebsiella pneumoniae: sequence homology, three-dimensional structure, and complement binding. Infect. Immun. , 1995. 63(3): p.903-10. 47. van der Ley, P., A. Bekkers, J. van Meersbergen, and J. A. Tommassen, Comparative study on the phoE genes of three enterobacterial species. Eur. J. Biochem., 1987. 164(2):469-75. 48. Werts, C., A. Charbit, S. Bachellier, and M. Hofnung., DNA sequence of the lamB gene from K. pneumoniae. Mol. Gen. Genet. , 1992. 233(3):372-8. 49. Garcia-Sureda, L., et al., Role of Klebsiella pneumoniae LamB Porin in antimicrobial resistance. Antimicrob Agents Chemother, 2011. 55(4): p. 1803-5. 50. Kaczmarek, F.M., et al., High-level carbapenem resistance in a Klebsiella pneumoniae clinical isolate is due to the combination of bla(ACT-1) beta-lactamase production, porin OmpK35/36 insertional inactivation, and down-regulation of the phosphate transport porin phoe. Antimicrob Agents Chemother, 2006. 50(10): p. 3396-406. 51. Achouak, W., T. Heulin, and J.-M. Pagès, Multiple facets of bacterial porins. FEMS Microbiology Letters, 2001. 199(1): p. 1-7. 52. STEVEN FORST, J.D., AND MASAYORI INOUYE, Phosphorylation of OmpR by the osmosensor EnvZ modulates expression of the ompF and ompC genes in Escherichia coli. Proc. Nati. Acad. Sci. USA, 1989. 86(16): p.6052-6. 53. Sleator, R.D. and C. Hill, Bacterial osmoadaptation: the role of osmolytes in bacterial stress and virulence. FEMS Microbiology Reviews, 2002. 26(1):49-71. 54. Feng, X., et al., OmpR Phosphorylation and Its Role in Signaling and Pathogenesis. American Society for Microbiology, 2003. 69(8): p. 390-395. 55. LA, E., P. H, and I. M., Signal transduction via the histidyl-aspartyl phosphorelay. Genes Cells, 1997. 2(3) : p.167-84. 56. Yang, F.C., et al., Characterization of ertapenem-resistant Enterobacter cloacae in a Taiwanese university hospital. J Clin Microbiol, 2012. 50(2): p. 223-6. 57. Lavigne, J.P., et al., An adaptive response of Enterobacter aerogenes to imipenem: regulation of porin balance in clinical isolates. Int J Antimicrob Agents, 2013. 41(2): p. 130-6. 58. Li, X.Z., P. Plesiat, and H. Nikaido, The challenge of efflux-mediated antibiotic resistance in Gram-negative bacteria. Clin Microbiol Rev, 2015. 28(2): p. 337-418. 59. Padilla, E., et al., Klebsiella pneumoniae AcrAB efflux pump contributes to antimicrobial resistance and virulence. Antimicrob Agents Chemother, 2010. 54(1): p. 177-83. 60. Martı´nez-Martı´nez, L., S. Herna´ndez-Alle´s, S. Abertı´, J. M. Toma´s, V. J. Benedı´, and G. A. Jacoby, In vivo selection of porin-deficient mutants of Klebsiella pneumoniae with increased resistance to cefoxitin and expanded-spectrum cephalosporins. Antimicrob. Agents Chemother., 1996. 40(2):342-8. 61. Doumith, M., et al., Molecular mechanisms disrupting porin expression in ertapenem-resistant Klebsiella and Enterobacter spp. clinical isolates from the UK. J Antimicrob Chemother, 2009. 63(4): p. 659-67. 62. Shakib, P., et al., Prevalence of OmpK35 and OmpK36 porin expression in beta-lactamase and non-betalactamase- producing Klebsiella pneumoniae. Biologics, 2012. 6: p. 1-4. 63. Domenech-Sanchez, A., et al., Role of Klebsiella pneumoniae OmpK35 Porin in Antimicrobial Resistance. Antimicrobial Agents and Chemotherapy, 2003. 47(10): p. 3332-3335. 64. Hernandez-Alles, S., et al., Porin expression in clinical isolates of Klebsiella pneurnoniae. Microbiology, 1999. 145: p. 673-679. 65. Crowley, B., Benedı´, V. J. & Dome´ nech-Sa´nchez, A., Expression of SHV-2 b-lactamase and of reduced amounts of OmpK36 porin in Klebsiella pneumoniae results in increased resistance to cephalosporins and carbapenems. Antimicrob Agents Chemother 2002. 46(11): p.3679-82. 66. Elliott, E., Brink, A. J., van Greune, J., Els, Z., Woodford, N., Turton, J., Warner, M. & Livermore, D. M., In vivo development of ertapenem resistance in a patient with pneumonia caused by Klebsiella pneumoniae with an extended-spectrum b-lactamase. Clin Infect Dis, 2006. 42(11): p.e95-8 67. Jacoby, G.A., Mills, D. M. & Chow, N. , Role of b-lactamases and porins in resistance to ertapenem and other b-lactams in Klebsiella pneumoniae. Antimicrob Agents Chemother 2004. 48(8):3203-6. 68. Netikul, T. and P. Kiratisin, Genetic Characterization of Carbapenem-Resistant Enterobacteriaceae and the Spread of Carbapenem-Resistant Klebsiella pneumonia ST340 at a University Hospital in Thailand. PLoS One, 2015. 10(9): p. e0139116. 69. Nikaido, H., Outer membrane barrier as a mechanism of antimicrobial resistance. Antimicrob. Agents Chemother., 1989. 33 (11):1831-6. 70. Chiu, S.K., et al., National surveillance study on carbapenem non-susceptible Klebsiella pneumoniae in Taiwan: the emergence and rapid dissemination of KPC-2 carbapenemase. PLoS One, 2013. 8(7): p. e69428. 71. Performance standards for antimicrobial susceptibility testing. Twenty-first informational supplement. CLSI document M100-S21. CLSI,Wayne, PA. Clinical and Laboratory Standards Institute (CLSI) 2014. 72. AJ, L., P. D, and C. GM, Methods for Generating Precise Deletions and Insertions in the Genome of Wild-Type Escherichia coli: Application to Open Reading Frame Characterization. JOURNAL OF BACTERIOLOGY, 1997. 179 (20): p.6228-37. 73. Zhang Y, J.X., Wang Y, Li G, Tian Y, Liu H, Ai F, Ma Y, Wang B, Ruan F, Rajakumar K., Contribution of β-lactamases and porin proteins OmpK35 and OmpK36 to carbapenem resistance in clinical isolates of KPC-2-producing Klebsiella pneumoniae. Antimicrob Agents Chemother, 2014. 58(2): p.1214-7. 74. Benz R, J.K., Lauger P, Ionic selectivity of pores formed by the matrix protein (porin) of Escherichia coli. Biochim Biophys Acta, 1979. 551(2): p.238-47. 75. Jacoby, G.A., AmpC beta-lactamases. Clin Microbiol Rev, 2009. 22(1): p. 161-82. 76. Pan, Y.J., et al., Identification of capsular types in carbapenem-resistant Klebsiella pneumoniae strains by wzc sequencing and implications for capsule depolymerase treatment. Antimicrob Agents Chemother, 2015. 59(2): p. 1038-47. 77. M, S., M. M, and T. J., Carboxy-terminal phenylalanine is essential for the correct assembly of a bacterial outer membrane protein. Journal of Molecular Biology, 1991. 218(1): p.141-8. 78. Pournaras S, M.M., Spanakis N, Ikonomidis A, Tassios PT, Tsakris A, Legakis NJ, Maniatis AN, Spread of efflux pump-overexpressing, non-metallo-beta-lactamase-producing, meropenem-resistant but ceftazidime-susceptible Pseudomonas aeruginosa in a region with blaVIM endemicity. J Antimicrob Chemother., 2005. 56(4):761-4. 79. Sun, J., Z. Deng, and A. Yan, Bacterial multidrug efflux pumps: mechanisms, physiology and pharmacological exploitations. Biochem Biophys Res Commun, 2014. 453(2): p. 254-67. 80. Garcia-Sureda, L., et al., OmpK26, a novel porin associated with carbapenem resistance in Klebsiella pneumoniae. Antimicrob Agents Chemother, 2011. 55(10): p. 4742-7. 81. Guillier, M., S. Gottesman, and G. Storz, Modulating the outer membrane with small RNAs. Genes Dev, 2006. 20(17): p. 2338-48. 82. L, E. and D. N, The regulatory RNA gene micF is present in several species of gram-negative bacteria and is phylogenetically conserved. Mol Microbiol. , 1994. 12(4): p.639-46. 83. Vogel, J. and K. Papenfort, Small non-coding RNAs and the bacterial outer membrane. Curr Opin Microbiol, 2006. 9(6): p. 605-11. 84. Wu, K.M., et al., Genome sequencing and comparative analysis of Klebsiella pneumoniae NTUH-K2044, a strain causing liver abscess and meningitis. J Bacteriol, 2009. 191(14): p. 4492-501. | |
dc.identifier.uri | http://tdr.lib.ntu.edu.tw/jspui/handle/123456789/50629 | - |
dc.description.abstract | 臨床目前使用碳氫黴烯作為治療超廣效乙內醯胺酶抗藥株感染的藥物,近年來卻陸續出現抗藥菌株的報導,其中腸桿菌科的克雷伯氏桿菌所占比例甚高,由於此類抗藥菌株限縮用藥選擇,必須仰賴副作用較劇的後線藥物,故尋找可能的抗藥機制,對於公共衛生政策而言極其重要。本篇研究承實驗室先前的研究,分析來自台灣四家教學醫院分離出的克雷伯氏肺炎桿菌之碳氫黴烯抗藥菌株,以聚合酶連鎖反應排除帶有碳氫黴烯酶基因者後,再深入探討革蘭氏陰性菌特有的外膜孔蛋白表現,研究主要孔蛋白OmpK35或OmpK36的缺失或突變是否會影響克雷伯氏肺炎桿菌對於碳氫黴烯類抗生素之感受性。首先,將完整的孔蛋白基因ompK35及ompK36選殖進入質體,並以電穿孔的轉形方式分別補回抗藥性菌株內,測試對於亞胺培南之最低抑菌濃度,發現有34.5%菌株的最低抑菌濃度降低和OmpK35、OmpK36孔蛋白的功能喪失相關,代表菌株孔蛋白的表現改變,或是基因突變或缺失造成功能不彰,進而影響克雷伯氏肺炎桿菌對碳氫黴烯的感受性。故進一步定序分析OmpK35和OmpK36孔蛋白的胺基酸序列,雖有41.2%的菌株胺基酸序列和對碳氫黴烯有感受性的參考菌株一致,但另有41.2%的菌株孔蛋白基因出現插入或單一鹼基剔除,造成移碼突變或提早轉錄終止,對照預測的結構,認為可能影響孔蛋白鑲嵌於外膜。為了解OmpK35和OmpK36孔蛋白實際的表現情形,以基因剔除的方式建構孔蛋白突變株做為對照組,且將外膜蛋白萃取出來觀察其表現量,由於蛋白染色無法依分子量準確區分出特定孔蛋白的位置,還須參照西方墨點法的結果交叉比對,結果顯示21.1%可偵測到OmpK35、52.6%可偵測到OmpK36。因而用反轉錄定量聚合酶連鎖反應分析其核酸表現量,儘管在高滲透壓環境下負責調控孔蛋白的ompR表現增加,但與抗藥性較不具關聯。最後,測定無標記基因置換突變株的抑菌濃度,證實OmpK35近羧端序列對碳氫黴烯抗藥性影響甚鉅。 | zh_TW |
dc.description.abstract | In recent decades, carbapenem-resistant Klebsiella pneumoniae (CRKPs) emerged rapidly due to the treatment of extended spectrum β-lactamse (ESBLs). To understand whether major outer membrane proteins, OmpK35 and OmpK36, played important roles in carbapenem resistance, we harvested non-carbapenemase-producing and imipenem-resistant K. pneumoniae isolates from four medical centers in Taiwan. In this study, the function of OmpK35 and OmpK36 from 19 of 55 isolates (34.5%) with high MIC (≥ 4μg/mL) was confirmed by using plasmid complementation tests, respectively. Moreover, amino acid sequence analysis revealed that 41.2% (7/19) frame-shift mutation of OmpK35 were caused by one-base pair deletion, whereas 41.2% (7/19) of clinical isolates carried the same sequence of OmpK35 as reference strains. To investigate whether clinical CRKPs were porin-lost or not, the outer membrane proteins of these strains were extracted by Western blots. We found that 21.1% of OmpK35 and 52.6% of OmpK36 were detected. In addition, we used real-time RT-PCR to analyze the transcriptional expression level of ompK35, ompK36 and their response regulator, ompR. The transcriptional expression of ompK35 was downregulated in low osmolality, and the expressions of ompK36 and ompR were upregulated in high osmolality environment. Finally, unmarked replacement OmpK35mutants restored their sensitivity against imipenem. In conclusion, there is no correlation between porins OmpK35 and OmpK36 expression and resistant to imipenem. And this study provides the evidence for importance of carboxyl terminus of OmpK35 in K. pneumoniae’s porin function. | en |
dc.description.provenance | Made available in DSpace on 2021-06-15T12:49:38Z (GMT). No. of bitstreams: 1 ntu-105-R03445105-1.pdf: 6976638 bytes, checksum: c24b0936e624d8618fa3e2abe6233570 (MD5) Previous issue date: 2016 | en |
dc.description.tableofcontents | 口試委員會審定書 I
誌謝 II 中文摘要 III 英文摘要 IV 目錄 V 表目錄 VII 圖目錄 VIII 第一章、 緒論 1 1.1 簡介克雷伯氏肺炎桿菌 1 1.2 抗乙內醯胺克雷伯氏肺炎桿菌的興起 2 1.3 抗碳氫黴烯克雷伯氏肺炎桿菌的出現 3 1.4 外膜孔蛋白簡介 5 1.5 外膜孔蛋白表現量之調控機制 6 1.6 抗藥性菌株和外膜孔蛋白缺失的關聯 7 1.7 研究動機 8 第二章、 材料與方法 10 2.1 材料 10 2.1.1 菌株與載體 10 2.1.2 培養基 10 2.1.3 抗生素 10 2.1.4 引子 (Primers) 11 2.2 方法 11 2.2.1 聚合酶鏈鎖反應 polymerase chain reaction (PCR) 11 2.2.2 萃取核醣核酸RNA Extraction 12 2.2.3 反轉錄定量聚合酶連鎖反應Reverse-transcription real-time PCR 13 2.2.4 補回試驗 Chromosomal complementation 14 2.2.5 建構無標記突變菌株unmarked mutation 17 2.2.6 製備勝任細胞 production of competent cells 20 2.2.7 電穿孔 electroporation 21 2.2.8 萃取外膜蛋白質 Outer membrane protein extraction 21 2.2.9 多株抗體製備 Polyclonal Antibody Production 23 2.2.10 西方點墨法Western blotting 25 2.2.11 蛋白質染色 Coomassie blue staining 28 2.2.12 定序分析 Sequencing Analysis 29 第三章、 結果 30 3.1 外膜孔蛋白 OmpK35 及 OmpK36補回試驗之抗藥性分析 30 3.2 CRKPs 之孔蛋白 OmpK35 和 OmpK36 基因序列分析 31 3.2.1 核苷酸序列比對 31 3.2.2 胺基酸序列比對 32 3.3 專一性檢視孔蛋白OmpK35 和 OmpK36之表現量 33 3.3.1 ompK35、ompK36基因剔除株之孔蛋白表現分析 33 3.3.2 臨床抗碳氫黴烯克雷伯氏肺炎桿菌之孔蛋白表現分析 34 3.4 偵測ompK35和ompK36與上游調控基因之核酸表現 35 3.5 無標記基因置換、剔除孔蛋白基因突變株對亞胺培南感受性的影響 36 3.5.1. 無標記基因剔除突變株無改變對亞胺培南的感受性 36 3.5.2. 無標記基因置換ompK35突變株恢復對亞胺培南的感受性 36 第四章、 總結與討論 38 第五章、 參考文獻 43 | |
dc.language.iso | zh-TW | |
dc.title | 探討克雷伯氏肺炎桿菌孔蛋白OmpK35和OmpK36與碳氫黴烯抗藥性之關聯 | zh_TW |
dc.title | The Role of Klebsiella pneumoniae porin OmpK35 and OmpK36 in Carbapenem Resistance | en |
dc.type | Thesis | |
dc.date.schoolyear | 104-2 | |
dc.description.degree | 碩士 | |
dc.contributor.oralexamcommittee | 周綠蘋(Lu-Ping Chow),董馨蓮(Shin-Lian Doong),蔡丰喬(Feng-Chiao Tsai) | |
dc.subject.keyword | 克雷伯氏肺炎桿菌,碳氫黴烯,抗碳氫黴烯克雷伯氏肺炎桿菌,外膜孔蛋白,抗藥機制, | zh_TW |
dc.subject.keyword | carbapenem-resistant Klebsiella pneumoniae,outer membrane protein,porin,OmpK35,OmpK36,carbapenam-resistance mechanism, | en |
dc.relation.page | 83 | |
dc.identifier.doi | 10.6342/NTU201600759 | |
dc.rights.note | 有償授權 | |
dc.date.accepted | 2016-07-21 | |
dc.contributor.author-college | 醫學院 | zh_TW |
dc.contributor.author-dept | 微生物學研究所 | zh_TW |
顯示於系所單位: | 微生物學科所 |
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