||Clostridium difficile infection: hospital-based epidemiology in Tainan and establishment of an animal model
||Institute of Clinical Medicine
Clostridium difficile infection
proton pump inhibitor
已知困難梭狀桿菌毒素會破壞腸壁，引起腸黏膜發炎及腸道破壞。除細菌毒素外，宿主免疫力缺失，也是困難梭狀桿菌感染增加的原因，因此宿主免疫防衛機制，也需要進一步研究。巨噬細胞如何辨識或吞噬困難梭狀桿菌的相關機制，仍未完全了解。Toll-like receptors (TLRs) 是先天性免疫重要組成，將研究TLRs 與困難梭狀桿菌關聯性。
獲得困難梭狀桿菌移生及感染的流行病學資料，將針對內科病房住院病患，以培養及毒素ELISA檢查糞便是否有困難梭狀桿菌移生。這些病人住院期間將持續追蹤，是否有發生困難梭狀桿菌感染的腸道炎及大腸炎。糞便中困難梭狀桿菌，將進一步做toxinotyping、 ribotyping。研究發現病患有困難梭狀桿菌移生，會有較高機會後續發生困難梭狀桿菌感染。氫離子阻斷劑和抗生素使用，都是困難梭狀桿菌移生或感染危險因子。另也證實台灣本土有高毒性困難梭狀桿菌菌株- 核糖核酸體分型126和027- 能造成嚴重感染；還發現特定TLRs基因多型性，與發生困難梭狀桿菌移生或感染有關。
為瞭解困難梭狀桿菌感染病理機制，將建立老鼠腸道感染模型。臨床上氫離子幫浦抑制劑使用，可能促成困難梭狀桿菌感染，藉由老鼠腸道感染模型，檢視氫離子幫浦抑制劑之角色與機轉，發現先前使用氫離子阻斷劑，將使老鼠的困難梭狀桿菌感染變得更嚴重。進一步以此老鼠模式，作為致病機制研究平台，探討TLR扮演的角色，用TLR2或 TLR4基因缺失老鼠，比較野生種老鼠及TLR2或 TLR4缺失老鼠，感染後腸道病理變化及發炎情況差異，結果發現TLR2或TLR4缺失老鼠發炎情形較嚴重, 因此推論TLRs在困難梭狀桿菌感染中，扮演保護性角色。
Clostridium difficile is a major cause of nosocomial antibiotic-associated diarrhea and has increasing incidence and fatality rates. The incidence of C. difficile infection (CDI) in Taiwan is unclear. C. difficile colonization is an independent risk factor for C. difficile-associated diarrhea. However, the impact of C. difficile colonization in hospitalized patients is not known. Gaps in our understanding of the pathogenesis of CDI present major challenges to the development of better preventive and therapeutic strategies against this problem.
C. difficile toxins breach the intestinal barrier and trigger musical inflammation and intestinal damage. In addition to the effects of bacterial toxins, impaired host immunity is also associated with increased risks of CDI, suggesting that the pathogenesis of CDI needs to be further characterized. The receptor-mediated mechanisms by which macrophages recognize and phagocytize bacteria, including clostridia, have not been well analyzed. Toll-like receptors (TLRs) are the major components of innate immunity. The relationship between TLRs and CDI was analyzed in this work.
An epidemiological study was performed in the medical wards of a regional hospital. Fecal culture and toxin ELISA tests were performed for C. difficile colonization in all hospitalized patients; during hospitalization, these patients were followed up for the development of CDI. All clinical C. difficile isolates were further characterized by toxinotyping and ribotyping. C. difficile colonization was noted to cause a higher risk of CDI. Prior exposure to proton pump inhibitors or antimicrobial agents was a risk factor for C. difficile colonization and infection. During the study period, some hypervirulent C. difficile strains, such as ribotype 126 and 027, were discovered in Tainan. Patients with a specific TLR polymorphism had a higher risk of C. difficile colonization and infection than those without the polymorphism.
To analyze the pathophysiology of CDI, a mouse model of CDI was designed to investigate the predisposing role of PPI in CDI. Mice developed severe CDI after PPI use. The mouse model of CDI could be used to analyze the contribution of innate immunity in CDI. To investigate the role of TLRs in the pathogenesis of CDI, Tlr2-/- or Tlr4-/- mice were infected by C. difficile. The mice then were examined for changes in the severity in clostridial colitis. Colon pathology findings in C. difficile-infected wild mice and Tlr2-/- or Tlr4-/- mice were compared. Tlr-/- mice presented more severe CDI than wild mice, indicative of a protective role of TLRs in CDI.
The first epidemiological information of C. difficile colonization and infection rates in Taiwan were obtained in the described clinical studies. In addition, basic research findings increased our understanding of TLRs and how they mediate the inflammatory responses against CDI. By combining these results, translational studies will likely draw the attention of medical professionals who deal with CDI. Furthermore, the described CDI mouse model can be utilized to develop preventive or therapeutic strategies for combating CDI.
English summary Ⅱ
Table legends Ⅶ
Figure legends Ⅷ
Chapter 1. Introduction 1
Chapter 2. The epidemiology and clinical impact of Clostridium difficile colonization and infection 4
Materials and methods 7
Chapter 3. Establishment of a mouse model of Clostridium difficile infection to investigate the role of a proton pump inhibitor and Toll-like receptors 25
Materials and methods 27
Chapter 4. Summary and suggestions for future research 41
Clostridium difficile study- related publications 43
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