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標題: 豬霍亂沙門氏桿菌cAMP受體蛋白對第三型分泌系統之功能性調控作用
Effect of cAMP Receptor Protein on Regulation of Type III Secretion System of Salmonella enterica Serotype Choleraesuis
作者: 陳正文
Chen, Zeng-Weng
關鍵字: Salmonella
Type Three Secretion System
cAMP receptor protein
cAMP receptor protein
出版社: 微生物暨公共衛生學研究所
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摘要: 豬沙門氏桿菌症(salmonellosis)是豬隻重要細菌性疾病之一。眾多沙門氏桿菌血清型中,豬霍亂沙門氏桿菌(Salmonella Choleraesuis)因會造成豬隻敗血症而受到重視。沙門氏桿菌症的預防上,活毒減毒疫苗是預防此疾病最有效的方法之一。根據文獻得知,具有cAMP受體蛋白(cAMP receptor protein; CRP)基因突變的豬霍亂沙門氏桿菌,不僅呈現高度減毒且可引起宿主良好免疫反應的菌株,該菌株常被應用在沙門氏桿菌疫苗的開發上,然而其毒力減弱的機轉仍尚未被瞭解。因此本研究之目的即是構築一豬霍亂沙門氏桿菌crp突變株,並探討crp突變株的減毒機制。研究結果顯示,豬霍亂沙門氏桿菌crp基因剔除後,突變菌株對於酸性環境及膽鹽,反而較親代株具有更高的耐受性,顯示此兩種殺菌條件與crp突變株的毒力減弱無關。而以豬隻ligated ileal loop動物實驗模式評估細菌入侵能力之結果發現,crp突變株在迴腸上皮及腸繫膜淋巴結的入侵能力,卻明顯較親代株降低。此外,沙門氏桿菌的腸道入侵與第三型分泌系統 (Salmonella pathogenicity island 1 (SPI-1) encoded type III secretion system; SPI-1 T3SS)有密切關係,因此本研究亦針對SPI-1 T3SS的分泌功能進行檢驗。結果顯示,crp突變株之SPI-1 T3SS無法正常分泌與細菌入侵有關的SopB (Salmonell outer protein B)分泌蛋白,且與巨噬細胞死亡(pyrotosis)有關SipB (Salmonella invasion protein B)蛋白之分泌,亦同時受到抑制。以上研究結果顯示,豬霍亂沙門氏桿菌crp基因突變後,SPI-1 T3SS之分泌功能會受到抑制,而相關所造成的腸道入侵能力減低及巨噬細胞死亡的減少,是豬霍亂沙門氏桿菌crp突變株毒力減弱的原因之一。
Salmonella enterica serovar Choleraesuis (S. Choleraesuis) causes a lethal systemic infection (salmonellosis) in swine. Live attenuated S. Choleraesuis vaccines are effective in preventing the disease, and isolates of S. Choleraesuis with mutations in the cAMP receptor protein (CRP) gene (S. Choleraesuis ∆crp) are the most widely used, although the basis of the attenuation remains unclear. The objectives of this study were to construct a S. Choleraesuis crp gene mutant and to determine whether the attenuated phenotype of S. Choleraesuis ∆crp was due to alterations in susceptibility to gastrointestinal factors such as pH and bile salts, ability to colonize or invade the intestine, or cytotoxicity for macrophages. The results indicated that, although the survival of S. Choleraesuis ∆crp at low pH or in the presence of bile salts did not differ significantly from the parental strain, the ability of the mutant to invade intestinal epithelia was significantly decreased. In examining the role of CRP on the secretory function of the Salmonella pathogenicity island 1 (SPI-1) encoded type III secretion system (T3SS), it was found that S. Choleraesuis ∆crp was unable to secrete the SPI-1 T3SS effector proteins, SopB and SipB, which play an important role in Salmonella intestinal invasiveness and macrophage cytotoxicity, respectively. In addition, caspase-1 dependent cytotoxicity for macrophages was significantly reduced in S. Choleraesuis ∆crp. Collectively, this study demonstrated that the CRP affects the secretory function of SPI-1 T3SS resulting in reduced ability to invade host intestinal epithelium that is a critical element in the pathogenesis of S. Choleraesuis.
其他識別: U0005-1901201111573600
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