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標題: Construction and characterization of capsule gene knockout mutants of Avibacterium paragallinarum
作者: 杜姿億
Tzu-Yi Tu
關鍵字: 雞傳染性鼻炎菌
Avibacterium paragallinarum
gene knockout
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摘要: Infectious coryza (IC) is an acute upper-respiratory disease associated with growth retardation and reduced egg production in chicken. This disease caused by infection with Avibacterium paragallinarum (Av. paragallinarum). Infectious coryza is distributed worldwide and causes significant economic loss of poultry industry. Av. paragallinarum is a Gram negative, facultatively anaerobic, rod shaped bacteria in the family of Pasteurellaceae. Capsule is located on the surface of bacteria and also one of the most important virulence factors of pathogenic bacteria. Capsule can prevent phagocytosis from host and defeat cell-mediated cytotoxicity. The hctABCD gene cluster encodes proteins that form an export complex of capsule polysaccharides, and it has high conserved sequence in different serotypes of Av. paragallinarum. In this study we used mobile group II intron (Targetron) to construct the hctA mutant strain. The hctA gene encodes ATP-binding protein component for polysaccharides export. We found that the amout of polysaccharides produced by mutant strain is less than the wild type strain. The mutant strains had a great ability than the wild type to form biofilms and to adhere to DF-1 cell. Taken together, there results showed that capsule from Av. paragallinarum is important for adherence and establishment of infection. On the other hand, the formation of capsule can prevent bacteria from cell-medated cytotoxicity and assist the spread of bacteria in the host. This discovery could be valuable for understanding the pathogenesis of Av. paragallinarum.
雞傳染性鼻炎菌 (Avibacterium paragallinarum, Av. paragallinarum) 為引起雞傳染性可利查 (Infectious coryza, IC) 之病原菌,此病又可稱作雞傳染性鼻炎,感染雞隻造成上呼吸道疾病,此疾病造成肉雞淘汰率增加,蛋雞產蛋功能下降,因此造成經濟上嚴重的損失。雞傳染性鼻炎菌為革蘭氏陰性菌,有許多重要的毒力因子,位在細胞最外側的莢膜多醣體結構具有保護菌體免於被宿主吞噬、抵抗補體媒介毒殺等功能,因此被視為重要的毒力因子。莢膜生合成基因包含了hctABCD 四個基因,具有負責運輸多醣體至細胞表面與組裝功能基因,此基因序列在不同血清型具有高度保留性。本研究使用商業化套組 Targetron 以 mobile group II intron 嵌入的原理將 hctA 基因不活化,由於 hctA 基因位在莢膜合成基因下游位置所以 hctA 基因的不活化不會影響其他莢膜合成基因的表現。本研究構築 Av. paragallinarum 之 hctA 基因缺損突變株及互補株,並比較野外株、突變株與互補株在多種特性之分析工作,包含了莢膜萃取分析、血球凝集試驗分析、生物膜形成分析、細胞吸附分析以及補體毒殺試驗分析。結果發現突變株之莢膜多醣體分子量低於野外株以及互補株,且突變株之細胞黏附能力與形成生物膜之能力皆高於野外株與互補株。另一方面,先前也有研究指出莢膜可以保護菌體免於被吞噬且對於補體毒殺具有抵抗性。上述結果顯示,對 Av. paragallinarum 而言,莢膜之減少,可能有助於細菌初期之吸附與感染,而莢膜之形成,則有助於細菌對補體毒殺之抵抗性與後期之擴散。
文章公開時間: 2017-12-16
Appears in Collections:微生物暨公共衛生學研究所



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