Please use this identifier to cite or link to this item: http://hdl.handle.net/11455/66313
標題: 家禽霍亂巴斯德桿菌絲狀血球凝集素基因之分析與應用
Characterization of the Filamentous Haemagglutinin Genes of Pasteurella multocida
作者: 李宜珊
Li, Yi-Shan
關鍵字: fowl cholera
家禽霍亂
P. multocida
filamentous haemagglutinin
pfhB1
pfhB2
recombinant protein
巴斯德桿菌
絲狀血球凝集素
重組蛋白
出版社: 微生物暨公共衛生學研究所
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Sthitmatee, N., Numee, S., Kawamoto, E., Sasaki, H., Yamashita, K., Takahashi, N., Kataoka, Y., Sawada, T., 2008. Protection of chickens from fowl cholera by vaccination with recombinant adhesive protein of Pasteurella multocida. Vaccine 26, 2398-2407. Tan, H.Y., Nagoor, N.H., Sekaran, S.D., 2010. Cloning, expression and protective capacity of 37 kDa outer membrane protein gene (ompH) of Pasteurella multocida serotype B:2. Tropical Biomedicine 27, 430-441. Tatum, F. M., Yersin, A.G., Briggs, R.E., 2005. Construction and virulence of a Pasteurella multocida fhaB2 mutant in turkeys. Microb. Pathog. 39, 9-17. Tatum, F.M., Tabatabai, L.B., Briggs, R.E., 2009. Protection against fowl cholera conferred by vaccination with recombinant Pasteurella multocida filamentous hemagglutinin peptides. Avian Dis. 53, 169-174. Townsend, K.M., Boyce, J.D., Chung, J.Y., Frost, A.J., Adler, B., 2001. 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摘要: 家禽霍亂 (Fowl cholera,FC)為家禽養殖業之重要疾病,此疾病之病原為家禽霍亂巴斯德桿菌 (Pasteurella multocida)。P. multocida藉由宿主上呼吸道的黏膜入侵宿主組織並繁殖,進而產生嚴重的系統性疾病。百日咳博徳氏菌的主要黏附素 (adhesin)為絲狀血球凝集蛋白 (Filamentous haemagglutinin,Fha),此蛋白是革蘭氏陰性菌的主要分泌蛋白之一。Fha可被運送至細胞表面或分泌到細胞外,並可促進細菌的散佈與聚生 (colonization)。先前研究發現,P. multocida菌株Pm70之基因存在兩個絲狀血球凝集素基因,pfhB1 (pm0057)、pfhB2 (pm0059),本研究針對4株禽源P. multocida的pfhB1及pfhB2基因作進一步探討。首先以PCR增幅出P. multocida菌株的pfhB1和pfhB2基因,並進行定序後比較其序列,比較結果發現不同菌株的PfhB1胺基酸序列具有98.5%~99.3%的相似性,而PfhB2胺基酸序列則有64.5~99.4%相似性,接著利用Pfam程式分析不同菌株之PfhB1、PfhB2,結果發現所有菌株的PfhB1與PfhB2蛋白皆含有一個hemagglutinin activity domain、一個peptidase C58 (Yersinia cystein protease_YopT) domain,以及多個haemmagglutinin repeats domain。本研究利用重組蛋白技術表現包含上述domain之PfhB與PfhB2蛋白片段,並探討其免疫原性與保護效力。本研究設計實驗之結果顯示包含PfhB1之peptidase_C58 domain的重組蛋白X73_pfhB1-2286-2613具有切割RhoA蛋白之protease活性。本研究之結果可協助了解P. multocida之致病機制並作為研發家禽霍亂疫苗之參考。
Pasteurella multocida is the causative agent of fowl cholera (FC), which is an economically important disease of the poultry. The filamentous hemagglutinin (Fha) is the major adhesin of the Bordetella pertussis and proteins showing sequences homology to Fha are found in a number of Gram-negative bacteria including P. multocida. The Fha is transported to the bacterial surface or secreted to the extracellular environment and could enhance bacterial dispersion and colonization. Analysis of the P. multocida Pm70 genome sequence identified two predicted filamentous haemagglutinin genes, pfhB1 (pm0057) and pfhB2 (pm0059). In this study, pfhB1, pfhB2 genes from four avian strains of P. multocida were characterized. PCR was used to amplify pfhB1 and pfhB2 genes from P. multocida. Sequence analyses showed that the amino acid sequences of PfhB1 from the four strains have 98.5-99.3% sequence identity, whereas the amino acid sequences of PfhB2 have only 64.5~99.4% sequence identity. Pfam search showed that both PfhB1 and PfhB2 contain a hemagglutinin activity domain, a peptidase C58 (Yersinia cystein protease_YopT) domain, and a numbers of haemmagglutinin repeats domains. Recombinant proteins containing these domains of PfhB1 and PfhB2 were produced and antigenicity and protection efficacy coferred by these recombinant proteins were investigated. In this study, we designed an experiment to determine whether the recombinant protein containing the peptidase_C58 domain of PfhB1 have the protease activity to cleave RhoA protein. The results show that the recombinant protein X73_pfhB1-2286-2613 have the protease activity to cleave RhoA protein. Results from this study are useful for understanding the biological functions of the Fha proteins and development of a vaccine against P. multocida.
URI: http://hdl.handle.net/11455/66313
其他識別: U0005-0407201116162000
文章連結: http://www.airitilibrary.com/Publication/alDetailedMesh1?DocID=U0005-0407201116162000
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