細菌高變異性及多血清型的特性是交叉保護疫苗研發的主要瓶頸。先
前研究中我們實驗室藉由免疫蛋白質體學技術發現發光桿菌(舊稱巴斯德
桿菌) (Photobacterium damselae subsp. Piscicida)之HSP60、enolase、GAPDH，屬於具有保護力之重要抗原。自台南地區養殖魚塭篩得之弧菌病原坎氏弧菌 (Vibrio campbellii)採用同樣的技術也篩得此三免疫原，兩病原菌的三個免疫原具84 %以上核酸相同度及94 %以上胺基酸相似
度。在這兩個菌種中，認為此三個抗原的胺基酸具高度保留性。
本篇探討此三個蛋白發展成為交叉保護性疫苗的可行性。使用大腸桿
菌表現之巴斯德桿菌及坎氏弧菌rHSP60、rEnolase 及rGAPDH 來免疫點
帶石斑魚，再以坎氏弧菌進行攻毒。結果顯示，巴斯德桿菌之rHSP60、
rEnolase 及rGAPDH 雖與坎氏弧菌之rHSP60、rEnolase 及rGAPDH 具94
％以上胺基酸相似度，但卻不具保護力。
由同源塑型結構預測結果顯示這三對不同來源之免疫原，其胺基酸變
異點皆位於結構表面。我們認為高變異的表面結構是決定免疫反應的關
鍵，影響保護力的差異。

High diversity and multiple serotypes in bacteria are the main bottleneck of cross-protective vaccine development. In previous studies, our lab isolated three protective immunogens, HSP60, enolase and GAPDH in Photobacterium damselae subsp. Piscicida (Ph. d. p.) by immunoproteomics
approach. In other studies, Vibrio campbellii was identified as a pathogen in fish farm in Tainan. These three immunogens were also isolated from this V. campbellii by the same approach. Preliminary data showed that the same immunogens derived from Ph. d. p. and V. campbellii shares at least 84 % nucleotide identity and 94 % amino acid homology. HSP60, enolase and GAPDH were considered as high conserved immunogens.
We evaluated the feasibility of using these three immunogens as universal vaccine candidates in this research. Orange-spotted grouper were immunized with E.coli expressing rHSP60, rEnolase and rGAPDH derived from Ph. d. p. and V. campbellii, and challenged with V. campbellii. Results indicated that rHSP60, rEnolase and rGAPDH of Ph. d. p. did not protect V. campbellii challenge, indicating antigens even contain 94 % amino acid similarity, still could not provide cross-protectivity.
Homology modeling prediction results indicated that the difference of amino acids sequences of these three pairs of proteins located on their surface. We suggested the variable surface structure may be the key in determining the response of immunity.

林煜晟 (2009)。 石斑魚養殖上強毒株坎氏弧菌免疫抗原分析與疫苗開發，國立成功大學生物科技研究所碩士論文

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