豬霍亂沙門氏桿菌(Salmonella enterica serovar Choleraesuis)長期潛伏於豬場，感染豬隻，造成豬隻下痢、失重、生長率下降及全身性敗血症，嚴重者甚至會死亡，好發於保育至肥育前期(8週至14週)的豬群。近年來，由於抗生素之濫用及抗藥性菌株產生，且豬霍亂沙門氏桿菌會寄生於細胞內，不易被抗生素殺滅，造成防治上的困難。因此發展安全有效之疫苗，為防治豬霍亂沙門氏桿菌之重要課題。鞭毛蛋白是構成鞭毛的重要元素，能誘導宿主產生體液性及細胞性免疫反應。本實驗以引子對FliC-F和FliC-R增幅fliC基因，並比較標準菌株和近4年16株野外分離株的鞭毛蛋白序列，顯示菌株間之鞭毛蛋白序列無明顯改變。將標準菌株之fliC基因構築於pGEX4T-1載體，轉型至E. coli BL21大量表現鞭毛蛋白，將純化的鞭毛蛋白以不同濃度進行腹部注射免疫接種BALB/c小鼠。小鼠每週固定抽取眼窩血，以血清進行ELISA測試，二次免疫50 μg組的小鼠血清抗體力價為1:160，與陽性對照組(減毒疫苗, Suisaloral®)的抗體力價相同。另外，培養免疫接種小鼠的脾臟細胞，利用ELISA測試培養液內IL-2、IFN-γ和TNF-α三種細胞激素含量，二次免疫50 μg組的小鼠脾臟細胞分泌量分別為529 pg/ml、849 pg/ml、392 pg/ml，最接近陽性對照組。在攻毒試驗中，二次免疫50 μg組的小鼠存活率為100%且沒有臨床症狀產生，與陽性對照組的結果相同。綜合上述實驗結果，豬霍亂沙門氏桿菌的鞭毛蛋白具有引發小鼠體液性和細胞性免疫反應的能力，並能誘使小鼠體內產生保護力，顯示鞭毛蛋白可當作次單位疫苗或免疫佐劑的選擇，進而應用於防治豬霍亂沙門氏桿菌。

Salmonella enterica serovar Choleraesuis is one of the predominant pathogens in Taiwanese piggeries. Typical clinical manifestations are septicemia, diarrhea, weight loss, growth rate decreasing, and even death. It causes disease in weaning pigs and in the early stage of finishing pigs (during 8 weeks to 14 weeks). Salmonella enterica serovar Choleraesuis belongs to Enterobacteriaceae. In recent years, due to abusing antibiotics in the field, antibiotic resistence has been a serious problem. Moreover, Salmonella enterica serovar Choleraesuis, which is an intracellular pathogen, is resistant to antibiotic treatment. It is important to develop a safe and more effective vaccine. Flagellum is composed with flagellin, which can induce humoral and cellular immune resposes. In this study, FliC-F and FliC-R were used as primers to amplify fliC gene. We align the flagellin sequences of the standard strain and 16 recently isolated strains. It is shown that the fligellin sequences were similar in these strains. The gene was constructed into pGEX4T-1 and transformed to E. coli BL21 to express the flagellin. The recombinant flagellin was immunized to the BALB/c with three different doses of flagellin via intraperitoneal route. Blood samples were weekly collected by orbital bleeding, and antibody titers were determined by ELISA. The group of 50 μg immunized twice has the same results of antibody titer (1:160) as the group of attenuated vaccine. Spleen cells of immunized mice were cultured and amount of cytokines (IL-2、IFN-γ and TNF-α) in medium were determined by ELISA. The cytokine secretion were 529 pg/ml、849 pg/ml、392 pg/ml in group of 50 μg immunized twice. It is similar with the result of the group of attenuated vaccine. Immunized mice were challenged by Salmonella enterica serovar Choleraesuis to observe the efficacy of flagellin. The group of 50 μg immunized twice has the same survival rate (100%) as the group of attenuated vaccine. The results suggested that the flagellin could provide the protection against Salmonella enterica serovar Choleraesuis and had implications for development of a subunit vaccine for Salmonella enterica serovar Choleraesuis.

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