鴿環狀病毒類病毒顆粒疫苗免疫效力評估

鴿環狀病毒(Pigeon Circovirus, PiCV)在1980年代末被發現，至今遍布全球。鴿環狀病毒會攻擊鴿隻免疫系統，特別是華氏囊，並造成免疫抑制，進而發生細菌、病毒、原蟲等二次感染。幼鴿與成鴿皆會受到感染，但好發於四到六個月大之幼鴿，因此被又稱為幼鴿疾病綜合症(young pigeon disease syndrome, YPDS)。類病毒顆粒(virus-like particles, VLPs)是由病毒的結構蛋白的自組裝產生的，相似於天然病毒，卻沒有病毒的基因體。比起傳統次單位疫苗能夠更有效地被抗原呈遞細胞識別、誘發更強的免疫反應。本實驗將鴿環狀病毒外殼蛋白基因轉殖到pET32a及pGS21a載體上，利用大腸桿菌系統大量誘導生產重組鴿環狀病毒外殼蛋白，外殼蛋白會自組成類病毒顆粒。純化經西方點墨法確認抗原性後，皮下注射進行免疫評估。西方點墨法結果顯示，在大於180kDa處有明顯的條帶，推測為類病毒顆粒。可溶性蛋白生產量pET32a-picv優於pGS21-picv。在30℃下誘導所形成的類病毒顆粒會多於37℃下誘導形成的。動物實驗結果顯示，施打類病毒顆粒的組別血清中的抗體相較於控制組具有顯著性的差異，在補強後能誘發更快更高的抗體免疫反應。

關鍵字

鴿環狀病毒；外殼蛋白；類病毒顆粒；原核表現系統

並列摘要

Pigeon Circovirus (PiCV) has been reported worldwide ever since it was reported in the 1980s. The virus attacks the pigeons' immune system specifically the bursa, which can cause immunosuppression and subsequently lead to secondary bacterial, viral or protozoan infections. PiCV infects young and old pigeons, but easily found in four to six months old pigeons therefore it is also known as young pigeon disease syndrome (YPDS). Virus-like particles (VLPs) are produced by the self-assembly of structural proteins of viruses. It is similar to natural viruses but without the viral genome. Compared with the traditional subunit vaccine, the VLP vaccine can be more effectively recognized by the antigen-presenting cells and induce a stronger immune response. In this study, the PiCV capsid protein gene was cloned into the pET32a and pGS21a vectors. Escherichia coli expression system was used to produce the recombinant PiCV capsid protein. The capsid protein will assemble into VLPs. Following purification, western blot analysis was used to confirm the antigenicity. Then, vaccine was injected via subcutaneous injection. Western blot assay showed significant band at greater than 180 k Da thus, it was hypothesized that it is a virus-like particle. Soluble protein production in pET32a-picv is better than in pGS21-picv. Induced PiCV capsid protein, more VLPs assembled at 30°C than in 37°C. The animal test showed the antibody titer of immunized with VLPs is higher than the control group. After the animal boost, they would have faster and stronger immune response.