鲤疱疹病毒2型(CyHV-2)灭活疫苗制备及其免疫效力测定

鲫造血器官坏死病是由鲤疱疹病毒2型(CyHV-2)引起的危害我国养殖鲫鱼(Carassius auratus)最严重的病毒性疾病。为制备有效预防鲫造血器官坏死病的灭活疫苗,以CyHV-2代表流行株CNDF-TB2015为疫苗株,以CSC细胞系进行病毒增殖,并采用不同浓度的病毒对不同规格鲫鱼进行攻毒,建立攻毒模型。结果表明,培养的病毒滴度达10¹⁰ TCID₅₀/mL,以4×10⁷TCID₅₀/尾的剂量腹腔注射感染体重300 g的鲫鱼可造成100%的死亡率。对病毒灭活剂甲醛、β-丙内酯和BEI灭活条件进行筛选优化,对疫苗免疫佐剂763A、IMS1312、铝胶和M402进行筛选,结果表明,浓度为10^8.3 TCID₅₀/mL的病毒液最佳灭活条件为终浓度10 mmol/L的BEI 37 ℃灭活72 h,疫苗制备最佳佐剂为水佐剂IMS1312。对灭活疫苗的最小免疫剂量、最佳免疫剂量和有效免疫产生期进行测定,结果表明,要获得70%以上保护率的最小免疫剂量为8×10⁶ TCID₅₀/尾,最佳免疫剂量为4×10⁷ TCID₅₀/尾,在免疫4周后保护率可达80%, 5周后保护率达到100%。制备的灭活疫苗为鲫造血器官坏死病的免疫预防提供了有力手段,也为其他水产动物病毒灭活疫苗的制备提供了借鉴。

PREPARATION AND EFFICACY OF INACTIVATED VACCINE AGAINST CYPRINID HERPESVIRUS-2

Herpesviral haematopoietic necrosis disease (HVHN) is a viral disease of crucian carp, caused by Cyprinid herpesvirus-2 (CyHV-2), which causes huge losses on aquaculture in China. In order to control the occurrence of HVHN, the inactivated vaccine was developed based on the vaccine strain CNDF-TB2015 and the immune efficacy was assessed. The results of CyHV-2 culture showed that the titer of virus cultured with CSC cells reached 10¹⁰ TCID50/mL, and the challenge model test showed the dose of 4×10⁷ TCID50/ind could cause 100% mortality in crucian carp within 14 d post injection with the CyHV-2 CNDF-TB2015. The results of inactivated vaccine preparation test showed that the optimal condition for CyHV-2 inactivation with 10^8.3 TCID50/mL was 10 mM BEI at 37 °C for 72 h and the best adjuvant was IMS1312. The protection rate with the minimum immune dose of 8×10⁶ TCID50 /ind could reach more than 70%, with the optimal immune dose of 4×10⁷ TCID50/ind could reach 100%. The protection rate reached 80% at week 4 postimmune and 100% at week 5 postimmune. This study indicated that the inactivated CNDF-TB2015 strain of CyHV-2 could serve as a potential vaccine candidate to prevent the prevalence of HVHN, also provides reference for the preparation of other inactivated viral vaccines for aquaculture.