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编码草鱼呼肠孤病毒VP5与NS38 B细胞表位的DNA疫苗 壳聚糖纳米口服制剂的研制与评价
唐庆权,张子涵,毛颖睿,鲁子怡,陶敏慧,徐婷婷,曾杭,樊慧敏,许晓牧,彭开松,朱若林,鲍传和
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(安徽农业大学动物科技学院水生健康与公共卫生实验室,合肥 230036;安徽农业大学动物科技学院水生健康与公共卫生实验室,合肥 230036; 安徽农业大学新农村研究院江淮分水岭(定远)综合试验站,定远 233200; 安徽省现代农业产业技术体系(水产)健康养殖研究室,合肥 230036)
摘要:
以编码II型GCRV VP5和NS38的B细胞线性表位的cDNA串联构建pcDNA3.1(+)-Bs5-10(GenBank登录号为MH234474),并包被成纳米级壳聚糖颗粒;无GCRV草鱼(11~12 cm、25~30 g)随机分为空白对照组、空白对照-GCRV攻毒组、裸空载体注射组、壳寡糖包被空载体口服组、裸DNA疫苗注射组(每尾鱼注射10 μg载体,1 d和29 d各免疫1次)、壳寡糖包被DNA疫苗口服组(每尾鱼大约投喂50 μg载体,1~3 d、15~17 d和43~45 d各口服免疫1次),在28℃水温下评价疫苗保护力和特异性抗体水平。结果显示,裸DNA疫苗注射组和壳寡糖包被DNA疫苗口服组的相对保护率分别为66.67%和50%;两组血清抗VP5和抗NS38的IgM水平均随加强免疫而显著升高(P<0.05);且在对应时间点(22 d或50 d),前组特异性IgM水平均显著高于后者(P<0.05)。结果表明,壳聚糖包被的DNA疫苗口服免疫能在一定程度上抵抗GCRV攻击,且体液免疫参与了该保护机制。
关键词:  草鱼呼肠孤病毒  多基因DNA疫苗  口服免疫;注射免疫
DOI:10.13610/j.cnki.1672-352x.20190716.003
基金项目:安徽省省级大学生创新创业训练计划项目(201610364015), 安徽省重点研究与开发计划(1704g07020123), 安徽省现代农业产业技术体系(皖农科[2016]84号)和安徽农业大学大学生创新基金(2017年度)共同资助。
Preparation and evaluation of a DNA vaccine oral nanoparticle oral preparation for encoding grass carp reovirus VP5 and NS38 B cell epitopes
TANG Qingquan,ZHANG Zihan,MAO Yingrui,LU Ziyi,TAO Minhui,XU Tingting,ZENG HANG,FAN Huimin,XU Xiaomu,PENG Kaisong,ZHU Ruolin,BAO Chuanhe
(Aquatic Health and Public Health Laboratory, School of Animal Science And Technology, Anhui Agriculture University, Hefei 230036;Aquatic Health and Public Health Laboratory, School of Animal Science And Technology, Anhui Agriculture University, Hefei 230036; Jianghuai Watershed (Dingyuan) Comprehensive Experimental Station, Anhui Agriculture University New Rural Research Institute, Dingyuan 233200; Anhui Modern Agriculture Industry Technology System (Aquaculture) Healthy Breeding Laboratory, Hefei 230036)
Abstract:
In this study, pcDNA3.1 (+) -Bs5-10 (GenBank login number is MH234474) was constructed in series of B cell linear epitopes of II type GCRV VP5 and NS38, and was packaged into nanoscale chitosan particles. Grass carp (11-12 cm, 25-30 g) without GCRV were randomly divided into blank control group, blank control group-GCRV attack group, bare empty carrier injection group, Chitosan oligosaccharide coated empty carrier oral group, naked DNA vaccine group (at day 1 and 29, each tail fish was injected 10 μg pcDNA3.1 (+) -Bs5-10), and Chitosan oligosaccharide coated DNA vaccine oral group(each tail fish took orally 50 μg pcDNA3.1 (+)- Bs5-10 everyday, which had been carried between 1-3 days, 15-17 days and 43-45 days). Then the protective ability and specific antibody level of vaccine need to be evaluated at the temperature of 28 ℃. The results showed that the relative percent survival of the naked DNA vaccine group and the Chitosan oligosaccharide coated DNA vaccine oral group were 66.67 % and 50 % respectively, and the level of serum anti VP5 and anti NS38 IgM in these two groups increased significantly with the enhanced immunization (P<0.05). Furthermore, the specific IgM level in the former group was significantly higher than the latter (P<0.05) at the corresponding time point (22 days
Key words:  grass carp reovirus  multi-gene vaccine  oral immunity  injection immunity

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