重组α1-giardin减毒沙门菌口服DNA疫苗的构建与鉴定  

The construction of a DNA vaccine with α1-giardin against Giardia delivered by attenuated Salmonella typhimurium elicits

作　　者：李瑶[1] 张宏梅[1] 时文艳[1] 唐文琴[1] 相明[1] 冯宪敏[1] 

机构地区：[1]吉林医药学院病原生物学教研室,吉林吉林市132013

出　　处：《中国兽医学报》2016年第4期600-603,612,共5页Chinese Journal of Veterinary Science

基　　金：国家自然科学基金资助项目(81301450;31572262);吉林省科技厅国际合作资助项目(20160414050GH);吉林市杰出青年基金资助项目(201464037)

摘　　要：以贾第虫的c DNA作为模板,进行PCR扩增得到目的片段,克隆入p VAX1载体,构建重组质粒p VAX1-α1-giardin;以电击转化的方式转入减毒沙门菌株SL7207,构建重组沙门菌SL7207/p VAX1-α1-giardin;通过体外连续培养测定重组菌株的载体携带稳定性;30只清洁级雌性BALB/c小鼠,随机分为3组,每组10只,分别以1×106的SL7207/p VAX1-α-giardin、SL7207/p VAX1和无菌1×PBS(p H 7.0)口服免疫小鼠,8周后处死,检测各组小鼠肠系膜淋巴结中α1-giardin的表达及血清中和小肠灌洗液中特异性Ig G和SIg A的水平。结果表明,重组减毒沙门菌SL7207/p VAX1-α1-giardin连续培养168 h后,质粒携带率为90%;口服免疫小鼠8周后,SL7207/p VAX1-α1-Giardin免疫组,肠系膜淋巴结可见α1-Giardin的高表达,血清中特异性Ig G抗体水平和小肠灌洗液中特异性SIg A水平均明显增高,分别为PBS组的3.6(P<0.01)和5.7倍(P<0.01)。本研究尝试了以减毒沙门菌为载体携带α1-giardin DNA的抗贾第虫口服疫苗的构建及免疫试验,为进一步的疫苗保护性试验和制剂研究奠定了基础。The DNA encoding α1-giardin was cloned into an expression plasmid pVAX1. The recom- binant plasmid pVAX1-α1-giardin and pVAX1 were then transformed into attenuated Salmonella typhimurium strain SL7207 by electroporation. The stability of the plasmid maintained in S. typhimurium was assessed by continuous culture in antibiotic or antibiotic-free medium and enzyme digestion. Ten BALB/c in each group were orally immunized with 1 × 10^6 SL7207 transformed with pVAX1-α1-giardin,the same number of the SL7207 transformed with empty vector pVAX1 only and the PBS control, respectively. Eight weeks after the first immunization, all mice were sacrificed and the sera were collected to detect anti-α1-giardin IgG,intestinal flush was collected to detect al- giardin-specific secretory mucosal IgA,and mesentery was coolected to detect the expression of α1- giardin in lymph node. The results showed 90% of bacteria maintained pVAX1-al-giardin after cultured for 168 hours. The serological total IgG was increased by 3.6 times and mucosal total IgA increased by 5.7 times in mice orally immunized with SL7207/pVAX1-α1-giardin compared with mice given the PBS control. There is no significant difference in both antibody concentration be- tween mice immunized with SL7207/pVAX1 and PBS control group,indicating α1-giardin DNA vaccine delieved by attenuated Salmonella typhimurium could effectively induce serological IgG and mucosal IgA response.

关 键 词：蓝氏贾第鞭毛虫 α1-贾第素 减毒沙门菌 口服疫苗 

分 类 号：S852.72[农业科学—基础兽医学]