【作者】 吴绍强； 蒋金书； 朱引洁； 刘群；

【Author】 WU Shao-qiang, JIANG Jin-shu, ZHU Yin-jie, LIU Qun(1. College of Veterinary Medicine, South China Agricultural University, Guangzhou, Guangdong 510642,China; 2.College of Veterinary Medicine, China Agricultural University, Beijing 100094, China)

【机构】 华南农业大学兽医学院； 中国农业大学动物医学院；

【摘要】 将E.tenella BJ株的保护性抗原基因TA4插入真核表达载体中,然后在其上游融合方式插入EtlA基因,构建成核酸疫苗并进行体外表达。IFAT检测表明,pcDT转染细胞的荧光亮度高,pcDET载体大,转染效率低,荧光较弱。动物保护性试验中,间隔一周两次免疫鸡后攻虫发现,核酸疫苗对球虫感染具有明显的保护作用,可显著减轻球虫感染引起的机体增重下降,其中,重组干扰素联合pcDT中剂量(50μg)免疫时,鸡的增重效果最为明显,盲肠病变值也最轻微,ACI达167.2。试验还发现,50μg pcDET可达到100μg pcDT的抗球虫效果,ACI在160以上,说明融合插入两种基因构建的核酸疫苗对球虫感染的保护效果更佳。更多还原

【Abstract】 In order to construct DNA vaccines against Eimeria infection, the gene TA4 of Eimeria tenella BJ strain was ligated to the mammalian expression vector pcDNA3.1/Zeo (+), and then, EtlA, another gene of Eimeria tenetta. BJ strain, was ligated to its upstream, aiming at their expression in fusion form. The IFAT method was established to examine the in vitro expressions of the constructed DNA vaccines in COS cells, all gave positive results. In the clinical experiment, the DNA vaccines were given to the broilers i.m. 10-15 min after the breasts had been pre-treated with 25% sucrose solution. Vaccinated twice at 1st and 2nd week of age, chickens were challenged with 3×l04 sporulated oocysts of E. tenetta BJ strain at the 3rd week of age. Chickens were slaughtered and the lesion scores of ceca, the relative body weight gains and the numbers of oocysts in the ceca of each group of chickens were assessed and compared at the 8th day post inoculation. The anti-coccidial index (ACI) was used as a criterion to compare the effects of each treatment.Results indicated that both pcDNA3.1-TA4 (pcDT) and pcDNA3.1-EtlA-TA4 (pcDET) could induce protective immunity against coccidial challenge. Their use could obviously alleviate the body weight decrease of chickens following coccidial infection. When 50 μg pcDT was given to chickens together with the recombinant chicken IFN-γ, chickens had the maximum body weight gain (108.8% of that of the unchallenged control group) and the minimum lesion score in all the inoculated groups. The corresponding ACI of this group was 167.2, while the ACI of 160 was achieved with 50μg pcDET or 100μg pcDT treatment. This indicates that the fusion expression of two protective antigen genes in the same plasmid can spare the use of the constructed piasmid, and then decrease the manufacturing expenses of DNA vaccine.更多还原