日本血吸虫中国株琥珀酸脱氢酶铁硫蛋白真核重组体的构建及其免疫小鼠的保护性研究

目的研究日本血吸虫琥珀酸脱氢酶铁硫蛋白(SjSDISP)全长编码基因DNA疫苗诱导小鼠的免疫保护性效果。方法将全长SjSDISP基因克隆到真核表达载体pcDNA3上,构建真核表达质粒pcDNA3-SjSDISP,用重组裸DNA免疫昆明小鼠3次,间隙2W,末次免疫后第二周进行日本血吸虫尾蚴攻击感染,感染后42 d处死小鼠,剖杀冲虫,计算减虫率和减卵率。结果经双酶切、PCR和测序验证,表明真核表达质粒pcDNA3-SjSDISP构建成功。动物免疫保护性实验结果显示:pcDNA3-SjSDISP疫苗组减虫率较低,与对照组相比减虫效果不明显(P>0.05);但在每克肝卵、每克粪卵和每雌子宫内卵方面,与对照组相比差异均具有显著性(P<0.01)。结论日本血吸虫琥珀酸脱氢酶铁硫蛋白DNA疫苗可能主要在抗血吸虫卵胚发育或抗生殖起作用,具有潜在的疫苗研究与开发价值。

Construction of the Eukaryotic Recombinant vector for succinate dehydrogenase iron-sulfur protein of Schistosoma japonicum Chinese strain and its protective immunity in mice

To develop succinate dehydrogenase iron-sulfur protein DNA vaccine of Schistosoma japonicum(SjSDISP)Chinese strain and to determine its protective efficacy in infected Kunming mice,the full-length cDNA of SjSDISP amplified from adult S.japonicum cDNA library was cloned into a eukaryotic expression vector pcDNA3 to construct the recombinant pcDNA3-SjSDISP.The naked recombinant pcDNA3-SjSDISP was used to immunize the mice three times with an interval of two weesks.and the pcDNA3 group was set up as control.All mice were challenged with 40 cercariae percutaneously on abdominal skin at week 8,killed and calculated the worms and eggs 42 days after infection.The recombinant plasmid pcDNA3-SjSDISP was confirmed by double endonucleases digestion,PCR and sequencing.Mice vaccinated with recombinant plasmid pcDNA3-SjSDISP revealed significant differences on liver eggs per gram(LEPG),fecal eggs per gram(FEPG) and intrauterine eggs reduction rate as compared with the control groups.It is evident the the full-length cDNA of SjSDISP was successfully cloned into eukaryotic expression plasmid pcDNA3 that could elicit moderate significant levels of anti-fecundity or anti-egg immunity against S.japonicum,indicating its potential as a promising vaccine candidate.