表达猪源GM-CSF重组PRRSV疫苗对同源高致病性毒株的免疫保护效力评价

Evaluation of Immune Protective Effect of Recombinant PRRS Attenuated Vaccine Strain Expressing Porcine GM-CSF Against Homologous HP-PRRSV

为研究表达猪源粒细胞-巨噬细胞集落刺激因子(GM-CSF)的重组猪繁殖与呼吸综合征病毒(PRRSV)在动物体内的免疫调节特性及对其的保护效力评价,本研究将15头30日龄仔猪随机分成4组,空白对照组(DMEM)4头、疫苗对照组(HuN4-F112株)4头、疫苗组(rPRRSV-GM-CSF株)3头和攻毒对照组(DMEM^(+)HuN4株)4头。疫苗对照组肌注免疫HuN4-F112株10^(5) TCID_(50)/头、疫苗组肌注免疫rPRRSV-GM-CSF株10^(5) TCID_(50)/头,实验空白组和攻毒对照组肌注DMEM 2 mL/头,免疫后28 d,疫苗组、疫苗对照组和攻毒对照组肌注HuN4株(10_(5) TCID_(50)/头)。攻毒后的试验结果表明,免疫rPRRSV-GM-CSF重组病毒组和疫苗株HuN4-F112组获得完全保护,阴性对照组全部死亡;通过IDEXX试剂盒检测仔猪血清中PRRSV抗体水平可知,在免疫14 d后,疫苗组抗体水平显著高于疫苗对照组(P<0.05);由流式细胞术分析体内免疫细胞亚群比例可知,疫苗组同疫苗对照组相比,疫苗组的重组疫苗株能够引起免疫记忆细胞亚群CD4^(+)CD8^(+)T在免疫后28 d显著升高,以及引发攻毒后其抗原递呈细胞显著增多,进而促进CD4-CD8^(+)T的增殖以发挥抗病毒免疫应答。本研究筛选出了一株具有改善HuN4-F112弱毒疫苗株免疫效果的重组病毒rPRRSV-GM-CSF,为进一步研发广谱通用的新型疫苗奠定基础。

To evaluate the immunoregulatory characteristics and protection of the recombinant PRRS virus expressing porcine GM-CSF(rPRRSV-GM-CSF)in animals,fifteen 30-day-old piglets were randomly divided into 4 groups.Four piglets in vaccine control group were injected with 10^(5) TCID_(50) HuN4-F112 strain,3 piglets in experimental group were injected with 10^(5) TCID_(50) rPRRSV-GM-CSF strain,and 4 piglets in each of blank group and challenge control group were injected with 2 mL DMEM.At 28 days after immunization,HuN4 strains(10_(5) TCID_(50))was intramuscularly injected into each piglet in experimental group,vaccine control group and challenge control group.Piglets were observed for 28 days post challenge.The results showed that the rPRRSV-GM-CSF group and HuN4-F112 group were completely protected and all piglets in control group died after challenge.Serum samples were collected from piglets at day 14 post immunization and tested for antibody levels with IDEXX ELISA kit.The antibody level of the rPRRSV-GM-CSF group was significantly higher than that of the HuN4-F112 group(P<0.05)and remained high until the later stage post challenge.Blood samples were also tested in flow cytometry for analysis of the proportion of immune cell subsets.As compared with the HuN4-F112 group,the immune memory cell subsets CD4^(+)CD8^(+)T in the rPRRSV-GM-CSF group significantly increased at day 28 post immunization and the antigenpresenting cells also significantly increased post challenge.In this study,the recombinant virus rPRRSV-GM-CSF was proofed to deliver better immune protection than the currently used HuN4-F112 attenuated vaccine strain,laying a foundation for the further research and development of a new vaccine with broad spectrum cross-protection.