Zhang Huawei, Qian Ping, Peng Bo, Shi Lin, Chen Huanchun, Li Xiangmin
State Key Laboratory of Agricultural Microbiology, Huazhong Agricultural University, Wuhan 430070, Hubei, China; Laboratory of Animal Virology, College of Veterinary Medicine, Huazhong Agricultural University, Wuhan 430070, Hubei, China.
State Key Laboratory of Agricultural Microbiology, Huazhong Agricultural University, Wuhan 430070, Hubei, China; Laboratory of Animal Virology, College of Veterinary Medicine, Huazhong Agricultural University, Wuhan 430070, Hubei, China.
Vaccine. 2015 May 15;33(21):2449-56. doi: 10.1016/j.vaccine.2015.03.090. Epub 2015 Apr 8.
Porcine circovirus type 2 (PCV2) causes porcine circovirus-associated disease. Capsid (Cap) protein of PCV2 is the principal immunogenic protein that induces neutralizing antibodies and protective immunity. GM-CSF is an immune adjuvant that enhances responses to vaccines. In this study, recombinant baculoviruses Ac-Cap and Ac-Cap-GM-CSF expressing the Cap protein alone and co-expressing the Cap protein and porcine GM-CSF, respectively, were constructed successfully. The target proteins were analyzed by western blotting and IFA. Further, these proteins were confirmed by electron microscopy, which showed that Cap proteins could self-assemble into virus-like particles having diameters of 17-25nm. Animal experiments showed that pigs immunized with Cap-GM-CSF subunit vaccine showed significantly higher levels of PCV2-specific antibodies and neutralizing antibodies than pigs immunized with the Cap subunit vaccine and a commercial vaccine (Ingelvac CircoFLEX; P<0.05). After PCV2 wild strain challenged, Pigs receiving the Cap-GM-CSF subunit vaccine showed significantly higher average daily weight gain after wild-type PCV2 challenge than pigs receiving the other three vaccines (P<0.05). None of PCV2 DNA was detected in all immunized animals, except control animals immunized with phosphate-buffered saline. These results indicated that GM-CSF was a powerful immunoadjuvant for PCV2 subunit vaccines because it enhanced humoral immune response and improved immune protection against PCV2 infection in pigs. Thus, the novel Cap-GM-CSF subunit vaccine has the potential to be used as an effective and safe vaccine candidate against PCV2 infection.
猪圆环病毒2型（PCV2）可引发猪圆环病毒相关疾病。PCV2的衣壳（Cap）蛋白是诱导中和抗体和保护性免疫的主要免疫原性蛋白。GM-CSF是一种可增强疫苗反应的免疫佐剂。在本研究中，成功构建了分别单独表达Cap蛋白以及共表达Cap蛋白和猪GM-CSF的重组杆状病毒Ac-Cap和Ac-Cap-GM-CSF。通过蛋白质免疫印迹法和间接免疫荧光法对目标蛋白进行了分析。此外，通过电子显微镜对这些蛋白进行了确认，结果表明Cap蛋白可自组装成直径为17-25nm的病毒样颗粒。动物实验表明，与接种Cap亚单位疫苗和商业疫苗（英特威猪圆环病毒疫苗；Ingelvac CircoFLEX）的猪相比，接种Cap-GM-CSF亚单位疫苗的猪体内PCV2特异性抗体和中和抗体水平显著更高（P<0.05）。在受到PCV2野生毒株攻击后，接种Cap-GM-CSF亚单位疫苗的猪在野生型PCV2攻击后的平均日增重显著高于接种其他三种疫苗的猪（P<0.05）。除用磷酸盐缓冲盐水免疫的对照动物外，在所有免疫动物中均未检测到PCV2 DNA。这些结果表明，GM-CSF是PCV2亚单位疫苗的一种强大免疫佐剂，因为它增强了体液免疫反应，并改善了猪对PCV2感染的免疫保护。因此，新型Cap-GM-CSF亚单位疫苗有潜力作为一种有效且安全的候选疫苗用于预防PCV2感染。
