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目的:从临床样本中分离发热伴血小板减少综合征布尼亚病毒(severe fever with thrombocytopenia syndrome bunyavirus,SFTSV)毒株,经过充分比较研究后,确定1株适合疫苗生产的毒株。方法:使用Vero细胞从发热伴血小板减少综合征患者临床样本中分离病毒株,经噬斑纯化鉴定后进一步适应传代,比较研究各毒株在细胞中的传代适应性、免疫原性、交叉保护性以及遗传稳定性等,筛选并确定1株疫苗生产用毒株,按照要求建立三级种子库。结果:共获得7个SFTSV分离株,各分离株病毒均可以在Vero细胞中良好复制,病毒滴度均可达到7.0 lg CCID50·m L~(-1)以上,其中AH12株滴度可达9.0 lg CCID50·m L~(-1)。毒株适应传代后滴度稳定,灭活病毒免疫动物均可产生针对本株及其他毒株的中和抗体,其中HB29株和AH12株免疫后具有较高的中和抗体水平以及良好的交叉保护性。基于AH12株建立的三级种子库经全面检定,结果符合《中国人民共和国药典》三部(2010版)要求。结论:经过对不同SFTSV毒株的比较研究,确定AH12株为疫苗生产用毒株并成功建立三级种子库。
OBJECTIVE: To isolate a strain of severe fever with thrombocytopenia syndrome bunyavirus (SFTSV) isolated from clinical samples. After a thorough comparative study, a strain suitable for vaccine production was identified. Methods: The virus strains were isolated from clinical samples of patients with fever and thrombocytopenia syndrome by Vero cells. After purified by plaque purification, the strains were further adapted to passage. The passage adaptability, immunogenicity and cross-protection of each strain in the cells were comparatively studied And genetic stability, screening and identification of a strain of vaccine production, in accordance with the requirements of the establishment of three seed banks. Results: A total of 7 SFTSV isolates were obtained. The viruses of all isolates could be replicated well in Vero cells. The titer of virus could reach above 7.0 lg CCID50 · m L -1, and the titer of AH12 strains reached 9.0 lg CCID50 · m L ~ (-1). The strain was stable after passage, and the neutralizing antibody against this strain and other strains could be produced by inactivated virus. Among them, HB29 strain and AH12 strain had higher neutralizing antibody level and good cross protection Sex. The third-grade seed bank established based on AH12 strain was fully tested and the results were in conformity with the requirements of the third part (2010 edition) of the Pharmacopoeia of the People’s Republic of China. Conclusion: After comparing the different SFTSV strains, AH12 strains were confirmed as vaccine production strains and the third seed bank was established successfully.