RESULTS: The 2BS cell-adapted rabies virus strain 2aG4-B40 was established by passage for 40 generations and selection of plaques in 2BS cells. RNA sequence analysis revealed that mutations in 2BS cell-adapted strains were not located at key sites that regulate the production of neutralizing antibodies or virulence in the aG strain (GQ412744.1). The gradual increase in virulence (remaining above 7.0 logLD50/ml from the 40th to 55th generation) and antigen further indicated that these mutations may increase the affinity of the adapted strains for human diploid cells. Identification tests revealed that the 2BS cell-adapted virus strain was neutralized by anti-rabies serum, with a neutralization index of 19,952. PrEP and PEP vaccination and the NIH test further indicated that the vaccine prepared with the 2aG4-B40 strain had high neutralizing antibody levels (2.24 to 46.67 IU/ml), immunogenicity (protection index 270) and potency (average 11.6 IU/ml).
CONCLUSIONS: In this study, a 2BS cell-adapted strain of the 2aG4 rabies virus was obtained by passage for 40 generations. The results of sequencing analysis and titre determination of the adapted strain showed that the mutations in the adaptive process are not located at key sequence regions of the virus, and these mutations may enhance the affinity of the adapted strain for human diploid cells. Moreover, vaccines made from the adapted strain 2aG4-B40 had high potency and immunogenicity and could be an ideal candidate rabies virus strain for inactivated vaccine preparation.
结果:通过传代40代并在2BS细胞中选择噬斑,建立了适应2BS细胞的狂犬病病毒株2aG4-B40。RNA序列分析揭示2BS细胞适应菌株中的突变不位于调节aG菌株中的中和抗体产生或毒力的关键位点(GQ412744.1)。毒力(从第40代到第55代保持在7.0logLD50/ml以上)和抗原的逐渐增加进一步表明,这些突变可以增加适应菌株对人二倍体细胞的亲和力。鉴定试验显示,抗狂犬病血清中和了适应2BS细胞的病毒株,中和指数为19,952。PrEP和PEP疫苗接种和NIH试验进一步表明,用2aG4-B40菌株制备的疫苗具有较高的中和抗体水平(2.24至46.67IU/ml),免疫原性(保护指数270)和效力(平均11.6IU/ml)。
结论:在这项研究中,通过传代40代获得2aG4狂犬病病毒的2BS细胞适应株。适应株的测序分析和滴度测定结果表明,适应过程中的突变不位于病毒的关键序列区域,这些突变可以增强适应菌株对人二倍体细胞的亲和力。此外,由适应株2aG4-B40制成的疫苗具有较高的效力和免疫原性,可以作为灭活疫苗制备的理想候选狂犬病病毒株。