A 2BS cell adapted China fixed rabies virus vaccine strain 2aG4-B40 and its application prospect in human rabies vaccine

Abstract

Abstract BACKGROUND Rabies virus is a fatal zoonotic disease which the pathogenesis has not been fully elucidated, vaccination is the only effective method to protect against rabies virus infection. Most inactivated vaccines are produced using Vero cells (African green monkey kidney cells) for its large-scale productivity. However, there is a potential carcinogenic risk due to the non-human DNA contamination. Thus, replacing Vero cells with human diploid cells is a better solution. In this study, we invented a 2BS adapted rabies virus strain and analyzed its sequence, virulence and immunogenicity to prepare a new strain that may be used in the production of human diploid cell inactivated vaccine. METHODS AND RESULTS The human diploid adapted rabies virus strain 2aG4-B40 was established at the 40th generation by the combination of passage and pick spot in 2BS cells. The RNA sequence analysis showed that the mutations in 2BS adapted strains were not located on the key sites that affect the production of neutralizing antibodies and virulence compared to aG strain (GQ412744.1). The gradually increased virulence (remained above 7.0LogLD50/ml from the 40th to 55th generation) and antigen further indicated that these mutations may enhance the affinity of adapted strains to human diploid cells. Identification tests showed that the 2BS adaption virus strain was neutralized by anti-rabies serum with a neutralization index of 19952. PrEP and PEP vaccination and the NIH test further indicated that the vaccine prepared with the 2aG4-B40 strain had high levels of neutralizing antibody (2.24 to 46.67 IU/ml), immunogenicity (protection index 270) and potency (average 11.6 IU/ml). CONCLUSIONS In this study, 2BS adapted strain of 2aG4 rabies virus was obtained at the 40th generation. The results of sequencing analysis and titer determination of the adapted strains 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 adapted strains to human diploid cells. Meanwhile, vaccines made from the adapted strain 2aG4-B40 had high potency and better immunogenicity, which could be an ideal candidate rabies virus strain for inactivated vaccines preparation.