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Usage of influenza vaccine is the best choice measure for preventing and conclusion of influenza virus infection. Although it has been used of chicken embryo to produce influenza vaccine, following with WHO recommended vaccine strain, there were uncontrollable factors and its deficiencies, specially, during an influenza pandemic in the world. The V...
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... the intraperitoneal injection (i.j.), animal assay was used for anaphylatoxin testing. After a sensitization and excitation per- iod, the symptoms of systemic allergic reaction criteria were evaluated (Table 1) and scored for the guinea pigs. These Figure 1. ...
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... of positive reaction Incidence of extremely strong positive reaction - + ++ +++ ++++ YN/05BVca 20 18 2 0 0 0 10% 0% BSA/08Vca 20 19 1 0 0 0 5% 0% BSA/08wt 20 0 1 16 2 1 100% 5% PBS 20 20 0 0 0 0 0% 0% Figure 4. Determination of weight changes for safety evaluation. The experi- mental animals' weights increased, and no significant differences was observed between the Vca groups and PBS control group (P > 0.5). ...
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Citations
... We employed the reverse genetics technology to combine eight gene fragments from the A/Yunnan/1/2005(H3va) [1] used as hyv and att strain donor strain and A/Jilin-Chaoyang/16/2016 (H3wt) used as wide type highly virulent influenza virus strains, which finally prepared 36 reassortment virus strains by specific gene fragment order. In this study, using mice weight changes and viral loading on the lower respiratory tract to decide the attenuation characteristics; using virus titter changes serially passaged eighteen times on Vero cells to decide the Vero cells high yield characteristics; using the neuraminidase inhibition (NI) and hemagglutination inhibition (HI) titter in the nasal immunized mice serum to decide the immunogenicity characteristics. ...
... On the other side, the advantage of Vero cells derived vaccine had been discussed by our previous study [1,9] . However, PR8 could not grow efficiently in Vero cells. ...
... These five gene constellation could generate new reassortant virus with att, hyv and imm characteristics. In the meantime, the immune effect of them were also determined by HI and NI testing, which was consistent with our previous research [1,9,17,18] . ...
It described the “ladder distribution” of live attenuated influenza vaccine (LAIV) reassortment between Vero cells adaption attenuated strain and wide type-high virulent influenza virus strains by the reverse genetics approach. That gene constellation could save the immunogenicity (imm) of the parental strain and make new reassort virus strain attenuation (att) characteristics for further vaccine using, rapidly, not only seasonal vaccine but also pandemic flu vaccine.
... This finding might be a of great practical benefit to modify reassortment method with vero cell adaption (va) parental strain and wide type (wt) virus for improved growth of vaccine 'seed' viruses, which could help influenza disease prevention and intervention initiatives. We employed the reverse genetics technology to combine eight gene fragments from the A/Yunnan/1/2005(H3va) [1] used as hyv and att strain donor strain and A/Jilin-Chaoyang/16/2016 (H3wt) used as wide type highly virulent influenza virus strains, which finally prepared 36 reassortment virus strains by specific gene fragment order. In this study, using mice weight changes and viral loading on the lower respiratory tract to decide the attenuation characteristics; using virus titter changes serially passaged eighteen times on Vero cells to decide the Vero cells high yield characteristics; using the Neuraminidase Inhibition (NI) and Hemagglutination Inhibition (HI) titter in the nasal immunized mice serum to decide the immunogenicity characteristics. ...
... Now, the quadrivalent inactivated influenza virus vaccine produced using the Madin Darby canine kidney cell line has been approved in the EU (Flucelvax® Tetra) and USA (Flucelvax Quadrivalent®; QIVc hereafter) for the prevention of influenza in adults and children [15]. On the other side, the advantage of Vero cells derived vaccine had been discussed by our previous study [1,9]. However, PR8 could not grow efficiently in Vero cells. ...
... These five gene constellation could generate new reassortant virus with att, hyv and imm characteristics. In the meantime, the immune effect of them were also determined by HI and NI testing, which was consistent with our previous research [1,9,17,18]. ...
It described the ladder distribution of Live Attenuated Influenza Vaccine (LAIV) reassortment between Vero cells adaption attenuated strain and wide type high virulent influenza virus strains by the reverse genetics approach. That gene constellation
... To improve growth efficiency of influenza viruses in Vero cells, some techniques have been reported, including using internal genes from adapted viruses [6,[34][35][36][37] or introducing Vero cell-adapted mutations [38,39]. In 2015, Ping et al. established random mutation libraries of six internal genes to select MDVs for both egg-based and cell-based platforms from over thousand candidate viruses [8]. ...
The embryonated egg-based platform currently produces the majority of seasonal influenza vaccines by employing a well-developed master donor virus (MDV, A/PR/8/34 (PR8)) to generate high-growth reassortants (HGRs) for A/H1N1 and A/H3N2 subtypes. Although the egg-based platform can supply enough seasonal influenza vaccines, it cannot meet surging demands during influenza pandemics. Therefore, multi-purpose platforms are desirable for pandemic preparedness. The Vero cell-based production platform is widely used for human vaccines and could be a potential multi-purpose platform for pandemic influenza vaccines. However, many wild-type and egg-derived influenza viruses cannot grow efficiently in Vero cells. Therefore, it is critical to develop Vero cell-derived high-growth MDVs for pandemic preparedness. In this study, we evaluated two in-house MDVs (Vero-15 and VB5) and two external MDVs (PR8 and PR8-HY) to generate Vero cell-derived HGRs for five avian influenza viruses (AIVs) with pandemic potentials (H5N1 clade 2.3.4, H5N1 clade 2.3.2.1, American-lineage H5N2, H7N9 first wave and H7N9 fifth wave). Overall, no single MDV could generate HGRs for all five AIVs, but this goal could be achieved by employing two in-house MDVs (vB5 and Vero-15). In immunization studies, mice received two doses of Vero cell-derived inactivated H5N1 and H7N9 whole virus antigens adjuvanted with alum and developed robust antibody responses.
... The Madin-Darby canine kidney (MDCK) cell line is sensitive to the influenza A and B viruses [8], but the potential tumorigenicity of this cell line is believed to be problematic by some scientists and vaccine scholars, including within our department. The second option is the African green monkey (Vero) cell line, which has been used to generate the polio and rabies vaccines [9]. However, there are no Vero cell-derived seasonal attenuated influenza A or B vaccines at present. ...
... Thus, BSL-3 was utilized in this virus generation procedure, which is safer than BSL-2. The reassortant virus was developed using the classical method, which has been described previously [9,16]. In this study, the multiplicity of infection (MOI) of each virus strain at each stage in the Vero cells was determined as needed. ...
... An analysis of animal weight changes (n.j.), showed that they grew slowly and steadily, which may provide an additional level of safety for a live influenza B vaccine derived from the YN/05BVca backbone [9,16]. ...
Background: It was to generate a new Vero and cold-adapted live attenuated influenza B vaccine with enough safety and immunogenicity.
Methods: According to modified classical reassortment method, the donor strain was B/Yunnan/2/2005Vca(B), and the parental virus strain was B/Brisbane/60/2008wt. After co-infection in Vero cells, the prepared antibody serum inhibited the donor strain growth, and screening conditions inhibited the parental virus growth, which induced the growth of the new reassortant virus B/Brisbane/60/2008Vca(B) grow. Through intraperitoneal injection (i.j.) and intranasal injection (n.j.) we evaluated the safety and immunogenicity of the vaccine.
Results: A high-yield of the reassortant virus was produced in Vero cells at 25°C, similar to the donor strains. After sequencing, it was found that B/Brisbane/60/2008Vca(B) Hemagglutinin (HA) and Neuraminidase (NA) gene fragments were from B/Brisbane/60/2008wt, while the other 6 gene fragments were from B/Yunnan/2/2005Vca(B). The n.j. immune pathway experiments showed no significant differences between the treatment and the PBS control group with respect to weight changes (P > 0.5). Furthermore, the new strain had a sufficient geometric mean titter (GMT) against B/Brisbane/60/2008wt.
Conclusion: The new reassortant live attenuated influenza B vaccine was safe and having enough immune stimulating ability.
We generated plasmid pools for the rapid preparation of candidate vaccine strains, which could grow in the Vero cells at low temperature. Firstly, we cloned in the pHW2000 plasmid each of the eight gene segments (PB2, PB1, PA, hemagglutinin [HA], neuraminidase [NA], NS, NP, M) of two master donor strains (MDS), respectively, A/Yunnan/1/2005Vca(H3N2) and B/Yunnan/2/2005Vca(By), which had Vca phenotype (cold-adapted phenotype in Vero cells). Secondly, the similar operation was implemented with each of the HA, NA and NP segments of circulating strains with epidemic potential (parental strains). The virus rescue techniques were employed in this study, according to the homology rate of HA segments between MDS and parental strains. Then, we harvested amount of new Vca virus strains. By transmission electron microscope, it could observe new viruses' diameter and length were from 100 to 120 nm. Importantly, these reassortant viruses could get high-yield production in Vero cells at 25℃ from the beginning to the fourth generation, which was significantly differ from their original parental viruses. Additional, these production 16 new Vca strains could maintain enough antibody binding capacity and attenuation phenotype, which consisted with their MDS. So these plasmid pools constructed by mount of different influenza A and B virus gene fragments could present desired working performance and provide convenience and realization for more Vca reassortant virus as candidate vaccine strain if needing.
