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Construction and identification of influenza plasmid pool imparting high yields to candidate vaccine viruses in Vero cell at low temperature.

ABSTRACT: 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.

SUBMITTER: Liu Z

PROVIDER: S-EPMC7576294 | biostudies-literature | 2020 Oct

REPOSITORIES: biostudies-literature

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Construction and identification of influenza plasmid pool imparting high yields to candidate vaccine viruses in Vero cell at low temperature.

Liu Ze Z Geng Xingliang X Cui Zhaohai Z Li Weidong W Ou Xia X Liao Guoyang G

Journal of cellular and molecular medicine 20200909 19

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 eac ...[more]

PMID: 32902192

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Project description:The development of influenza candidate vaccine viruses (CVVs) for pre-pandemic vaccine production represents a critical step in pandemic preparedness. The multiple subtypes and clades of avian or swine origin influenza viruses circulating world-wide at any one time necessitates the continuous generation of CVVs to provide an advanced starting point should a novel zoonotic virus cross the species barrier and cause a pandemic. Furthermore, the evolution and diversity of novel influenza viruses that cause zoonotic infections requires ongoing monitoring and surveillance, and, when a lack of antigenic match between circulating viruses and available CVVs is identified, the production of new CVVs. Pandemic guidelines developed by the WHO Global Influenza Program govern the design and preparation of reverse genetics-derived CVVs, which must undergo numerous safety and quality tests prior to human use. Confirmation of reassortant CVV attenuation of virulence in ferrets relative to wild-type virus represents one of these critical steps, yet there is a paucity of information available regarding the relative degree of attenuation achieved by WHO-recommended CVVs developed against novel viruses with pandemic potential. To better understand the degree of CVV attenuation in the ferret model, we examined the relative virulence of six A/Puerto Rico/8/1934-based CVVs encompassing five different influenza A subtypes (H2N3, H5N1, H5N2, H5N8, and H7N9) compared with the respective wild-type virus in ferrets. Despite varied virulence of wild-type viruses in the ferret, all CVVs examined showed reductions in morbidity and viral shedding in upper respiratory tract tissues. Furthermore, unlike the wild-type counterparts, none of the CVVs spread to extrapulmonary tissues during the acute phase of infection. While the magnitude of virus attenuation varied between virus subtypes, collectively we show the reliable and reproducible attenuation of CVVs that have the A/Puerto Rico/9/1934 backbone in a mammalian model.

Dataset Information

Construction and identification of influenza plasmid pool imparting high yields to candidate vaccine viruses in Vero cell at low temperature.

Publications

Construction and identification of influenza plasmid pool imparting high yields to candidate vaccine viruses in Vero cell at low temperature.

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