Evaluation of a genetically modified foot-and-mouth disease virus vaccine candidate generated by reverse genetics

Foot-and-mouth disease virus (FMDV) is known to employ the conserved Arg–Gly–Asp (RGD) tripeptide located on the variable βG–βH loop of the VP1 capsid protein for binding to cells. Coxsackievirus A9 (CAV9) also carries an RGD sequence, but on a short C-terminal extension of its VP1 and in a different amino acid context. This apparent relationship raised the question of whether insertion of the heterologous CAV9 sequence into FMDV would influence infection by the genetically modified FMDV. Four VP1 mutants were generated by PCR mutagenesis of a full-length FMDV cDNA plasmid. After transfection of BHK-21 cells, viral protein synthesis and virus particle formation could be detected. Two of the four mutants, mV9b and mV9d, could be propagated in BHK-21 cells, but not in CV-1 cells. Both of these mutants contained 17 amino acids of the C terminus of CAV9 VP1. Infection of BHK cells could be specifically inhibited by rabbit immune serum raised against a synthetic peptide representing the amino acid sequence of the C-terminal extension of CAV9 VP1. This demonstrated the direct involvement of the inserted sequence in cell infection. In fact, genetically modified FMDV O1K was capable of employing the VP1 C-terminal RGD region of CAV9 for infection of BHK cells. In addition, these results show that, even in cell culture-adapted viruses, the RGD-containing βG–βH loop plays an important role in virus infectivity.

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Enhanced Stability of Inactivated Foot-And-Mouth Disease Virus Vaccine With A Novel Solution Buffer.

10.21203/rs.3.rs-1035449/v1 ◽

2021 ◽

Author(s):

Jing Li ◽

Yanming Wei ◽

Rong Zhang ◽

Huiqing Yang ◽

Xuerong Liu

Keyword(s):

Disease Virus ◽

Virus Vaccine ◽

Vaccine Development ◽

Foot And Mouth Disease ◽

Mouth Disease ◽

Size Exclusion ◽

Basic Solution ◽

The Novel ◽

Mouth Disease Virus ◽

Foot And Mouth

Abstract Objectives: Stability is vital for potency of food-and-mouth disease virus vaccine preparation. However, the assembly of inactivated foot-and-mouth disease virus is poor stable and prone to dissociate into 12s under mild acidic or heating conditions, especially emulsified with oil-adjuvant. Thus, it is crucial to explore a suitable medium and condition to improve the stability and efficiency of inactivated FMDV vaccine. Results: In this study, the basic solution buffer and a serious of potential stabilizers, such as carbohydrate, amino acid, antioxidant, salt and antioxidant were screened for evaluating stable effect on FMDV antigen with aid of high performance size exclusion chromatography (HPSEC). On this basis, orthogonal experiment was performed to optimize and finally confirm the formulation. Anti-aging test were carried out to asses the efficiency of formulation on vaccine stability and the results showed that the vaccine was more stabler either stored at 37℃or 4℃. Moreover, physicochemical monitoring revealed that formulation had no influence on the properties of vaccine. The combined results suggested that the novel solution buffer would lower degradation and prolong shelf life of vaccine. In a word, the novel buffer is beneficial to make FMD vaccine more stable and effective, reducing the dependence on cold delivery and storage. This study also provides insight into the processes of optimization and inactivated vaccine development.

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