Self Replicating Gene Vaccine Carrying P1-2A Gene of FMDV Serotype O and its Effects on the Immune Responses of Cattle

Abstract

DNA vaccines are considered as alternatives to live attenuated ones for those diseases like foot-and-mouth disease (FMD) where the production and application of live vaccines have been found unsuccessful. However, stability of DNA and the quantity of antigen expressed are the major limitation with naked DNA vaccines. To address these issues self replicating gene vaccine construct was made for foot-and-mouth disease virus (FMDV) type 'O' and studied. The vector for vaccine construct, designated as pSinCMVVac carried CMV promoter and Poly(A) signal sequences at 5 0 and 3 0 end of Sindbis replicase gene respectively. Gene for structural protein precursor (P1-2A) of FMDV serotype 'O' was inserted into pSinCMVVac under subgenomic promoter. 5 0 UTR (untranslated region) of FMDV was introduced upstream of P1-2A to enhance the level of expression of cloned gene. Functionality of the vaccine construct was confirmed in vitro and in vivo. The self-replicating gene vaccine construct was tested in cattle in comparison with naked DNA vaccine carrying P1-2A and 3CD (pUP3CD). Humoral immune response by ELISA and SNT and cellular response by lymphoproliferation assay using MTT were studied. The default approach of using self replicating gene vaccine in high dose and multiple injection in cattle as followed in our studies might result in immunosuppression as this was observed in our subsequent experiments in guinea pigs. Hence based on dose response studies, vaccine strategy needs to be decided. However, the approach of using Sindbis polymerase gene and UTR in FMDV vaccine is the first report and shows future scope of developing such vaccines.

Immunization of chicken with flagellin adjuvanted Salmonella enteritidis bacterial ghosts confers complete protection against chicken salmonellosis

“…The CMI responses against antigen stimulation were assessed by PBMC proliferation by 3-(4, 5-dimethylthiazol-2-yl)-2, 5-diphenyl tetrazolium bromide ( MTT ) assay ( Nagarajan et al, 2011 ). The PBMCs were separated by Histopaque 1077 (Sigma, St. Louis, MO) density gradient centrifugation.…”

Sindbis Virus Replicase‐based DNA Vaccine Construct Encoding FMDV‐specific Multivalent Epitope Gene: Studies on its Immune Responses in Guinea Pigs

“…Sindbis virus replicase‐based vector pSinCMV‐Vac [33] was used for constructing the replicase‐based DNA vaccine for FMD. The plasmid pET‐MEG990 [34] was used as a source of the FMDV‐specific multivalent epitope gene (MEG990).…”

“…Construction of the replicase‐based DNA vaccine encoding FMDV multiple epitopes, pSinCMV‐Vac‐MEG990. Construction of the SINV replicase‐based vector, pSinCMV‐Vac [33], and the linking of C‐terminal halves of VP1 of serotype O, A and Asia 1 by viral 2A sequences into MEG990 gene were described earlier [34]. In the present study, the FMDV‐specific MEG990 gene was inserted into the pSinCMV‐Vac for use as a DNA vaccine for FMD.…”