H. J. Dechamma

DNA prime-protein boost strategy with replicase-based DNA vaccine against foot-and-mouth disease in bovine calves.

The limited efficacy of DNA vaccines against foot-and-mouth disease (FMD) in cattle and other natural hosts has prompted a search for a more effective vaccination regimen. In this study we tested a DNA prime-protein boost vaccination strategy against FMD in bovine calves. We used purified recombinant FMDV specific multi-epitope protein (rMEG990) and an optimized sindbis virus replicase-based DNA vaccine expressing this protein (pSinCMV-Vac-MEG990). We demonstrate that vaccination with a low dose of pSinCMV-Vac-MEG990 (10 μg/animal) and subsequently boosting with rMEG990 resulted in induction of neutralizing antibodies, IFN-γ production and protection against homologous virus challenge. However, vaccination with a high dose of pSinCMV-Vac-MEG990 (100 μg/animal) and boosting with rMEG990 resulted in significantly lower immune responses and more severity to the challenge test. Additionally, we show that the post-vaccinal IFN-γ levels in animals correlated positively to their protection against FMDV challenge. These findings suggest that a replicase-based DNA vaccine in proper prime-boost combination may offer an efficient vaccine strategy against FMDV and that IFN-γ could be used as an additional immune parameter to predict protection against FMDV infection.

Cationic microparticle [poly(d,l‐lactide‐co‐glycolide)]‐coated DNA vaccination induces a long‐term immune response against foot and mouth disease in guinea pigs

Foot and mouth disease (FMD) can be controlled by regular vaccination and restriction of the movement of infected animals in the endemic countries. Although presently used, tissue culture inactivated vaccine gives protection, it has several limitations, including a short duration of immunity. DNA vaccine delivered through microparticles could comprise an alternative approach to conventional vaccine when aiming to circumvent these limitations.

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

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′ and 3′ 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′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.

Improved immune response by ID-pVAC: A secretory DNA vaccine construct delivered by PLG micro particles against foot and mouth disease in guinea pigs

Foot and mouth disease (FMD) outbreaks usually have devastating effects on the economy of countries were disease is endemic due to direct and indirect cost; most of them related to international trade embargoes of animals and animal products. Although currently used inactivated vaccine provides protection, it has several drawbacks like short duration of immunity, and the requirement for containment facilities. A DNA vaccine construct which expresses the secretary antigens, delivered through micro particles could be one of the alternate approaches to overcome these limitations. Present study is envisaged to prepare a DNA vaccine construct containing the VP1 sequence of FMDV serotype O in pVAC vector. DNA vaccine was formulated by adsorbing plasmid DNA construct on cationic micro particles and administered in guinea pigs @25 μg DNA vaccine construct per animal intramuscularly. Sera samples collected were analyzed by sandwich ELISA and SNT, shown enhanced immune response in PLG adjuvanted DNA vaccine. MTT and 3H Thymidine incorporation have shown good CMI responses to PLG adjuvanted DNA. When challenged with 100 gpid50 of homologous virus 5 of the six animals were protected.

DNA vaccine (P1-2A-3C-pCDNA) co-administered with Bovine IL-18 gives protective immune response against Foot and Mouth Disease in cattle

Foot-and-mouth disease (FMD) is a highly contagious disease of cloven-hoofed animals causing considerable economic loss in the affected countries. Presently used tissue culture inactivated vaccine protects the vaccinated animals for a short duration. DNA vaccines along with appropriate adjutants is one of the approach for the development of alternative vaccine. In the present study, we constructed P1-2A-3CpCDNA (containing P1-2A-3C coding sequences of FMDV Asia-1 Ind 63/72) and bovine IL-18 pCDNA plasmids and evaluated in cattle. Four groups of calves each group containing six calves were vaccinated with 200μg of plasmid DNA vaccine P1-2A-3CpCDNA, P1-2A-3CpCDNA+ bIL-18pCDNA and inactivated vaccine respectively where as fourth group was unvaccinated. P1-2A-3CpCDNA+bIL-18pCDNA vaccinated animals have shown higher levels of neutralizing antibodies and specific T-cell proliferation responses. Higher levels of CD4(+) and CD8(+) cells were observed in these animals. Similarly, IL-18 adjuvanted group has shown increased Th1 and Th2 cytokine responses. All the vaccinated animals were challenged with cattle adapted FMD homologous Asia1 virus two weeks after the booster dose. IL18 co administered DNA vaccine construct has protected four out of six animals challenged with homologous virus.

Adjuvantation of inactivated Foot and Mouth Disease Virus vaccine with IL-15 expressing plasmid improves the immune response in Guinea Pigs

Foot and mouth disease is a highly contagious disease affecting cloven footed animals. Vaccination using inactivated virus is followed to control the disease. As the immune response conferred by the inactivated vaccine is short lived, there is a need for an alternate vaccine with increased duration of immunity. Inclusion of adjuvant which enhances B and T cell responses is one of the strategies to increase the duration of immune responses of the vaccine. Interleukin 15 is one such a cytokine which improves the cell mediated immune response and also involved in the maintenance of memory T and B cells. In the present communication, we evaluated the role of bovine IL-15 as an adjuvant to inactivated FMD vaccine in guinea pig model. Animals injected with FMD inactivated vaccine and IL-15 plasmid showed improved levels of neutralizing antibodies which were maintained up to 6 months (as the level of neutralizing antibodies is more >1.5 which is considered to give protection). Increased Th1 and Th2 responses (by measuring the level of IL-4 and IFN- gamma responses) were seen in IL-15 adjuvanted guinea pigs compared to animals injected with inactivated vaccine alone.

Expression of bovine (Bos indicus) interleukin-18 in Escherichia coli and its biological activity

IL‐18 modulates immune functions by inducing IFN‐γ production and promoting Th1 immune responses. In the present study, we amplified and cloned the sequence (582 bp) encoding full‐length bovine IL‐18 from PBMC stimulated with PHA. The nucleotide and the deduced amino acid sequence of Bos indicus IL‐18 showed an identity of 86–98% compared with IL‐18 sequences of other ruminants. The insert was subcloned into a pET 32a vector and expressed in Escherichia coli as a fusion protein and the matured protein was obtained by caspase I treatment. The specificity of these proteins was confirmed by western blotting. The biological activity of the purified protein was analyzed by its ability to induce IFN‐γ production in PBMC measured by ELISA and qPCR.