ABSTRACT
BACKGROUND: 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. METHODS: We constructed the expression plasmid (pVAC-1D) containing 1D gene FMD virus serotype Asia 1. Poly(D,L-lactide-co-glycolide) (PLG) microparticles were prepared and coated with the pVAC-1D plasmid. Guinea pigs were vaccinated with PLG-coated and naked DNA vaccine constructs intramuscularly. The humoral response was measured by an enzyme-linked immunosorbent assay (ELISA) and the serum neutralization test (SNT). Analysis of the persistence and the expression of pVAC-1D plasmid construct was carried out by quantitative polymerase chain reaction (qPCR). RESULTS: The humoral response lasted for 1 year, as measured by ELISA and SNT. Analysis of the persistence and the expression of pVAC-1D plasmid construct by qPCR has shown that pVAC-1D expression was seen for a longer duration compared to the naked DNA vaccine. Microparticles coated plasmid DNA-injected guinea pigs were protected when challenged with FMD virus. CONCLUSIONS: The present study has shown that the delivery of plasmid coated on cationic PLG microparticles enhance the duration of immunity of the DNA vaccine constructs.
Subject(s)
Capsid Proteins/immunology , Foot-and-Mouth Disease , Genetic Vectors/administration & dosage , Immunity, Active/genetics , Vaccines, DNA , Animals , Capsid Proteins/genetics , Cell-Derived Microparticles/chemistry , Foot-and-Mouth Disease/genetics , Foot-and-Mouth Disease/therapy , Foot-and-Mouth Disease Virus/genetics , Foot-and-Mouth Disease Virus/immunology , Genetic Vectors/chemistry , Guinea Pigs , Injections, Intramuscular , Lactic Acid/chemistry , Lactic Acid/immunology , Polyglycolic Acid/chemistry , Polylactic Acid-Polyglycolic Acid Copolymer , Vaccines, DNA/administration & dosage , Vaccines, DNA/immunologyABSTRACT
Calcium phosphate nanoparticles provide safe and easily manufactured vaccine adjuvant and delivery system for DNA vaccines. In the present study FMDV "O" P1-3CD DNA vaccine was encapsulated in calcium phosphate nanoparticles of size 50-100 nm diameters. The maximum loading and entrapment efficiency of nanoparticles were studied by spectrophotometer, as well as agarose gel electrophoresis. In vitro transfection efficiency of these calcium phosphate nanoparticles was found to be as good as commercial transfecting reagent lipofectamine. In vivo analysis of the calcium phosphate nanoparticle P1-3CD (CaPNP1-3CD) FMDV "O" vaccine in mice and guinea pigs could induce significant cell mediated and humoral immune response. Also, immunized mice and guinea pigs were protected against the challenge virus.
Subject(s)
Foot-and-Mouth Disease Virus/genetics , Foot-and-Mouth Disease/prevention & control , Nanoparticles/therapeutic use , Vaccines, DNA/therapeutic use , Animals , Calcium Phosphates , DNA, Viral/genetics , Female , Foot-and-Mouth Disease/virology , Genes, Viral/genetics , Guinea Pigs , Immunity, Cellular , Immunity, Humoral , Mice , Reverse Transcriptase Polymerase Chain Reaction , Vaccines, DNA/administration & dosageABSTRACT
Both polyacrylamide gel electrophoresis (PAGE) and iso-electric focusing (IEF) have been standardized using the sucrose density gradient purified 146S particles of FMD virus subtypes A10 and A22. Differences in the molecular weights of structural proteins (VP1, VP2 and VP3 of two subtypes (A10 and A22) of FMDV have been revealed in PAGE but no appreciable differences in the pI of VP1, VP2 and VP3 is found in IEF.
Subject(s)
Aphthovirus/chemistry , Viral Structural Proteins/analysis , Viral Structural Proteins/isolation & purification , Centrifugation, Density Gradient , Electrophoresis, Polyacrylamide Gel , Isoelectric Focusing , Molecular WeightABSTRACT
Hexokinase (B.C. 2.7.1.1) activity as a marker enzyme during FMD viral infection has been observed spectrophotometrically in a system coupled with glucose-6-phosphate dehydrogenase, in supernatants of BHK(21)Cl(13) suspension as well as anchored cell culture at a minimum of 10(4) infective virus particles/ml. Specific activity increased with virus concentration in culture supernatants and abruptly decreased with a fall in virus titer, as has been noted by TCID/50,146 S concentration, and enzyme-linked immunosorbent assay (ELISA) readings.
