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Optimization of DNA-based Vaccination in Cows Using Green Fluorescent Protein and Protein A as a Prelude to Immunization Against Staphylococcal Mastitis

Department of Animal Science, University of Vermont, Burlington 05405

Corresponding author:
D. E. Kerr; e-mail:
dkerr{at}zoo.uvm.edu.

Staphylococcus aureus is a contagious pathogen that often results in chronic intramammary infections in dairy cows. Current vaccine formulations are ineffective in preventing this infection. The objective of this study was to stimulate an immune response in dairy cows through injection of plasmid DNA designed to express staphylococcal Protein A in transfected cells. Intramuscular and intradermal vaccination sites were evaluated using a plasmid containing the human cytomegalovirus (CMV) promoter/enhancer directing expression of green fluorescent protein (pcDNA3/GFP). DNA was delivered by needle and syringe, or by high–, intermediate-, or low-pressure jet injections (Ped-o-Jet and LectraJet). Five cows per treatment were injected with 0.5 mg of plasmid DNA at 6, 4, and 2 wk prepartum. Serum antibody levels determined by ELISA indicated that intradermal high-pressure jet injection elicited a greater immune response compared to needle and syringe injection. Differences in antibody production among low-pressure and needle and syringe treatment groups were not significant. An expression plasmid containing the CMV promoter/enhancer driving expression of the Fc-binding domain of S. aureus Protein A was coinjected into cows by vulvamucosal vaccination using the high-pressure Ped-o-Jet. Beginning 6 wk prepartum, groups of cows (n = 5) were injected three times at 2-wk intervals with DNA in saline, DNA in aluminum phosphate adjuvant, or served as noninjected controls. A cellular immune response to Protein A was detected in 4 of 10 animals, while cellular responses to GFP were not detected. Humoral responses to Protein A were observed in 6 of 10 animals and to GFP in 2 of 10 animals. Aluminum phosphate adjuvant appeared to enhance antibody production in response to Protein A. In experiment 3, a protein boost injection of Protein A was given to six animals approximately 5 mo postpartum. Three animals were nonvaccinated controls, and three were among those stimulated to produce antibody in response to the DNA-based vaccine. These results showed that Protein A specific antibodies remained elevated as compared to nonvaccinated controls and were stimulated in response to the protein boost. However, the magnitude of the response in animals previously vaccinated with DNA was not different than that observed in the nonvaccinated controls. We have shown that a humoral and cellular immune response to abbreviated Protein A can be raised in dairy cows using intravulvamucosal jet injection of a DNA-based vaccine.

Key Words: Protein A • in vivo transfection • vulva mucosa • jet-injection

Abbreviation key: CMV = human cytomegalovirus, GFP = green fluorescent protein, HRP = horseradish peroxidase, MHC = major histocompatability complex, PBS-T = PBS containing 0.1% Tween 20, PBS-T-G = PBS containing 0.1% Tween 20 and 0.1% gelatin

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