Title:
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Studies on louping-ill virus vaccine
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Studies were undertaken to define the optimum dosage regimen for commercial loupingill virus vaccine, to investigate the susceptibility of vaccinated yet seronegative sheep to challenge with live louping-ill virus and to determine the storage characteristics of the vaccine at refrigerated and room temperatures. Vaccination studies confirmed that two 1ml doses were required to provoke an adequate immune response in the majority of animals and maximum antibody titres were produced when an interval of two to eight weeks between injections was employed. Vaccinated animals were clinically resistant to experimental challenge although in animals exhibiting minimal or undetectable post-vaccinal antibody titres viraemias of low intensity were demonstrated. Circulating virus was not detected in animals with pre-challenge antibody titres greater than 1/10. Studies concerning vaccine stability demonstrated that both the virus antigen concentrate and the emulsified final vaccine were immunologically unstable following storage for periods of less than six months. Production of louping-ill virus vaccine was therefore reappraised in an attempt to improve vaccine potency and stability and to produce a one-dose vaccine capable of consistently producing a satisfactory immune response in sheep. Optimal conditions for the growth of louping-ill virus in BHK-21 cell monolayer cultures were determined experimentally and using this technique under commercial conditions titres of harvests ranged from 8.5 log₁₀pfu/ml to 6.1 log₁₀pfu/ml. A direct correlation between the preinactivation infectivity titres of virus suspensions and the immunogenicity of resulting vaccines was demonstrated and a threshold titre of 8.0 log₁₀pfu/ml determined. Harvests with preinactivation infectivity titres below the threshold produced unacceptable vaccines. Concentration of virus antigen was therefore required to consistently produce potent louping-ill virus vaccine. The present commercial vaccine is manufactured by concentrating inactivated virus harvests by methanol precipitation. A study of the efficacy of this concentration technique demonstrated however that there was no increase in the immunogenicity of concentrated virus harvests compared to unconcentrated virus harvests and in several instances concentration resulted in vaccines of decreased potency. The low potency characteristic of the commercial vaccine may therefore be partly attributable to the ineffectiveness of this technique for concentrating viral antigen. Concentration by methods other than methanol precipitation were examined; these were ultrafiltration and precipitation with polyethylene glycol. Vaccines produced from virus antigen concentrated by ultrafiltration and prepared under both experimental and commercial conditions of production were consistently more immunogenic than corresponding vaccines prepared by concentrating viral antigen by methanol precipitation or polyethylene glycol precipitation. A single 1ml dose and two 1ml doses of vaccine prepared by ultrafiltration produced a satisfactory immune response in sheep and cattle respectively and antibody titres remained constant for at least six months. Stability studies with vaccine prepared by ultrafiltration and stored at refrigerated temperatures demonstrated that although the immunogenicity of the emulsified final vaccine progressively declined over a six month period, immunogenicity of the virus antigen concentrate remained stable. Further work therefore is required to improve stability of the final vaccine and it is suggested that this be designed to evaluate the use of alternate adjuvants, preservative agents and storage conditions.
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